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Modulating Ageing Anti-Ageing
Heart CentreUniversity Hospital Halle
Ernst-Grube-Str. 4006120 Halle (Saale)
From Friday,
September 1st, 2017, 6 pm
till Sunday,
September 3rd, 2017, 1:30 pm
Opening: Lion BuildingUniversitätsplatz 11, Halle (Saale)
Conference Site: Steintor-CampusAdam-Kuckhoff-Straße 35, 06120 Halle (Saale)
Keynote SpeakerClaudio FranceschiBologna, Italy
from Molecular Biology to Clinical Perspectives
German National Academy of Sciences Leopoldina
German Society of Gerontology and Geriatrics (DGGG)
Interdisciplinary Centreon Ageing Halle (IZAH)
RTG 2155: ProMoAge
Modulating Ageing / Anti-Ageing:
from Molecular Biology to Clinical Perspectives
September 01st
– 03rd
2017
Heart Centre University Hospital Halle (Saale)
in cooperation with
German National Academy of Sciences Leopoldina
German Society of Gerontology and Geriatrics
(DGGG)
Interdisciplinary Centre on Ageing Halle (IZAH)
RTG 2155: ProMoAge
Index
Page
4 Running Program
5 Abstracts Speakers
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Aging and diseases: common mechanisms but different rates Claudio Franceschi
Defining the target group: risk prediction of ageing trajectories Andrea Maier
Do biomarkers of ageing reflect real age? Andreas Simm
Challenges understanding aging through biomarkers across populations: the example of calcium Tamàs Fülöp
Motor and cognitive intervention to improve functional mobility and reduce falls in older adults and patients with neurodegeneration Anat Mirelman
Age-dependent posttranslational protein modification – driving force in neurodegeneration? Hans-Ulrich Demuth
Protein oxidation and aggregation in aging and senescence Tilman Grune
Proteasome modulation as an anti-aging and antiaggregation strategy Niki Chondrogianni
Glycated proteins in nutrition: friend or foe Katarina Sebekova
Cardiac ageing enhances atrial fibrillation or vice versa? – Impact of altered miRNA expressional profile and metabolic signalling for treatment and persistence of atrial fibrillation? Bernd Niemann
Protective effects of caffeine in the cardiovascular system depend on mitochondrial p27 – in aging and disease Judith Haendeler
Oral anticoagulation and polypharmacy in older adults with atrial fibrillation Markus Gosch
Gender related aspects of the cardiovascular system and its impact on progression and aging Sandra Eifert
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Historical overview of research on lifespan extension and its ethical implications Georg M. Martin
Rejuvenation biotechnology: why age may soon no longer mean aging Aubrey de Grey
The shape of lives to come Ian Richard Ground
Ethical aspects of biogerontological research Zoé Rheinsberg
Epigenetic stress response and stem cell aging Lenhard Rudolph
Understanding muscle stem cell regenerative decline with aging Pura Munoz-Cánove
Bioengineered matrices to regenerate the heart – The Zurich LifeMatrix programm Maximilian Emmert
Mesenchymal stem cells-on-chips/organ-on-chips/chips in mesenchymal organs – monitoring healing kinetics: an ageing perspective Günter Lepperdinger
Effects of pharmacological inhibition of aging-associated pathways on Drosophila lifespan Alexey Moskalev
The Path towards translating senolytic drugs and other interventions targeting aging processes into clinical treatments James Kirkland
Cellular senescence and aging Judith Campisi
DNA damage: impact on aging, neurodegeneration and the effect of nutritional interventions Jan Hoeijmakers
58 Posters ( in alphabetical order)
114 List of speakers
118
119
Organizers
Involved Societies
120 Authors Index
125 List of Sponsors
126 Map of Halle (Saale)
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R u n n i n g P r o g r a m
Modulating Ageing / Antiageing: From Molecular Biology to Clinical Perspectives
- Meeting language English -
Friday September 01st 2017 18:00 Opening
Andreas Simm
Address M. Gekle, Dean of the Medical Faculty,
Martin-Luther-University Halle-Wittenberg T. Klöss, Medical Director of the University Hospital Halle (Saale) H. Treede, Heart Centre of the University Hospital Keynote lecture and Schober award
Laudation Claudio Franceschi by Judith Campisi, Novato, USA
Keynote lecture: Claudio Franceschi University of Bologna, Bologna, I
“Aging and diseases: common mechanisms but different rates”
20:00 Come Together (Löwengebäude – Lion Building)
5
Saturday September 02nd 2017
08:00 – 10:00 Session 1
Setting up the problem, predictors and markers Chair: Claudio Franceschi, Andreas Simm Defining the target group: risk prediction of ageing Andrea Maier trajectories Do biomarkers of ageing reflect real age? Andreas Simm Challenges understanding aging through biomarkers Tamás Fülöp across populations: the example of calcium Motor-Cognitive Interventions to reduce the risk of falls Anat Mirelman in older adults and neurodegenerative conditions
10:00 – 10:30 Coffee Break
10:30 – 12:30 Session 2
Protein damage and Aggregation Chair: Günter Lepperdinger, Lenhard Rudolph
Age-dependent posttranslational protein modification – Hans-Ulrich Demuth
driving force in neurodegeneration?
Protein Oxidation and Aggregation in Aging and Tilman Grune Senescence Proteasome modulation as an anti-aging and anti- Niki Chondrogianni aggregation strategy Glycated proteins in nutrition: friend or foe Katarina Sebekova
12:30 – 13:30 Lunch Break
6
Saturday September 02nd 2017
13:30 – 15:30 Session 3
Cardiovascular System
Chair: Tamàs Fülöp, Maximilian Emmert Cardiac ageing enhances atrial fibrillation or vice versa? – Bernd Niemann Impact of altered miRNA expressional profile and metabolic signalling for treatment and persistence of atrial fibrillation? Protective effects of caffeine in the cardiovascular system Judith Haendeler depend on mitochondrial p27 - in aging and disease Oral anticoagulation and polypharmacy in older adults Markus Gosch with atrial fibrillation Gender related aspects of the cardiovascular system and Sandra Eifert its impact on progression and aging
15:30 – 16:30 Poster Session / Coffee Break
16:30 – 18:30 Session 4
Panel discussion on ethical issues Chair: Jutta Schnitzer-Ungefug Historical overview of research on lifespan extension George M. Martin and its ethical implications Rejuvenation biotechnology: Aubrey de Grey why age may soon no longer mean aging The shape of lives to come Ian Richard Ground Ethical aspects of biogerontological research Zoé Rheinsberg
20:00 Conference-Dinner (Halloren- und Salinemuseum)
7
Sunday September 03rd 2017
08:30 – 10:30 Session 5
Stem Cells and Regeneration Chair: Judith Campisi, Judith Haendeler Epigenetic stress response and stem cell aging Lenhard Rudolph Understanding muscle stem cell regenerative decline Pura Munoz-Cánove with aging Bioengineered matrices to regenerate the heart – Maximilian Emmert The Zurich LifeMatrix programm Mesenchymal stem cells-on-chips/organ-on-chips/chips Günter Lepperdinger in mesenchymal organs – Monitoring healing kinetics: an ageing perspective
10:30 – 11:30 Poster Session / Coffee Break
11:30 – 11:45 Posterprice
11:45 – 13:45 Session 6
Anti-Aging Therapies Chair: Andrea Maier, George M. Martin Effects of pharmacological inhibition of aging-associated Alexey Moskalev pathways on Drosophila lifespan The path towards translating senolytic drugs and other James Kirkland interventions targeting aging processes into clinical treatments Cellular senescence and aging Judith Campisi DNA damage: impact on aging, neurodegeneration and Jan Hoeijmakers the effect of nutritional interventions
13:45 – 14:15 Farewell
8
Aging and diseases: common mechanisms but different rates
Claudio Franceschi
The new GEROSCIENCE is based on the strong epidemiological evidence that aging is the
major risk factor for age-related diseases, and assumes that they share a common set of basic
mechanisms. This assumption is supported by a large panoply of experimental data and its
major consequence is that the primary target of Medicine is to combat aging instead of any
single age-related disease one by one, as favored by the present fragmentation into hundreds
of specialties and sub-specialties. If the same molecular and cellular mechanisms underpin both
aging and age-related diseases, a major question emerges: which is the difference between
aging and diseases? This is an old question that was addressed starting from Terentius and
Cicero who argued pros and cons, respectively, whether “senectus ipsa est morbus”, and
recently this topic was addressed by several scholars. On the basis of a variety of recent
experimental data and of the conceptualizations I proposed in the last years (the “remodeling”
theory of aging; inflammaging and garbaging; liquid immune self and immunobiography) I will
illustrate the hypothesis that age-related diseases - including the not so well-defined geriatric
syndromes and conditions such as frailty, metabolic syndrome and mild cognitive impairment -
can be conceptualized as ACCELERATED aging. Within such a perspective centenarians
represent one extreme (healthy aging/extreme longevity) characterized by DECELERATED
aging. At the other extreme there are patients who suffered one or more severe age-related
pathologies in their sixties, seventies and eighties, and died 20-40 years before people of the
same cohort who reached 100 years or more. These patients can be considered as affected by
ACCELERATED aging. In between these two extremes there is a bunch of intermediate
trajectories representing a sort of grey area. Thus, clinically different, classically-defined age-
related diseases such as diabetes, cardiovascular diseases, cancer, dementia, among others,
are indeed the result of a peculiar combination of alterations regarding the same, above-
mentioned shared, limited set of basic mechanisms. Thus, as anticipated by Lazlo Barabasi
some years ago with the conceptualization of “diseasome”, apparently/clinically different
diseases share common mechanisms and genes. According to this integrated view the question
whether aging is a disease in itself becomes meaningless because aging and age-related
diseases are part of a continuum where precise boundaries do not exist. Whether an individual
will follow a trajectory of accelerated or decelerated aging will depend on his/her genetic
background (population genetics) which will interact lifelong with environmental and lifestyle
factors, in a rapidly changing world. Within this scenario it is easy to understand how important
and critical is the search for biomarkers capable of distinguishing between chronological and
biological age and predict/anticipate the rate/trajectory of aging.
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Defining the target group: risk prediction of ageing trajectories
Andrea Maier
Science behind ageing trajectories is rapidly emerging, showing differences in the rate
of ageing already at early ages. This indicates that organ system changes occur even
before diseases become prevalent. Measurements quantifying the ageing process are
not yet defined but highly warranted to disentangle the chronological age from the
biological age, expressing the rate of ageing, in humans.
The EU funded projects PreventIT and PANINI both aim to 1) identify individuals at risk
for future detrimental outcome and 2) tailored approaches to prevent negative
outcomes. Latest evidence on how to measure biological age and its prediction for
ageing trajectories will be presented for different age categories.
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Do biomarkers of ageing reflect real age?
Andreas Simm
Average life span and life expectancy in the industrial nations have grown dramatically
in the last century. Degenerative diseases will become of major interest with age being
an independent risk factor in most of these diseases. Clinicians know that the
chronological age of a patient does not really represent the ‘real’ age of the person.
Biomarkers of ageing should help to characterize the individual biological age and could
be subsequently used to identify individuals at high risk of developing age-associated
diseases or disabilities. A biological age scale can further help to evaluate the success
of preventive actions.
Unfortunately, there are many problems by creating / identifying such biomarkers. We
do not have a gold standard for measuring biological age but do correlate proposed
biomarkers with chronological age. Most of these markers were identified within blood,
which does not reflect all organs.
Skin-autofluorescence related to the accumulation of advanced glycation endproducts is
one of the proposed biomarkers of ageing. On the cellular levels, treatment of
endothelial with AGE inducing dicarbonyls induce cellular senescence. Accumulation of
AGEs in tissue collagen correlate with age and the outcome of patients after cardiac
surgery. Skin autofluorescence reflects tissue AGE modification, correlates with age,
predicts the outcome of patients and can be modified by preventive actions like
exercise. Whereas these data are promising, AGE skin autofluorescence like all other
proposed biological age scales can be used in cohorts only but are unable to monitor an
individual risk.
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Challenges understanding aging through biomarkers across populations: the example of calcium
Tamàs Fülüp
Recent studies have shown contradictory associations between calcium levels and
health outcomes. We suspected these conflicting results were the consequence of more
general issues with how biomarkers are analyzed in epidemiological studies, particularly
in the context of aging. To demonstrate the risks of typical analyses, we used three
longitudinal aging cohort studies and their demographic subsets to analyze how calcium
levels change with age and predict risk of mortality and frailty. We show that calcium
levels either increase or decrease with age depending on the population, and positively
or negatively predict frailty depending on the population; both age and frailty results
showed substantial heterogeneity. Mortality analyses revealed few significant
associations but were likely underpowered. Variation in population composition
(demographics, diseases, diet, etc.) leads to contradictory findings in the literature for
calcium and likely for other biomarkers. Epidemiological studies of biomarkers are
particularly sensitive to population composition both because biomarkers generally have
non-linear and often non-monotonic relationships with other key variables, notably age
and health outcomes, and because there is strong interdependence among biomarkers,
which are integrated into complex regulatory networks. Consequently, most biomarkers
have multiple physiological roles and are implicated in multiple pathologies. We argue
that epidemiological studies of aging using biomarkers must account for these factors,
and suggest methods to do this.
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Motor-Cognitive Interventions to reduce the risk of falls in older adults and neurodegenerative conditions
Anat Mirelman
Gait impairments and falls are ubiquitous among older adults and patients with common
neurological diseases. Normal and safe mobility depends on intact sensory and motor
systems, but there is a growing body of research that specifically links the cognitive sub-
domains of attention and executive function to gait alterations and fall risk. These
domains apparently play a critical role in the regulation of gait especially under
challenging conditions where decisions need to be made in real-time and constant
adaptation is required to manage internal and external factors such as occur in
everyday life situations. Current therapeutic approaches tend to provide interventions
that target a single domain, however, therapeutic interventions, which focus on a
combined motor-cognitive approach, may improve gait and functional everyday mobility
and decrease the risk of falls in older adults and patients with neurological disorders.
Dr. Mirelman will present an overview on the links between motor and cognitive function
and their relationship to mobility, everyday life activity, and falls in ageing. Evidence
from the V-TIME project, a recently completed multi-modal randomized controlled trial,
aimed at improving mobility and fall risk in older adults and patients with Parkinson’s
disease will be presented as an example for a motor-cognitive intervention. The study
compared the effects of a combined treadmill training with virtual reality with a similar
intensity motor training alone on mobility and fall frequency in a diverse group of 302
older adults with a history of falls. The study demonstrated that both interventions were
effective in reducing fall frequency over a 6-month follow up period but the effects of the
treadmill- virtual reality training was significantly larger than that of the treadmill alone.
The presentation will provide a summary of advantages and disadvantages of such an
approach, the fidelity of the method used, the suitability of such intervention for this
population, and the utility of these types of interventions in community settings.
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Age-dependent posttranslational protein modification – driving force in neurodegeneration?
Hans-Ulrich Demuth
The Alzheimer syndrome is the most frequent age-related neurodegenerative disorder which
affects today more than 35million patients world-wide. In Germany alone this number will
exceed 3million people in 2050.
Up to now, there is no medication available which would hold or even reverse the progressive
disease. In recent decades attempts to find disease-modifying drugs have failed. Many of them
have been directed against the deposited Abeta peptides (Aβ), which are product of the
turnover of a membrane protein (APP) of neurons. Some drug approaches where developed to
slow down this protease-driven turnover using small molecule inhibitors, some to immunize
against the Aβ peptides.
However, all of them did not specifically target the very different posttranslational modifications
of Aβ or the Amyloid Precursor Protein (APP). Recent research uncovered among these
modifications very aggregation prone forms which in a prion-like manner induce the formation of
soluble and metastable oligomeric aggregates, which are acknowledged today as the most
neurotoxic agents in the human brain.
Among them, pyroglutamate-3 amyloid-β (pGlu-3 Aβ) is an N-terminally truncated and
pyroglutamate-modified Aβ species which has been shown to be a major component of Aβ
deposited in the plaques and blood vessels of Alzheimer’s disease and Down syndrome brains
(Saido et al., 1995; Lemere et al., 1996).
Formation of pGlu-3 Aβ is a multi-step process whereby the first two N-terminal amino acids,
aspartate and alanine, are cleaved off, exposing glutamate at position 3 on the N terminus of
Abeta. Subsequently, glutamate is post-translationally modified to N-terminal pyroglutamate by
cyclization of the exposed glutamate residue by glutaminyl cyclase and resulting altered
biochemical properties including increased hydrophobicity, high aggregation propensity and
neurotoxicity (Russo et al., 2002; Schilling et al., 2006).
Prevention and therapeutic trials previously conducted in different laboratories showed that
prevention of pGlu-3 Aβ formation by inhibiting the enzyme Glutaminyl Cyclase and pGlu-3 Aβ
mAb immunizations resulted in a reduction in general Aβ and pGlu-3 Aβ pathology (Nussbaum
et al., 2012; Frost et al., 2012). This suggests that presence of pGlu-3 Aβ contributes to
increased plaque burden. Both approaches have recently reached clinical development stage.
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Protein Oxidation and Aggregation in Aging and Senescence
Tilman Grune
Reactive oxygen species (ROS) are formed continuously in the organism even under
physiological conditions. Proteins are prominent targets of various modifications such as
oxidation, glycation, or conjugation with products of lipid peroxidation, leading to the
alteration of their structure and biological function. Under steady state conditions such
modifications are leading to an enhanced degradation of the damaged protein, however,
if the degradation machinery is overwhelmed, protein aggregates are formed.
Such protein aggregates are often randomly formed, are highly reactive, grow with time
and have their own pathophysiological effects which are not yet well understood. These
pathophysiological functions include the formation of reactive species, inhibition of parts
of the ubiquitin-proteasomal system and the induction of lysosomal instability. A large
part of the existing protein aggregates are localized in the autophagosomes of
mammalian cells, but cannot be degraded after fusion with lysosomes, leading to
lysosomal rupture and free protein aggregates, affecting the cellular metabolism. In
order to reduce the effects of free protein aggregates, chaperones assist the formation
of aggresome-like structures which can again be taken up into autophagosomes.
Therefore, protein aggregates underlie a constant fluctuation in the senescent cell and
exert a multitude of effects in the aged cell.
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Proteasome modulation as an anti-aging and anti-aggregation strategy
Niki Chondrogianni
Proteasomes are constituents of the cellular proteolytic networks that maintain protein
homeostasis through regulated proteolysis of normal and abnormal (in any way)
proteins. Proteasome activation in cell lines has been shown to result to cellular lifespan
extension and to exert protein anti-aggregation activity.
Using Caenorhabditis elegans as a model, we analyzed in detail the proteasome status
upon the progression of ageing and Alzheimer's disease (AD) and we investigated the
effects of enhanced proteasome activities on the progression of the above mentioned
phenomena. The obtained results were validated in human and murine cells of neuronal
origin.
Proteasome activation in C. elegans either through genetic means or through
compounds resulted in enhanced levels of proteasome activities that led to a SKN-1-
and proteasome activation-dependent lifespan extension. The elevated proteasome
function conferred lower paralysis rates in various AD nematode models accompanied
by decreased Aβ deposits thus ultimately decelerating the progression of AD
phenotype. More importantly, similar positive results were also delivered in human
neuroblastoma cells and in murine cortical neurons.
Our results suggest that proteasome activation with downstream positive outcomes on
ageing and AD, an aggregation-related disease, is feasible in both a genetic and a non-
genetic manipulation manner in a multicellular organism. Moreover they unveil the need
for identification of anti-ageing and anti-amyloidogenic compounds among the nutrients
found in our normal diet.
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Glycated proteins in nutrition: friend or foe
Katarina Sebekova
During thermal processing of foods, non-enzymatic amino-carbonyl reactions lead to
formation of Maillard reaction products (MRPs) - brownish substances, rendering foods
attractive color, odor and taste. Only a few MRPs are chemically defined yet. In vivo
analogues of MRPs are termed advanced glycation end products (AGEs). AGEs are
formed mainly on long-lived proteins, during ageing, and under conditions of
hyperglycemia, enhanced oxidative and carbonyl stress. AGE-modification alters protein
structure and function. Interaction of AGEs with their receptor RAGE may result in
diabetogenic, inflammatory, or pro-atherogenic responses. Studies in rodents showed
that dietary LMW MRPs are partially absorbed into circulation. Their intake is reflected
by increase in plasma AGEs, their urinary excretion, and accumulation in different
tissues. Dietary MRPs (dMRPs) are the main source of circulating AGEs. Majority, but
not all, studies in rodents and humans showed that diets high in MRPs associate with
negative health effects (e.g. elevated inflammatory markers, impairment in insulin
sensitivity, late complications of diabetes). However, vegetarians, who present more
favorable cardiometabolic risk profile compared with omnivores, consume more dMRPs
and present higher circulating AGE levels. Thus, cause-and-effect relationships
between dMRPs intake and health effects are far from clear by now, as rodents are not
accustomed to ingestion of heated food, thus might not be the ideal model for effects of
dMRPs in humans, and human studies generally use different culinary approach to
prepare high- vs. low-MRP-diets. Formation of MRPs in foods may decrease the
bioavailability of proteins and their susceptibility to gastrointestinal degradation. Other
potentially toxic substances (e.g. acrylamide, aromatic amines) are formed during heat-
treatment of foods. On the other hand, some MRPs may exert antioxidative effects, and
formed melanoidins may exert positive health effects. Recent food habits (e.g.
substitution of breast-milk with infant formulas which contain 100-folds higher amounts
of MRPs; increasing consumption of fast-foods and snacks), and the increase in life
expectancy rates (accompanied by the appearance of novel age-related pathologies, in
which AGEs seem to be implicated) point to the need of further research in this area.
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Cardiac ageing enhances atrial fibrillation or vice versa? Impact of altered miRNA expressional profile and metabolic signalling
for treatment and persistence of atrial fibrillation?
Bernd Niemann
Ageing is said to induce atrial fibrillation (AF) and to reduce therapeutic success during
medical and ablation therapy. Moreover, senescent patients often suffer from perma-nent
AF resulting in the need of oral anticoagulation. Accordingly, elderly patients are at risk for
bleeding complications due to anticoagulation. Therefore, freedom from AF and freedom
from anticoagulation might be an important quality marker for perioperative cardiac surgery
success in aged patients despite indication for anti-arrhythmic therapy is weakened in these
patients following latest guidelines. However, data are lacking that answer the question if
AF is the result of ageing or if AF is causing age-associated atrial changes. Addressing AF
might be as well addressing pathological ageing of the atria and help to improve cardiac
function in patients undergoing cardiac surgery.
Atrial fibrillation from paroxysmal to persistent AF is accompanied by atrial structural
remodeling. Pro-fibrotic signals lead to interstitial fibrosis, which is considered a structural
basis for the development and sustaining of AF and might be understood as to be the
substrate of atrial ageing. Cardiac microRNAs (miRNAs) and adipokines are important
regulators of this interstitial fibrosis. We have characterized patterns of miRNA-signaling
and metabolic signaling (adipokines) in patients suffering from paroxysmal, persistent and
new onset atrial fibrillation.
Clinical and molecular patterns observed in patients with new-onset postoperative AF are
similar to patterns in patients with paroxysmal AF, but are more pronounced in patients with
persistent AF but absent in patients (and elderly patients) with sinus rhythm. Thus,
calendrical age per se might not offer a highly potent substrate and trigger for induction and
persistence of atrial fibrillation, which allows for therapeutic success even in elderly. On the
other hand a pathological environment can perpetuate AF and reduce therapeutic success
in cardiac surgery. Addressing these pathological patterns might offer additional therapeutic
tools to improve therapeutic success and quality of life in aged patients. These senescent
patients might benefit more than younger patients from the opportunity to terminate
anticoagulation and to gain cardiac function due to sinus rhythm.
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Protective effects of caffeine in the cardiovascular system depend on mitochondrial p27 - in aging and disease
Judith Haendeler
Epidemiological studies demonstrate that habitual coffee consumption is associated
with a lower risk for adverse cardiovascular events. We describe a novel mechanism by
which caffeine improves functions of the endothelium and the heart. The effects depend
on p27, known as a cell cycle inhibitor, and moreover, on its caffeine-induced
translocation to mitochondria. Only mitochondrial p27 phosphorylated on two specific
residues enhances migration of endothelial cells. Furthermore, it protects
cardiomyocytes against apoptosis. p27-deficiency and aging decrease respiration in
heart mitochondria and the latter is rescued by caffeine, which induces transcriptome
changes in a p27-dependent manner affecting mostly genes relevant for mitochondrial
processes. Caffeine also reduces infarct size after myocardial infarction in prediabetic
mice, and increases mitochondrial localization of p27. Moreover, p27 is necessary for
myofibroblast differentiation, and thus, proper infarct healing. Our data directly link the
protective effects of caffeine in the cardiovascular system to mitochondrial p27.
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30
Oral anticoagulation and polypharmacy in older adults with atrial fibrillation
Markus Gosch
Atrial fibrillation is the most frequent cardiac arrhythmia. The prevalence is around 1 to 2
% and increases with age up to 5 to 15 % in people aged over 80 years. It is one of the
strongest risk factors for stroke. In case of atrial fibrillation oral anticoagulation is a well
known treatment strategy that leads on one hand to a significant reduction of strokes in
these patients, on the other hand to a higher risk of bleeding. Regarding the guidelines
the indication of oral anticoagulation treatment in patients with atrial fibrillation seems to
be quite easy. However, older and very old patients are underrepresented in clinical
trials and we get only some information about this group of patients in a very short
chapter called „special situations“. Many questions in the management of older and very
old patients remain unanswered. It is not only the question about anticoagulation itself,
since few years it is more a discussion on the choice of treatment. Beside vitamin K-
antagonists, four different DOAC’s are available. Regarding the recommendations of the
guidelines DOAC‘s should be preferred compared to Vitamin K antagonists.
As mentioned in the guidelines we need special considerations for older and very old
adults. Multimorbidity, polypharmacy and geriatric syndroms have a high prevalence in
this age group and an enormous impact on management of oral anticoagulation.
Patients above 80 has a significant risk of bleeding compared to younger age groups,
but even this age group benefits from oral anticoagulation. What we need is an
individual approach based on guidelines, current literature and clinical assessment.
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Gender related aspects of the cardiovascular system and its impact on progression and aging.
Sandra Eifert
Mortality rates in cardiovascular disease differ highly between men and women,
independent of treatment options. Applying the gender lens to cardiovascular aging is of
major clinical interest. That accounts for the entire course of disease; from risk factors
via onset to therapeutic options. They may play a leading role in cardiovascular disease
progression and aging. Cardiovascular risk factors including renal failure, and hormonal
influences were reviewed and their gender-specific distribution isreported on.
The levels of sex hormones decrease with age and thus, may potentially contribute to
cardiovascular aging and the adaption to certain physiologic or non physiologic
conditions. Sex-specific factors implicating endothelial function/ dysfunction will be
recognized because of its clinical relevance to preventive and therapeutic strategies.
Outcomes of men and women in relation to age will be reported in aortic disease, aortic
stenosis as well as heart failure.
Female gender has become an independent risk factor of survival after the majority of
cardiosurgical procedures. Severely impaired left ventricular ejection fraction
independently predicts survival in men, whereas age does in females.
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Historical Overview of Research on Lifespan Extension and its Ethical Implications
George M. Martin
The human aspiration to greatly enhance one’s lifespan probably dates to the early history of Homo
sapiens and the evolution of his neocortex. Recent research on the comparative genomics of our
species and that of our closest cousin, the chimpanzee, in fact points to the emergence of specific
genetic loci responsible for our extensive six-layered cerebral cortex (KS Pollard Nature, 2006). The
idea that scientific research might bring these wishes to fruition might be dated to the 16th century
when, for example, individuals such as Robert Boyle (1627–1691), a founding member of the Royal
Society, is said to have proposed experiments as "to replace the blood of the old with the blood of
the young" (https://en.wikipedia.org/wiki/Life_extension ) – a remarkably contemporary enterprise
(https://www.technologyreview.com/s/603242/questionable-young-blood-transfusions-offered-in-us-
as-anti-aging-remedy/ )!
Most contemporary geroscientists, however, would probably point to the landmark dietary restriction
experiments on rats as the beginning of the modern era of lifespan extension research (C.M McCay,
et al. Scientific Monthly, 1934). Numerous species have since been shown to respond to such
interventions, although much more research is needed to establish dose-response curves as
functions of genetic backgrounds.
A huge volume of basic research has finally led to what may well be the first US Food and Drug
Agency approved clinical trial of an agent (Metformin) for the enhancement of lifespan
(https://www.wired.com/story/this-pill-promises-to-extend-life-for-a-nickel-a-pop). Meanwhile, a range
of substances are being consumed without such clinical trials because of their marketing as
Nutraceuticals. Ethical concerns about efficacy and about possible untoward effects in genetically
susceptible individuals who purchase “anti-aging” products need to be addressed in such cases,
however. The sale of hormones for such treatments was estimated to yield circa $50 billion in sales
back in 2009 (http://articles.chicagotribune.co2009-06-15/news/0906140132_1_anti-aging-
hormones-ama-council).
The major concern for legitimate research-based interventions to extend lifespan is the paucity of
evidence that such interventions would increase the ratios of “healthspan/lifespan” in most
individuals. One could imagine, for example, that an intervention that led to enhanced physiological
robustness and resilience might extend the duration of a lethal illness and thus decrease that ratio
(M. Kaeberlein et al., Science, 2015).
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36
Rejuvenation biotechnology: why age may soon no longer mean aging
Aubrey de Grey
People are living longer than they used to - no longer because of reductions in child
mortality, but because we are postponing the ill-health of old age. But you’ve seen
nothing yet: regenerative medicine and other new biomedical technologies will
eventually be so comprehensive that people will stay truly youthful however long they
live - which means they may mostly live very long indeed. The social and economic
consequences of this transition will pervade every aspect of our lives. I will discuss
recent progress towards the implementation of this comprehensive “rejuvenation
biotechnology”.
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The Shape of Lives to Come
Ian Richard Ground
The ethical challenges posed by clinical and technological advances in the
understanding of ageing are the subject of welcome controversy across political and
social, economic and environmental dimensions.
But ethics is not concerned not just with the good or bad, utopian or dystopian
consequences of our decisions. In a world facing all too many challenges, such
utilitarian calculations have too many variables to offer much certainty.
A rich philosophical tradition holds that Ethics also concerns our conception of what it is
to lead a flourishing – as the Greeks held – eudaemonic human life. A central idea is
that how we lead our life – our commitments, life decisions and so on - is partly
constituted by our conception of the shape of that life.
In my contribution to the panel discussion, I wish to highlight the ethical challenges
posed by the advances in understanding ageing to our conception of the shape of our
lives and what it means for such lives to flourish. If we come to think of ageing as a
disease, what are the consequences for the rest of our ethical and conceptual scheme?
How we reconcile that radical conceptual change in our thinking about ageing - that
many would have us enthusiastically embrace - with what may be a deeper truth: that it
is not merely our bodies, but we who age?
39
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40
Ethical questions of biogerontological research. An overview of the debate
Zóe Rheinsberg
Significant advances have been achieved in the field of biological ageing research.
They give raise to hopes in the context of challenges related to population ageing but
also to important ethical questions. These questions need to be addressed both by
scientific research and debate and by a broad discourse involving the general public.
The following sectors should be explored and discussed: (1) ethical, legal, and social
aspects of biogerontological research, and its methods and goals, (2) specific
challenges in research ethics, and (3) specific ethical implications for physicians. It is
crucial that the debate focuses on realistic scenarios of what could be achieved by
biogerontological research in closer future. A postponement of the onset of age-related
diseases and a moderate extension of the human lifespan seem to be a more suitable
basis for discussion than a radical life extension and the entire abolition of age-related
disease and suffering.
Concrete questions which need to be addressed relate to the necessity and dimension
of funding for biogerontological research, the accessibility of potential medical
applications, and the influence of biogerontological research on perception and images
of age, ageing, and old persons. While this is already happening in the scientific
community the public dialogue is still at its beginnings. Discourse projects can be one
approach to initiate and inform a public debate on ethical, legal, and social questions of
biogerontological research.
41
42
Epigenetic stress response and stem cell aging
Lenhard Rudolph
Adult tissue stem cells contribute to the lifelong maintenance of 0rgan homeostasis and
regeneration. However, the functionality of stem cells declines during aging and there is
emerging evidence for the clonal dominance of mutant stem cells. Both processes
contribute to the evolution of aging associated dysfunctions and diseases but molecular
mechanisms that impair the function of stem cells during aging remain incompletely
understood. Our recent work revealed that alterations in epigenetic stress responses
lead to an aberrant activation of developmental pathways that impair the self renewal
and regenerative capacity of muscle stem cells. Recent studies from the laboratory of
David Scadden indicate that heterogeneity of epigenetic memory in hematopoietic stem
cell is linked to the number of cell divisions and determines the heterogeneity in the
functionality of HSCs during aging. Interestingly, the vast majority of gene mutations
leading to clonal dominance of HSCs during aging affect epigenetic regulators.
Together, these data indicate that alteration in epigenetic memory and stress responses
represent driving forces for the decline of stem cell function and the selection of stem
cell mutation during aging. During my talk I will present new data on physiological
conditions and molecular mechanisms that may contribute to alterations of the
epigenome and gene regulation in aging stem cells and thus to disease development in
aging.
43
44
Understanding muscle stem cell regenerative decline with aging
Pura Munoz-Canove
Skeletal muscle has a remarkable capacity to regenerate by virtue of its resident Pax7-
expressing stem cells (satellite cells), which are normally quiescent in the adult. Upon
injury, quiescent satellite cells activate and proliferate, to subsequently differentiate and
form new myofibers or self-renew to restore the quiescent satellite cell pool. We have
shown that these long-lived resting stem cells attenuate proteotoxicity and avoid
senescence through basal autophagy. This protective “clean-up” system is lost during
aging, resulting in stem cell regenerative decline. We therefore propose that loss of
proteostasis is causally implicated in muscle stem cell aging, a process that can be
targeted for rejuvenation.
45
46
Bioengineered matrices to regenerate the heart – The Zurich LifeMatrix Programm
Maximilian Emmert
Cardiovascular disease represents a major cause of morbidity and mortality worldwide
accounting for the death of numerous patients. The concept of regenerative medicine
comprising cell-based therapies, bio engineering technologies and hybrid solutions has
been suggested as a next generation approach for affected patients. In particular,
cardiovascular tissue engineering has been demonstrated to be a promising concept to
generate living, autologous cardiovascular structures with the ability to remodel and to
grow which may be particularly beneficial for children. However, although these
regenerative concepts have shown great potential in experimental studies, the safe and
effective translation into a widespread clinical setting is still limited. Furthermore, the
currently utilized ``classical`` approaches are still too complex with regards to logistical
and financial effort. Therefore, significant simplification is needed and in particular, so
called off-the-shelf concepts may represent an attractive option to overcome the current
problems towards clinical translation.
47
48
Mesenchymal Stem Cells-on-Chips/Organ-on-Chips/Chips in Mesenchymal Organs – Monitoring Healing Kinetics:
an ageing perspective
Günter Lepperdinger
Stem cells play an important role during development and regeneration, and their
dysfunction is associated with a variety of diseases, such as sarcopenia, osteopenia, or
several classes of cancer. Activated stem cells bring forth specific progenitor cells,
which promote tissue remodeling, repair and regeneration (RRR). Adult human
mesenchymal stem cells (MSC) can be readily isolated from different sources, and
many clinical trials have been successfully performed previously. Consequently, many
therapies recapitulating RRR are currently being implemented in clinical settings. Major
tissue changes during RRR are constituted through the deposition of material into the
pericellular space, be it soft extracellular matrix (ECM) or dense load-bearing
substances such as collagenous hydroxyapatite. In vivo quantitative analyses thereof
are still demanding. We first developed impedance microsensors for in vitro use, which
allowed to assess cell attachment and viability as well as bipotential differentiation
capacity, thus proofing that impedance changes enables documentation of MSC quality
in a fast, efficient and reliable fashion. Based on this biochip technology we have
developed implantable titanium dioxide-coated bioimpedance sensors. After
implantation, the sensory system revealed in subcritical defects of rabbit calvaria distinct
differences between granulation and soft tissue formation within two weeks, thus
allowing the accurate assessment of bone healing prior the formation of a cortical bone
layer. Hence this technology enables diagnosis of early healing deficiencies or
decreasing implant stability. Accounting for that specific early information allows
commencing with and adjustment of appropriate medicinal interventions.
49
50
Effects of pharmacological inhibition of aging-associated pathways on Drosophila lifespan
Alexey Moskalwev
Mutations of the PI3K, TOR, iNOS, and NF-κB genes increase lifespan of model
organisms and reduce the risk of some aging-associated diseases. We studied the
effects of inhibitors of PI3K (wortmannin), TOR (rapamycin), iNOS (1400W), NF-κB
(pyrrolidin dithiocarbamate and QNZ), and the combined effects of inhibitors: PI3K
(wortmannin) and TOR (rapamycin), NF-κB (pyrrolidin dithiocarbamates) and PI3K
(wortmannin), NF-κB (pyrrolidine dithiocarbamates) and TOR (rapamycin) on
Drosophila melanogaster lifespan and quality of life (locomotor activity and fertility). Our
data demonstrate that pharmacological inhibition of PI3K, TOR, NF-κB, and iNOS
increases lifespan of Drosophila without decreasing quality of life. The greatest lifespan
expanding effect was achieved by a combination of rapamycin (5 μM) and wortmannin
(5 μM) (by 23.4%). The treatment of Drosophila melanogaster with 10 non-steroidal
anti-inflammatory drugs (NSAIDs: CAY10404, aspirin, APHS, SC-560, NS-398, SC-
58125, valeroyl salicylate, trans-resveratrol, valdecoxib, licofelone) leads to extension of
lifespan, delays age-dependent decline of locomotor activity and increases stress
resistance. The lifespan extending effects of APHS, SC-58125, valeroyl salicylate,
trans-resveratrol, valdecoxib, and licofelone were more pronounced in males,
valdecoxib and aspirin - in females. We demonstrated that lifespan extension effect of
NSAIDs was abolished in flies with defective genes involved in Pkh2-ypk1-lem3-tat2
pathway.
51
52
The Path towards Translating Senolytic Drugs and Other Interventions Targeting Aging Processes into Clinical Treatments
James Kirkland
Interventions targeted at fundamental mechanisms of aging hold promise for enhancing
healthspan by delaying, preventing, or alleviating a range of age-related diseases and
conditions—a theory termed the “geroscience hypothesis.” Among the more promising
interventions are senolytic agents, drugs that selectively eliminate senescent cells.
Early, proof-of-concept Phase 2 clinical trials will be a critical step in translating these
agents and others emerging from cell culture and animal preclinical studies into clinical
practice. The goals of such proof-of-concept trials for newer interventions include
generating preliminary signals of efficacy in age-related diseases or outcomes that will
test the value of proceeding to larger Phase 3 trials. Drugs that target aging processes,
such as metformin, for which there are already extensive clinical data related to other
indications, are at the point of being tested in Phase 3 trials, such as the proposed
TAME study. In addition to generating data about effectiveness, such trials need to be
designed to contribute data and biological samples to support basic research through
strategic networks, as well as furthering dialogue with regulatory agencies about
registration indications. Frameworks for proof-of-concept and later stage trials that
target age-related chronic diseases, geriatric syndromes, or resilience to stressors will
be considered. We propose that a strategic infrastructure and shared resources are
needed to accelerate translation of therapies targeting fundamental aging processes
into clinical treatments.
53
54
Cellular senescence and aging
Judith Campisi
Cellular senescence is a complex cell fate decision that entails an essentially
irreversible arrest of cell proliferation coupled to a multi-component secretory phenotype
(termed the senescence-associated secretory phenotype or SASP). The senescence
response, and particularly the SASP, is most likely an evolutionary trade-off. That is, it
likely evolved to benefit young organisms by promoting tissue repair and remodeling,
but becomes maladaptive and compromising to tissue homeostasis later in life. This
trade-off arises because the SASP includes a variety of pro-inflammatory cytokines,
chemokines, growth factors, proteases and bioactive lipids, many of which can cause or
contribute to local and systemic inflammation – a hallmark of aging. Several lines of
recent evidence, primarily from mouse models, suggest that the selective clearance of
senescent cells from aged individuals can ameliorate a variety of age-related
phenotypes and pathologies. The challenge for applications to humans will be to
preserve the beneficial effects of senescent cells, while eliminating their deleterious
effects later in life.
55
56
DNA damage: impact on aging, neurodegeneration and the effect of nutritional interventions
Jan Hoeijmakers
The molecular basis of aging (-related diseases) is one of the main unsolved questions in
biology. Aging appears remarkably plastic: e.g. suppressing insulin signalling extends lifespan
in worms, flies and mice. On the other hand, virtually all premature aging syndromes in man
provide links with genome instability. We have generated mouse models which strikingly mimic
human DNA repair deficiency disorders and display wide-spread accelerated aging, revealing a
strong link between unrepaired DNA damage and many features of aging. E.g. Ercc1∆/- mice
defective in ≥3 repair pathways show extensive premature multi-morbidity in post-mitotic and
proliferative tissues limiting lifespan to 4-6 month. Simultaneously these mice exhibit an anti-
aging ‘survival response’, which suppresses growth and enhances maintenance, resembling the
longevity response induced by dietary restriction (DR). Interestingly, subjecting these progeroid,
dwarf mutants to actual (30%) DR tripled remaining lifespan, and drastically retarded numerous
aspects of accelerated aging, with the neuronal system benefitting disproportionally. E.g. DR
animals retained 50% more neurons and maintained full motoric function, delaying motor
decline >20(!)-fold. Repair-deficient Xpg-/- mice also showing many premature aging symptoms
responded similarly. The DR response in Ercc1∆/- mice resembled wt DR including (further)
reduced insulin signaling. Interestingly, ad libitum Ercc1∆/- liver expression profiles showed
declining expression of long genes, consistent with genome-wide accumulation of stochastic,
transcription-blocking lesions, which affect long genes more than short ones. Similar findings
were made in human brain profiles upon aging, demonstrating relevance for normal aging in
humans. DR in repair-deficient mice alleviated this decline, indicating that DR prolongs genome
function. We will present phenotypes of conditional DNA repair models targeting aging to
selected organs and connections with the unfolded protein response and proteinopathies
(Alzheimer’s and Parkinson diseases). Our findings strengthen the link between DNA damage
and aging, establish Ercc1∆/- mice as powerful model for identifying interventions to promote
healthy aging, reveal untapped potential for reducing endogenous damage, provide new venues
for understanding the molecular mechanism of DR, and suggest a counterintuitive DR-like
therapy for human progeroid genome instability syndromes and DR-like interventions for
preventing neurodegenerative diseases.
57
58
Posters (in alphabetical order)
The Poster Award
Ceremony
Sunday 3rd
of September from 11:30 to 11:45
59
(1) Carbon nanoparticles cause loss of gap junctional intercellular
communication in the lung and the endothelium
Nilo Ale-Agha1, Tim Spannbrucker1,2, Klaus Unfried2, Yogi Altschmied1, Jojo Haendeler1
1Heisenberg group - Environmentally induced Cardiovascular Degeneration, Clinical Chemistry, University of
Duesseldorf, and IUF-Leibniz Research Institute for Environmental Medicine, Duesseldorf, Germany, 2Environmentally induced skin and lung aging, IUF-Leibniz Research Institute for Environmental Medicine,
Duesseldorf
Introduction: Gap junctional intercellular communication (GJIC) pathways are involved in a
variety of vascular and pulmonary functions. Gap junctions composed of connexin (Cx)
molecules are clusters of intercellular channels connecting adjacent cells. The loss of Connexin
43 (Cx43), one of the major connexins in the vascular and pulmonary systems, and GJIC was
suggested to be related to senescence and many cardiovascular and pulmonary disorders. It is
known that environmental factors like industrial or consumer derived pollution affect the
cardiovascular and pulmonary systems. Senescent endothelial cells are associated with
cardiovascular diseases, and lung cell senescence is supposed to impair lung function. We
have shown that non-cytotoxic, non-inflammatory concentrations of carbon nanoparticles (CNP)
induce senescence both in human primary endothelial cells (EC) and rat lung epithelial cells
(RLE).
Objective: The aim of this study was to investigate whether CNP affect GJIC in the lung and
endothelium and might therefore be involved in cardiovascular and pulmonary aging processes.
Material and Methods: CNP particle suspensions were applied to EC, RLE or C57Bl/6 mice
(pharyngeal aspiration). GJIC was measured by dye transfer assay. The localization of Cx43
was assessed by immunofluorescence staining in lung sections. Phosphorylation and protein
levels of Cx43 were determined by immunoblot.
Results: Two weeks incubation of cells with non-toxic CNP concentrations showed increased
levels of senescence-associated beta-galactosidase and the cell cycle inhibitor p21, two known
markers for cellular aging. The same concentration of CNP resulted in a dramatic reduction of
GJIC in both cell types, which was due to a significant change in phosphorylation and
internalization of Cx43. Thus, CNP-induced senescence is accompanied by loss of GJIC. CNP
applied in non-inflammatory concentrations into mice in vivo reduced Cx43 protein levels, while
the phosphorylation of Cx43 was upregulated in the lungs of CNP treated mice.
Conclusion: These data indicate that Cx43, which is essential for endothelial and pulmonary
functionality, is affected by CNP and that loss of GJIC is involved in CNP-induced senescence.
Further studies have to clarify whether a causal link between CNP-induced senescence and
loss of GJIC exists.
60
(2) Protective functions of mitochondrial TERT in age-associated
cardiovascular diseases
Yogi Altschmied, Christine Goy, Mark Zurek, Florian von Ameln, Niloofar Ale-Agha, Jojo Haendeler
Heisenberg group - Environmentally induced Cardiovascular Degeneration, Clinical Chemistry, University of Duesseldorf, and IUF-Leibniz Research Institute for Environmental Medicine, Duesseldorf, Germany
Introduction: Age-associated cardiovascular disorders are associated with mitochondrial
dysfunction, which causes reduced respiratory chain activity, decreased mitochondrial ATP
production, an increase in reactive oxygen species (ROS) and cell death. We have previously shown
that the catalytic subunit of telomerase, Telomerase Reverse Transcriptase (TERT) is not only
present in the nucleus, but also in mitochondria, where it binds to mtDNA and protects it from
damage. Furthermore, TERT improves mitochondrial respiration and reduces mitochondrial ROS
formation and both these functions can presumably ascribed to mitochondrial, rather than nuclear
TERT.
Objective: The aim of this study was to characterize the functions of mitochondrial TERT in age-
associated cardiovascular diseases ex vivo and in vivo.
Materials and Methods: To analyze specifically the role of mitochondrial TERT we transduced
different cardiovascular cells with lentiviral expression vectors for mitochondrially-targeted TERT.
Furthermore, we created a unique new mouse model, in which the animals contain TERT exclusively
in the mitochondria (mitoTERT mice). Apoptosis as well as cytosolic and mitochondrial ROS were
measured by flow cytometry and ATP with a luminometric assay. Respiratory chain activity was
measured by oxygraphy. Myofibroblast differentiation of isolated cardiac fibroblasts was assessed
measuring specific markers by immunoblotting and semi-quantitative real-time PCR. Experimental
myocardial infarction was set using ligation of the left ascending coronary artery in an open chest
model.
Results: Mitochondrial TERT showed protective functions in different cells of the cardiovascular
system, as it specifically reduced mitochondrial ROS levels, protected endothelial cells and
cardiomyocytes against apoptosis. Moreover, mitochondrial TERT was sufficient to compensate for
TERT-deficiency in mitochondrial respiration and ATP production in the heart as well as in disturbed
cardiac myofibroblast differentiation. In addition, while TERT-deficient animals showed worse
outcome after myocardial infarction compared to wildtype littermates, mitochondrial TERT was
sufficient to rescue this defect.
Conclusion: Our data show for the first time that mitochondrial TERT is required and sufficient to
protect cells of the cardiovascular system ex vivo and in vivo against damage and loss of
functionality commonly observed in age-associated cardiovascular diseases and complications, like
e.g. atherosclerosis and myocardial infarction.
61
(3) Human HSCs show changes in frequency, polarity and cell
division kinetics upon aging
Amanda Amoah, Anja Keller, Ramiz Emini, Markus Hönicka, Andreas Liebold, Carolina Florian, Hartmut Geiger.
Hematopoietic stem cells (HSC) are crucial for maintaining blood homeostasis
throughout the life of an organism. Recent findings using mouse models, however,
show distinct functional and phenotypic changes in HSCs upon aging, which are
reversible. To better explore the prospects of rejuvenation, we sought to characterize
aging-induced changes in human HSCs. Our results show that although the number of
total bone marrow (BM) cells remain unchanged, the number of multipotent
hematopoietic stem and progenitor cells (HSPC) (CD34+CD38-) and the number of
HSCs (CD34+CD38-CD90+) increase upon aging. Furthermore, we observed a delay in
aged HSCs in the response to cytokine stimulation ex vivo. Similar to murine HSCs,
aged human HSCs show a significant decrease in the frequency of cells that present
with a polar distribution of tubulin and the small RhoGTPase Cdc42 in the cytoplasm
and Ac-H4K16 in the nucleus. Collectively these findings identify functional and
phenotypic changes occurring in human HSCs upon aging.
62
(4) Dietary flavanol monomer (-) epicatechin enhances endothelial
function - causal role of SNAP23?
Karin Aufenvenne1, Christine Goy1, Christian Heiss2, Yogi Altschmied1, Jojo Haendeler1
1Heisenberg group - Environmentally induced Cardiovascular Degeneration, Clinical Chemistry, University of
Duesseldorf, and IUF-Leibniz Research Institute for Environmental Medicine, Duesseldorf, Germany, 2Division of Cardiology, Pulmonology, and Vascular Medicine, University of Duesseldorf
Introduction: Diet is a major determinant of health as well as healthspan. Cocoa
flavanols have emerged as important dietary bioactives that may be able to improve
cardiovascular health. However, the mechanisms of action are not established.
Objectives: In the present work, we aimed at identifying molecules activated by
flavanol related bioactives, e.g. (-) epicatechin using gene expression analyses and
confirmed the findings with endothelial cell models. We hypothesized that regular
consumption of plant flavanols may lead to improved endothelial function.
Material and Methods: Microarray based analyses of whole blood samples from
healthy volunteers. Hypothesis driven analyses of gene expression results; analysis of
SNAP23 by immunoblot; measurements of NO bioavailabilty and eNOS phosphorylation
were performed. Transient transfections as well as lentiviral transductions of endothelial
cells with expression vectors for SNAP23 were performed.
Results: We first performed microarray based expression analyses of whole blood
samples from healthy volunteers randomly and double blinded consuming cocoa
flavanols (450 mg bi-daily) or placebo over one month. In parallel with improved
vascular function, the analyses showed a striking upregulation of approximately 500
transcripts. One of the most prominent upregulated genes by flavanols is SNAP23. To
investigate a causal relationship ex vivo, we first investigated whether (-) epicatechin
increases SNAP23 protein levels. Indeed, SNAP23 was increased up to 3-fold. Along
the same line eNOS was activated and NO bioavailability was increased by (-)
epicatechin. Moreover, overexpression of SNAP23 in combination with (-) epicatechin
did not enhance eNOS phosphorylation further, suggesting the same pathway.
Conclusion: Flavanols increase flow-mediated dilation. This effect depends on NO and
on SNAP23 as demonstrated in endothelial cells.
63
(5) Novel Amide AGEs as Oxidative Stress Markers
T. Baldensperger, Halle (Saale)/D, Prof. Dr. M. A. Glomb, Halle (Saale)/D
Prof. Dr. M. A. Glomb, MLU Halle-Wittenberg, Institute of chemistry – Food chemistry, Kurt-Mothes Straße 2, 06120 Halle (Saale), D. marcus.glomb@chemie.uni-halle.de
The demographic change in Germany and improving medical care lead to progressive
ageing of our society. As a result an increase of age associated diseases like diabetes,
atherosclerosis and cancer is expected. In order to face these future problems a
detailed knowledge of molecular mechanisms leading to the decline of organ functions
and tissue homeostasis is crucial. There is rising evidence that posttranslational
modifications of proteins play a key role in ageing processes. One major aspect of
posttranslational modification in vivo is the enzymatic acylation of proteins. Recent
publications indicate that Maillard processes and fat metabolism also lead to
accelerated acylation of proteins. This means fundamentally important enzymatic
processes are paralleled by non-enzymatic pathways. The physiological importance and
correlation to ageing of such non-enzymatic acylation pathways is not understood yet.
In order to analyze non-enzymatic acylation a highly sensitive HPLC-MS/MS method
was developed. Authentic reference standards of the novel amide advanced glycation
endproducts (AGEs) N-glyoxylyl lysine and N-pyruvoyl lysine were synthesized. First
screenings revealed N-glyoxylyl lysine as a glyoxal and N-pyruvoyl lysine as a
methylglyoxal specific AGE. N-glyoxylyl lysine was established as a further product of
the N-carboxymethyl lysine (CML) reaction cascade and N-pyruvoyl lysine as a further
product of the N-carboxyethyl lysine (CEL) reaction cascade. Consequently we
proposed a mechanism including isomerization and oxidation of the central intermediate
of the CML/CEL reaction cascade in analogy to the oxidation step established for the
formation of carboxylic acids in the well-known Strecker degradation. In fact,
N-glyoxylyl lysine and N-pyruvoyl lysine were formed exclusively under aerated
conditions. In addition, artifact amide formation directly from glyoxylic acid, pyruvic acid
or from aldol condensation products were ruled out. Due to the strict correlation of
N-glyoxylyl lysine and N-pyruvoyl lysine formation to oxidative conditions we aim to
establish these novel amide AGEs as marker structures for oxidative stress in vivo.
64
(6) Impact of advanced glycation end products on macrophage activation.
Veronika Bezold1, Rüdiger Horstkorte1, Toni Ehrhardt2, Matthias Jung2, Kaya Bork1
1Institute for Physiological Chemistry, Martin-Luther-University Halle-Wittenberg, Halle (Saale), Germany
2Department of Psychiatry, Psychotherapy and Psychosomatics, Martin-Luther-University Halle-Wittenberg, Halle
(Saale), Germany
Advanced glycation end products (AGEs) and their receptor have been implicated in the
progressions of many diseases, such as diabetes and atherosclerosis, and are also
critical for pathologic changes in chronic degenerative diseases, such as Alzheimer's
disease and Parkinson's disease. AGEs increase with age and have been investigated
as a marker for aging. It is discussed if age-associated changes in immune system
development and function are related to an increased level of AGEs [1].
Differentiation of monocytes into macrophages is a key step in innate immune
response. The human monocytic cell line THP-1 can easily be differentiated into
macrophages using various chemical stimuli. Therefore this cell line is often used as a
model for differentiation studies. We treated undifferentiated and differentiated THP-1
cells with methylglyoxal (MGO) to increase the formation of AGEs and analyze the
activation of the cells. We could show that the increased formation of AGEs influences
the expression of the activation marker CD14, interleukin 1-beta, interleukin 6,
interleukin 8, interleukin 10 and TNF-alpha. In addition, we analyzed the expression of
the AGE-receptor RAGE after treatment with MGO. RAGE is a transmembrane receptor
expressed in various immune cells. The interaction between RAGE and its ligands is not
clearly understood regarding cellular signaling, though it is thought to result in pro
inflammatory gene activation [2].
References [1] G. Glorieux, R. Vanholder, N. Lameire, Eur. J. Clin. Invest. 2011, 12, 1015-1018.
[2] A. Bierhaus, S. Schiekofer, M. Schwaninger, M. Andrassy, P.M. Humpert, J. Chen, M. Hong, T. Luther, T. Henle, I. Klöting, M. Morcos, M. A. Hofmann, H. Tritschler, B. Weigle, M. Kasper, M. A. Smith, G. Perry, A.M. Schmidt, D. M.Stern, H.U.Häring, E. Schleicher, P.P. Nawroth, Diabetes 2001, 50, 2792-2809.
65
(7) Development of an age-specific genome-scale model of skeletal
muscle metabolism
Andrea Cabbia, MSc
Biomedical Engineering Eindhoven University of Technology (NL)
Skeletal myocytes are among the most metabolically active cell types, implicated in
nutrient balance, contributing to the insulin-stimulated clearance of glucose from the
blood, and secreting myokines that contribute in regulating inflammation and the ageing
process. The loss of muscle mass and strength with age (sarcopenia), is a risk factor for
cardiovascular and metabolic diseases, it increases the risk of falls, of developing frailty
and disabilities, and results in an impairment in the quality of life and autonomy of an
individual.
An active lifestyle is the most immediate and accessible treatment to prevent
sarcopenia, with a considerable impact on the ageing process: PANINI is a European
Training Network whose aim is understanding how lifestyle factors can influence healthy
ageing. In this context, we present the first age-specific genome-scale metabolic model
of the skeletal muscle, a mathematical representation of the myocyte metabolic network
in the elderly, built using RECON2, the human metabolic reconstruction, and gene
expression data, gathered from older adults' muscle tissue biopsies.
This model will be used to analyze patient-specific data for potential mechanisms able
to explain the different ageing paces of different individuals and to investigate the
effectiveness of different nutritional and physical exercise regimes in stimulating post-
exercise protein synthesis, which is often impaired in the elderly.
The aim is to identify an optimal and personalized lifestyle change intervention able to
prevent the onset of sarcopenia.
66
(8) Involvement of mitophagy in the elimination of damaged
mitochondria during the process of UVB-induced senescence
Maria Cavinato1, Rafal Koziel1, Sophia Wedel1, Brigitte Jenewein1, Martin Hermann2, Nikolaus Romani3, Pidder Jansen-Dürr1
1Institute for Biomedical Aging Research, Universität Innsbruck, Innsbruck, Austria
2Department of Anesthesiology and Critical Care Medicine, Medical University of Innsbruck, Innsbruck, Austria
3Department of Dermatology and Venerology, Medical University of Innsbruck, Innsbruck, Austria
Skin aging is the result of two types of aging, “intrinsic aging” an inevitable consequence
of physiologic and genetic determined changes and “extrinsic aging” which is dependent
on external factors like exposure to sunlight, smoking, dietary habits among others. UV
and other forms of ionizing radiation cause skin injury through the generation of free
radicals and other oxidative byproducts as well as DNA damage. The oxidized proteins
are generally degraded by the ubiquitin-proteasome system or by autophagy, and
alterations on these pathways lead to accumulation of damaged molecules. The activity
of proteasomes and autophagic organelles is regulated by overlapping signals, and
regulatory cross-talk between both quality control systems has been described.
Likewise, excessive ROS production leads to impairment of mitochondria with
consequences that are related to several age-related diseases as well as to the
physiology of normal aging. We have previously demonstrated that inhibition of
proteasomal degradation of damaged proteins and activation of autophagosome
formation are early events in UVB-induced senescence of human dermal fibroblasts
(HDF), dependent on UVB-induced accumulation of reactive oxygen species (ROS).
Here we show that UVB treatment of HDFs leads to impaired mitochondrial function,
damage to mitochondrial structure and disruption of mitochondrial network and that
these damaged organelles are eliminated by mitophagy. Under these conditions,
mitophagy receptor Bnip3L/Nix is differentially regulated in fibroblasts. Elimination of
Bnip3L/Nix increases cell proliferation and inhibits mitophagy activation upon UVB
treatment. These findings have potential implications for fundamental as well as
translational research into skin aging, and in particular photoaging.
67
(9) Investigating the relationship between protein glycation and
aggregation in cells and tissues
Simone Di Sanzo
1. Ori, A. et al. Integrated Transcriptome and Proteome Analyses Reveal Organ-Specific Proteome Deterioration in Old Rats. Cell Syst. 1, 224–237 (2015).
2. Salahuddin, P., Rabbani, G. & Khan, R. The role of advanced glycation end products in various types of neurodegenerative disease: a therapeutic approach. Cell. Mol. Biol. Lett. 19, (2014). 3. Vlassara, H. & Uribarri, J. Advanced glycation end products (AGE) and diabetes: cause, effect, or both? Curr. Diab. Rep. 14, 453 (2014).
4. Stinghen, A. E. M., Massy, Z. A., Vlassara, H., Striker, G. E. & Boullier, A. Uremic Toxicity of Advanced Glycation End Products in CKD. J. Am. Soc. Nephrol. 27, 354–370 (2016).
5. Snow, L. M., Fugere, N. A. & Thompson, L. V. Advanced Glycation End-Product Accumulation and Associated Protein Modification in Type II Skeletal Muscle With Aging. Journals Gerontol. Ser. A Biol. Sci. Med. Sci. 62, 1204–
1210 (2007).
The homeostasis of the proteome of cells is required to maintain the function of organs
and it was shown to decline during aging. The proteostasis is influenced by changes in
protein localization, protein abundance and post-translational modification1. The
formation of advanced glycation end products (AGEs) is a non-enzymatic
posttranslational modification that has been shown to influence the activity and
aggregation propensity of both extracellular and intracellular proteins. AGEs are
associated with oxidative stress and inflammation, processes that lead to cardiovascular
disease, diabetes, chronic kidney disease (CKD), and neurodegenerative diseases2–4.
Although a handful of specific AGE-modified proteins have been identified for example
in aging skeletal muscle5, a detailed characterization of the targets of AGEs and the
relationship between this modification and protein aggregation are currently missing. In
this project, we aim to identify the preferential protein targets of AGEs in cells and
tissues. For this purpose, we develop a proteomic workflow based on selective
enrichment of AGE-modified proteins coupled to mass spectrometry for protein
identification. In parallel, we plan to systematically investigate the relationship between
protein glycation and aggregation by monitoring proteome-wide changes in protein
aggregation propensity induced by exposure to glyoxal. The combined study will allow
us to investigate whether AGEs are mechanistically linked to changes in protein
aggregation propensity. In the future, we plan to extend our work to primary cells and
tissue to study the targets of AGEs during aging and the effect of dietary interventions
and obesity on the extent and specificity of protein glycation.
Keyword: proteostasis, ageing, AGEs, glycation, mass spectrometry, target, aggregation
68
(10) Analysis of Late-Onset Alzheimer’s Disease Using Patient-Specific In-Vitro Models
Ehrhardt T1, Jung M1, Bezold V2, Bork K2, Hartmann C1, Giegling I1, Rujescu D1 1 Martin Luther University Halle-Wittenberg, Department Psychiatry, Psychotherapy, and Psychosomatic Medicine
2 Martin Luther University Halle-Wittenberg, Department of physiological chemistry
Background and Aim of the Study: Alzheimer´s disease (AD) is the most common form of dementia.
Environmental and genetic factors contribute to the risk for AD, but the underlying disease mechanisms are poorly
understood. AD is a genetically complex disorder and shows heritability of up to 80%. Glia cells including astrocytes
and microglia are affected in AD patients contributing to the tau pathology and the accumulation of neurotoxic
amyloid as well. Genome-wide association studies (GWAS) allowed the identification of DNA variations associated
with an elevated risk for AD in recent years. Together with other groups, we identified a number of AD susceptibility
genes including CD33, SORL1, ABCA7, and TREM2, which highly recommends that the immune system plays a
major role in onset, progression and treatment of AD. Our research is part of the psychiatric university clinic in Halle
(UKH). Therefore, we have access to blood cultures of AD patients of our clinic suitable for the generation induced
pluripotent stem cells (iPS). Pluripotent stem cells can be differentiated into patient-specific microglia, which
represent the major part of the immune cells in the human brain. The diversity of microglia functions contributes to a
healthy immune system. Disturbed microglia function represents an important disease mechanism in AD. Activation
of inflammatory microglia leads to production of toxic substances and decreased phagocytosis. Besides the way of
activation, the polarization state mainly depends on the expression level of the two recently described surface marker
genes CD33 and TREM2. These genes are contrary and have to be in a homeostasis level for a controlled
phagocytosis. Mutations in this genes lead to an imbalance and can influence the amount of amyloid uptake.
Material and Methods: Blood was collected from AD patients carrying risk variants in AD susceptibility genes for the
generation of iPS cells. SNP variants in those genes were genotyped based on their potential function according to
literature and genomic localization (exonic, binding domain, or promoter affecting SNPs) and tested for association to
AD. Reprogrammed iPS cells carried the genetic background of a certain AD patient. Pluripotency was characterized
by alkaline phosphatase staining, the expression of pluripotency markers, and the differentiation into the three germ
layers. Crucial pluripotency markers are OCT4, SOX2, NANOG, KLF4, MYC, and LIN28. AD iPS cells were
differentiated into astrocytes and microglia and the differentiation status was characterized by the expression of
crucial glia cell markers. Further, AD susceptibility genes recently published in GWAS were analysed.
Results: The protein expression was successfully induced shown by IF and WB analysis. Cells were also screened
for the most efficient induction of the three germ layers and the induction of neural cell fates including glia cell fates.
The established AD-specific iPS cell lines from AD patients represent a powerful tool for the analysis of molecular
and cellular disease mechanisms. We established 4-step protocol for the generation of AD-specific microglia enabling
the focused analysis of AD-associated risk variants. The protocol was verified by morphology, FACS analysis, IF
analysis, and RNA expression of crucial microglia marker including hematopoetic lineage markers and the induction
of well-described microglia markers such as IBA1. The disease-specific analysis of the genetic background of AD
patients in iPS cells represents a completely new approach for the understanding of AD genetics and AD-associated
risk variants.
Discussion: Together, combining molecular genetics of AD for the investigation of risk variants and iPS cell
technology for the generation of patient- and disease-specific stem cells provide a promising approach to
characterize known disease mechanisms. These approaches contribute to the understanding of known disease
mechanisms and to the discovery of unknown disease aspects as well.
69
(11) Proteasome deficiency in endothelial cells is causally linked to senescence
Odeta Meçe1, Govind M. Pai1, Nderim Kryeziu1, Claudia Ender1, Tilman Grune2, Regine Heller1
1Institute for Molecular Cell Biology, Centre for Molecular Biomedicine (CMB), Jena University Hospital,
2Deutsches Institut für Ernährungsforschung, Potsdam - Rehbrücke (DIfE)
Senescence is thought to contribute to endothelial dysfunction, which ultimately leads to
development of cardiovascular disease. The current project characterizes proteasomal
protein degradation in endothelial cells undergoing replicative or stress-induced
senescence in vitro or chronological ageing in vivo.
Our results demonstrate that human endothelial cells become senescent in vitro within
25 cumulative population doublings. Senescence was verified by increased number of
senescence-associated β-galactosidase (SA-β-gal)-positive cells, increased cell size and
granularity and upregulation of the p53/p21 pathway. In addition, protein carbonylation
and nitration as well as reduced GSH levels were observed. Studies on the expression
of proteasomal subunits revealed a decline of the β5 and β2 subunits in senescent
cells. In parallel, trypsin- and chymotrypsin-like activities were decreased. Similar
observations were made in primary endothelial cells derived from old mice (24 months
old) when compared to young mice (2 months old). Furthermore, treatment of early
passage human endothelial cells with H2O2 leading to premature senescence was
characterized by a decline of proteasomal trypsin- and chymotrypsin-like activities
without loss of β5 and β2 subunits.
Transient inhibition of proteasomal activity in endothelial cells by MG132 or bortezomib
led to premature senescence characterized by SA-β-gal-positive cells and increased
expression of p53 and p21. In parallel, persistent accumulation of carbonylated proteins
was observed indicating a role of oxidative stress in the development of senescence.
From these results we suggest a relationship between proteolytic insufficiency and
senescence, which may contribute to vascular ageing.
70
(12) BENEFICIAL EFFECT OF ASCORBATE IN RENAL AGING
K Forman1,2, F Martínez1, M Cifuentes3,4, R Bertinat1, K Salazar1 and F Nualart1
1Center for Advanced Microscopy CMA BIO-BIO, Neurobiology and Stem Cell Laboratory. University of Concepcion,
Barrio Universitario s/n, Chile 2 Department of Nutrition and Dietetics, University of Concepcion, Barrio Universitario sn, Chile
3Department of Cell Biology, Genetics and Physiology, IBIMA, BIONAND, Andalusian Center for Nanomedicine and
Biotechnology, University of Malaga, Malaga, Spain. 4Networking Research Center on Bioengineering, Biomaterials and Nanomedicine, (CIBER-BBN), Malaga, Spain.
Renal aging might be due to damage caused by inflammation, apoptosis and oxidative
stress. Vitamin C (VC) or ascorbic acid (AA) is a major antioxidant and plays an
essential role in defending against cellular damage. AA transport is mediated by high
affinity sodium-dependent transporters (SVCTs). The purpose of this study was to
evaluate the protective effect of AA in the kidney of male senescence-accelerated mice
(SAMP8). Also the influence of AA on the morphological parameters and on the renal
SVCT1 expression were investigated.
SAMP8 mice at 2 (young) and 9 (aged) months of age were used (n=15). One group
was treated with AA (10mg/Kg-IP-6 months). The expression of SVCT1 were analyzed
by conventional immunohistochemical/multi-labeling immunofluorescence analysis and
laser capture microdissection coupled to qRT-PCR. The expression of interleukin 1(IL-
1b), inducible nitric oxide synthase (iNOS) and survivin were measured by qRT-PCR.
The morphological study was performed using Hematoxylin-eosin staining.
71
(13) Influence of glyoxal and methylglyoxal on mesenchymal stem cell differentiation
Susan Garbe1, Maria Schindler1, Juliane-Susanne Jung1, Alexander Navarrete Santos2, Tom Seeling1 and Anne Navarrete Santos1
1 Department of Anatomy and Cell Biology, Martin Luther University Halle-Wittenberg, Faculty of Medicine, Halle
(Saale), Germany 2Centre for Medical Basic Research, Martin Luther University Halle-Wittenberg, Faculty of Medicine, Halle (Saale),
Germany
Mesenchymal stem cells (MSC) differentiate into chondrocytes, bone, fat and muscle cells
and therefore play a crucial role in human development. About three quarters of the body
mass arises from MSC and after embryonic development they are important. The
dicarbonyls methylglyoxal (MGO) and glyoxal (GO) are inducers of advanced glycation end
products (AGEs). For AGEs an adverse influence in diabetes, renal dysfunction and vessel
stiffness has been described. The physiological concentrations of MGO and GO in human
plasma are 120 and 200 nM respectively. In the plasma of diabetic patients these
concentrations are up to six fold higher. Thus, we hypothesize that high MGO and GO
levels lead to failures in the cellular differentiation with consequences for mesenchymal cell
fate determination and multipotency.
The cell model C3H10T1/2, an immortalized multipotent mesenchymal stem cell line from
mouse embryos was used to investigate how MGO and GO affect the differentiation of
mesenchymal stem cells into adipocytes. Undifferentiated C3H10T1/2 cells were exposed
for 72 hours after reaching confluent stage to 50 and 100 µM of MGO and GO, alone and
simultaneously. The cellular formation of the AGEs MG-H1 (methylglyoxal-derived
hydroimidazolone 1) and carboxymethyllysine (CML) and the senescence and apoptosis
marker p21 was then analysed by flow cytometry. Treatment with 100 µM GO and MGO
significantly increased the expression of p21. CML and MG-H1 were detected after
exposure to MGO and GO alone and in combination. After then MGO/GO-treated
C3H10T1/2 cells were differentiated into adipocytes using induction media containing
insulin, dexamethasone and isobutylmethylxanthine (for 48 hours). The grade of adipogenic
differentiation was analysed by flow cytometry with Nile Red staining at day 13. An
exposure to MGO and GO for 72 hours did not significantly affect the differentiation of the
C3H10T1/2 cells into adipocytes.
We conclude that pathophysiological concentrations of MGO and GO affect mesenchymal
stem cells increasing AGE levels and p21 expression, whereas adipogenic differentiation
was not changed. Supported by DFG GRK 2155 ProMoAge
72
(14) Neuroblastoma Metastasis Driven by Methylglyoxal
VS.Gnanapragassam1, M.Nagasundaram1, M.Scheer1 and R. Horstkorte1. 1 Institute of Physiological Chemistry, Martin Luther University Halle-Wittenberg, Germany.
Neuroblastoma is one of the most frequent neuronal malignancy, affects the young
children’s worldwide. Highly heterogeneous and most of these tumors progressed to
metastasis through multiple mechanisms [1]. Methylglyoxal is a side product of
glycolysis, their influence on neuroblastoma sialylation and metastasis, is not yet
investigated in detail [2,3,4].
Neuroblastoma cell lines were treated with methylglyoxal at micromolar concentrations.
Significant increase in advanced glycation end products (AGES) and its receptor RAGE
on treatment with higher concentration of methylglyoxal. Increased phosphorylated
nuclear factor kappa B (NFkB) and extracellular signal–regulated kinases (ERK) were
also observed. This led to strong influence of altered sialic acid expression. Further
more the proliferation and migration properties of these cells were also increased on
methylglyoxal treatment. This results illustrates, the altered role of methylglyoxal in
enhancing metastatic property through modulation of sialylation.
References
1. Aminzadeh S., et al. (2015) Energy metabolism in neuroblastoma and Wilms tumor. 2. Gnanapragassam VS., et al. (2014). Sialic Acid Metabolic Engineering: A Potential Strategy for the Neuroblastoma Therapy. PLOS ONE 11(4): e0154289. doi: 10.1371/journal.pone.0154289 . 3. Allaman I., et al. (2015). " Methylglyoxal, the dark side of glycolysis." Front Neurosci 9: 23. doi: 10.3389/fnins.2015.00023. 4.Sullivan LB., et al. (2016). Altered metabolite levels in cancer: implications for tumor biology and cancer therapy." Nat Rev Cancer. 16(11):680-693.
73
(15) The anti-apoptotic properties of APEX1 in the endothelium
require the first twenty amino acids and converge on Thioredoxin-1
Christine Goy, Philipp Jakobs, Karin Aufenvenne, Yogi Altschmied, Jojo Haendeler
Heisenberg group - Environmentally induced Cardiovascular Degeneration, Clinical Chemistry, University of Duesseldorf,
and IUF-Leibniz Research Institute for Environmental Medicine, Duesseldorf, Germany
Introduction: The APEX nuclease (multifunctional DNA repair enzyme) 1 (APEX1) has a
disordered N-terminus, a redox and a DNA repair domain. APEX1 has anti-apoptotic
properties, which have been linked to both domains depending on cell type and experimental
conditions.
Objective: As protection against apoptosis is a hallmark of vessel integrity, we wanted to
elucidate whether APEX1 acts anti-apoptotic in primary human endothelial cells and if, what
the underlying mechanisms are.
Material and Methods: Domain mapping of APEX1. Transient transfection of endothelial cells
with Superfect (Qiagen). Apoptosis measurements were performed with Annexin V,
Caspase3/7 ELISA and immunoblots for Caspase 3 cleavage. Lentiviral transduction of
endothelial cells with APEX(1-20) and Trx-1 were performed. Carotid ligation of mice and
immunostaining of aortas were performed.
Results: APEX1 inhibits apoptosis in endothelial cells by reducing Cathepsin D cleavage,
potentially by binding to the unprocessed form. Diminished Cathepsin D activation results in
increased Thioredoxin-1 protein levels leading to reduced Caspase 3 activation. Consequently
apoptosis rates are decreased. This depends on the first twenty amino acids in APEX1,
because APEX1 (21-318) induces Cathepsin D activity, decreases Thioredoxin-1 protein
levels and thus, increases Caspase 3 activity and apoptosis. Along the same lines, APEX1 (1-
20) inhibits Caspase 3 cleavage and apoptosis. Furthermore, re-expression of Thioredoxin-1
via lentiviral transduction rescues endothelial cells from APEX1 (21-318)-induced apoptosis. In
an in vivo model of restenosis, which is characterized by oxidative stress, endothelial
activation and smooth muscle cell proliferation, Thioredoxin-1 protein levels are reduced in the
endothelium of the carotids.
Conclusion: APEX1 acts anti-apoptotic in endothelial cells. This anti-apoptotic effect depends
on the first twenty amino acids of APEX1. As proper function of the endothelium during life
span is a hallmark for individual health span, a detailed characterization of the functions of the
APEX1 N-terminus is required to understand all its cellular properties.
74
(16) Establishment of a blood-brain barrier model from patient-derived
induced pluripotent stem cells for the analysis of Alzheimer’s disease
Carla Hartmann, Matthias Jung, Toni Ehrhardt, Ina Giegling, Dan Rujescu, Department
of Psychiatry, Psychotherapy, and Psychosomatic Medicine, Martin Luther University
Halle-Wittenberg, Halle/Saale, Germany
aCenter for Molecular Biomedicine, Department of Biophysics, Friedrich Schiller University Jena & Jena University
Hospital, Hans-Knöll-Str. 2, D-07745 Jena, Germany bDepartment of Physiology, University of Pennsylvania, Philadelphia, USA
Dorsal root ganglia (DRG) neurons are important relay stations between the periphery
and the central nervous system and are essential for somatosensory signaling. Reactive
species are produced in a variety of physiological and pathophysiological conditions and
are known to alter electric signaling. Here we studied the influence of reactive species
on the electrical properties of DRG neurons from mice with the whole-cell patch-clamp
method. Even mild stress induced by either low concentrations of chloramine-T (10 µM)
or low-intensity blue-light irradiation profoundly diminished action potential frequency
but prolonged single action potentials in wild-type neurons. The impact on evoked
action potentials was much smaller in neurons deficient of the tetrodotoxin (TTX)-
resistant voltage-gated sodium channel NaV1.8 (NaV1.8–/–), the channel most important
for the action potential upstroke in DRG neurons. Low concentrations of chloramine-T
caused a significant reduction of NaV1.8 peak current and progressively slowed down
inactivation at higher concentrations. Blue light had a smaller effect on amplitude but
slowed down NaV1.8 channel inactivation. The observed effects were less apparent for
TTX-sensitive NaV channels, rendering NaV1.8 as an important reactive-species-
sensitive component in the electrical signaling of DRG neurons, potentially giving rise to
loss-of-function and gain-of-function phenomena depending on the type of reactive
species and their effective concentration and time of exposure. The mechanisms
described here might be relevant for alterations in rapid electrical signaling observed in
aged tissue or under age-related pathophysiological conditions.
75
(17) Delirium after cardiac surgery: non-invasive predictive parameters to identify patients at risk
Britt Hofmann1; Friederike Fee Barthel1; Pauline Eberlein1; Rüdiger Horstkorte2;
Andreas Wienke3; Hendrik Treede1; Andreas Simm
1Department of Cardiac Surgery, University Hospital Halle, Germany
2 Institute for Physiological Chemistry, Martin-Luther-University Halle-Wittenberg, Germany
3Institute of Medical Epidemiology, Biostatistics and Informatics, Martin-Luther-University Halle-Wittenberg, Germany
OBJECTIVES: Delirium after cardiac surgery is a problem with seriously consequences for
patients and healthcare. Preventive strategies from preoperative predictive non-invasive
assessments may reduce the incidence and severity of delirium. The present aim was to
explore preoperative non-invasive parameters for delirium in older patients undergoing cardiac
surgery with cardiopulmonary bypass.
METHODS: Patients (68.7 ± 9.3 years) scheduled for routine cardiac surgery were included (n
= 111). All patients were assessed pre-, peri- and postoperatively, and delirium was determined
using the confusion assessment method and the intensive care delirium screening checklist.
General cognitive and adaptive function was pre- and postoperatively evaluated with the clock
drawing test. The non-invasive parameters hand grip test (HGT) and skin autofluorescence
(sAF) were preoperatively recorded. All variables were analysed by uni-/multivariable logistic
regression.
RESULTS: Delirium was diagnosed in 24 patients (21.6%). Furthermore, univariable regression
analysis identified age (OR=1.09; 95%CI: 1.03-1.16; p=0.005), sAF (OR=3.9; 95%CI: 1.56-9.76;
p=0.004), non-dominant HGT (OR=0.93; 95%CI: 0.89-0.98; p=0.007) and dominant HGT
(OR=0.94; 95%CI: 0.89-0.98; p=0.005) as predictors of postoperative delirium. Finally, delirium
was independently associated with tissue glycation reflected by sAF (OR=2.82; 95%CI: 1.06-
7.49; p=0.038).
CONCLUSIONS: This study demonstrated for the first time a relationship between
postoperative delirium and the tissue glycation related skin autofluorescence. In addition,
postoperative delirium was associated with age and hand grip strength of the assessed cardiac
surgery patients.
76
(18) Detection of free Advanced Glycation End Products in vivo
during Hemodialysis
C. Hohmann, Halle (Saale)/D, C.Henning, Halle (Saale)/D, K. Liehr, Halle (Saale)/D, Prof. Dr.
M. A. Glomb, Halle (Saale)/D
Prof. Dr. M. A. Glomb, MLU Halle-Wittenberg, Institute of chemistry – Food chemistry Kurt-Mothes Straße 2, 06120 Halle (Saale), D, marcus.glomb@chemie.uni-halle.de
Patients suffering from uremia (advanced chronic kidney failure) have to undergo hemodialysis on a
regular basis to eliminate catabolic products. Uremia is understood as a status of chronic inflammation
linked to oxidative and carbonyl stress on the patients’ organism1.
Numerous studies have documented accumulation of advanced glycation end products (AGEs) in the
bloodstream of such patients2. AGE free adducts can originate from proteolysis, direct modification of
free amino acids or digestion of Maillard modified proteins ingested with the daily diet.
The amount of AGE free adducts in blood plasma and dialysis fluid is therefore an applicable parameter
to characterize the inflammatory status of uremic patients.
A highly selective LC-MS/MS-method was used for the analysis of AGE levels both in plasma and dialysis
fluid. A broad range of 19 amino acid modifications could be identified and quantitated.
Due to their low molecular weight between 174,1 Da (N6-formyl lysine) and 428 Da (glucosepane) it is
expected that the free amino acid modifications are successfully eliminated by the dialysis procedure
using membranes with a typical 35-45 kDa molecular weight cut-off. Indeed, this assumption proved to be
valid for all but one of the target analytes, with a mean clearance rate of 75%. Only pyrraline as the single
exception showed an opposite behavior, increasing by 59% during the treatment.
To find an explanation for this phenomenon, in-depth studies were conducted. De novo formation in blood
after the intake of the corresponding precursor compound 3-deoxyglucosone with the dialysis fluid or
catalytic effects on formation by the dialysis membrane were ruled out. To determine the role of
exogenous sources like glycated food proteins, a controlled dietary study with 8 dialysis patients was
carried out. In the first of two sessions, the patients were offered bread rolls with a mean pyrraline content
of 25.2 μmol/serving. In the second session the patients were asked to eat solely before the treatment. As
a result, the food intake during dialysis was directly correlated to an increase in plasma and in dialysate
verifying the exceptionally high bioavailability of food born pyrraline3.
Keywords: dialysis treatment, blood plasma, dialysate, free AGEs
References (1) Miyata, T.; Wada, Y.; Cai, Z.; Iida, Y.; Horie, K.; Yasuda, Y.; Maeda, K.; Kurokawa, K.; van Ypersele de Strihou, C. Implication of an increased oxidative stress in the formation of advanced glycation end products in patients with end-stage renal failure. Kidney Int. 1997, 51, 1170–1181. (2) Agalou, S.; Ahmed, N.; Thornalley, P. J.; Dawnay, A. Advanced glycation end product free adducts are cleared by dialysis. Ann. N. Y. Acad. Sci. 2005, 1043, 734–739. (3) Hohmann, C.; Liehr, K.; Henning, C.; Fiedler, R.; Girndt, M.; Gebert, M.; Hulko, M.; Storr, M.; Glomb, M. A. Detection of Free
Advanced Glycation End Products in Vivo during Hemodialysis. J. Agric. Food Chem. 2017, acs.jafc.6b05013.
77
(19) AGEs in hematopoietic development
Nicolas Huber, Karl Lenhard Rudolph
Advanced glycation end products (AGEs) are posttranslational protein modifications that
emerge from glycation, a non-enzymatic reaction between the protein and a sugar.
AGEs are reported to have negative effects on proteins, cells, tissues and organisms
and are linked to several age-related diseases such as Glycation is a non-enzymatic
reaction between a reducing sugar and a protein, which leads to the formation of
advanced glycation end products (AGEs). These posttranslational modifications are
often seen as protein damage as they can lead to aggregation, degradation or loss of
function. AGEs are linked to several age-related pathologies such as Alzheimer’s
disease, diabetes, inflammation signaling and cardiovascular diseases among others.
We investigate AGEs in the murine hematopoietic system, a tissue strongly relying on
stem cell function and differentiation which is compromised in older individuals. We find
that throughout hematopoietic development, populations show different AGE levels,
depending on their lineage and differentiation stage. We find these population-
dependent levels to be unchanged in old age and in dietary restriction, which indicates
that these levels are conserved and potentially inevitable or necessary. The differences
in AGE levels point towards a possible relationship between AGEs and differentiation
within the hematopoietic system. The question to be answered is if differentiation drives
AGE levels or if AGEs could actually contribute to differentiation. This would argue for
an important role of AGEs and could contribute to rethink the dogma that renders AGEs
only harmful to the organism.
Though focused on aging, the field of AGE research neither concentrates on stem cells
nor the hematopoietic system, thereby neglecting their importance. Our studies serve to
gain a better understanding of the biological role of glycation during aging and in stem
cells, starting with the hematopoietic system.
78
(20) Non-enzymatic modifications of the mineralocorticoid receptor during vascular aging
Ralf Hübschmann, Stefanie Ruhs, Claudia Großmann
Julius-Bernstein-Institut für Physiologie Halle
Numerous clinical studies showed that the mineralocorticoid receptor (MR), a ligand-
dependent transcription factor, mediates pathophysiological processes like
inflammation, fibrosis and endothelial dysfunctions in the cardiovascular system. An
important aspect of the deleterious MR effects is that the receptor and its natural ligand
aldosterone per se are not capable of eliciting pathophysiological effects but requires a
certain micro-milieu. Several studies indicate that a parainflammatory micro-milieu
created by a shift in redox state and/or altered level of oxidative and nitrosative stress
(ROS/RNS) represents a possible trigger to induce pathophysiological MR effects.
During ageing, vascular cells are increasingly exposed to such a parainflammatory
micro-milieu which then augments pathophysiological MR signaling, leading to age-
associated inflammatory processes (vasculopathy). Our experiments show that
nitrosative stress influences genomic MR activity. Whereas nitric oxide (NO) attenuates
aldosterone-induced transcriptional MR activity by reducing its binding to DNA
elements, peroxynitrite (ONOO–) acts as a ligand-independent MR activator. When
exploring the underlying molecular mechanism, we found that activation of the MR leads
to posttranslational modification of the receptor and that PTM of the MR may also occur
during conditions involving enhanced nitrosative stress. Co-immunoprecipitation and
Western blot experiments show an increased S-nitrosylation level of the MR after NO
treatment whereas ONOO– stimulation leads to an elevated amount of nitrated MR
tyrosines. In silico predicted modified cysteine and tyrosine residues will be validated by
mass spectrometry and then characterized by performing site-directed mutagenesis
experiments followed by functional analyses of mutated MR compared to wild type
receptor. Overall, these experiments will help to understand pathophysiological MR
signaling during nitrosative stress conditions which occur during ageing.
79
(21) Stem cells modelling Alzheimer's disease for the analysis of molecular aging of the brain
Matthias Jung, Toni Ehrhardt, Carla Hartmann, Jovita Schiller, Ina Giegling, and Dan Rujescu
Department of Psychiatry, Psychotherapy, and Psychosomatic Medicine, Martin Luther University Halle-Wittenberg
Aim: Alzheimer’s disease (AD) is the most common form of dementia caused by environmental and
genetic factors. The majority of patients are suffering from late onset AD in persons older than
65 years. One characteristic of AD is the accumulation of amyloid-β (Aβ) in the brain. Recently,
19 loci in the DNA of AD patients were associated with AD including CLU. CLU has been
described as a potential regulator of brain function during aging. To investigate molecular aging
mechanisms in AD, we established stem cell-derived cerebral in vitro models for the patient-
specific analysis of psychiatric genetics.
Methods: Episomal vectors were used for the generation of induced pluripotent stem (iPS) cell lines from
patient-specific B-lymphoblastoid cell lines (B-LCLs). Pluripotency was verified and a screening
protocol ensured a high neuronal differentiation potential towards neural and glial lineages.
Cells were also applied to generate self-renewing neural stem cells (NSCs) suitable to generate
neurons and astrocytes in 2D and 3D models.
Results Permanent iPS cell lines and NSCs were characterized by morphology and the presence of
stem cell-specific marker genes including OCT4 and NANOG in iPS cells and SOX2 and PAX6
in NSCs. Stemness markers were analyzed whether they represent appropriate aging markers.
NSCs were successfully differentiated into neurons and astrocytes. Astrocyte-specific markers
including EAAT1 demonstrated efficient astrocyte differentiation. The efficient differentiation of
mature neurons was shown by the presence of glutamate and GABA receptors. Mitochondrial
respiration was elevated in differentiated neurons. Patch clamp analysis verified the functionality
of GABAergic and glutamatergic neurons and astrocytes. 3D organoids obtained from NSCs
showed the expression of cortical markers such as TBR1 mimicking the developing human
cortex. Cells obtained from young and old patients showed different properties.
Together, stem cells modelling AD represent a powerful tool to study molecular aging of the
brain.
80
(22) Are AGE-modified amino acids proteinogenic?
Astrid Junk1,2, Anne Kummer1, Patrick Winterhalter1,2, Tim Baldensberger2,3, Marcus
Glomb2,3, Andreas Simm1,2
1Clinic of Heart Surgery, Martin-Luther-University Halle-Wittenberg, Halle (Saale),
2ProMoAge, GRK 2155,
3Institute of food chemistry, Martin-Luther-University Halle-Wittenberg, Halle (Saale)
The characteristic western diet contains heated and highly processed proteins. These
processes lead to various artificial modifications e.g the formation of Advanced
Glycation Endproducts (AGEs). The reaction is defined as a non-enzymatic addition of
reactive carbohydrates e.g. sugars to amino acids. The endogenous formation of AGEs
is a part of the normal ageing metabolism. High levels of AGEs in tissues and body
fluids can become pathogenic due to their ability to promote oxidative stress and
inflammation. Current research could show, that only a part of the absorbed AGEs are
excreted via the kidneys after 48 hours, the fate of the non-excreted AGEs remains
undetermined.
Within this project, the reuse of single AGE-modified amino acids, e.g.
Carboxymethyllysine (CML), in protein translation is tested.
In an in vitro translation approach based on Hela lysates and a GFP-expression vector,
the addition of CML induce an increase influorescence without any change in protein
amount of the expressed target protein. Control experiments exclude effects of pH,
charge and molarity as a cause of modified fluorescence. Furthermore, due to a total
amino acid analysis of the produced proteins, incorporation of CML could be detected.
Thus, this first results indicates hint at a low rate of translational incorporation of CML.
The next step will be a mass spectrometric analysis to clarify which amino acids are
substituted. In addition, cells will be fed with CML-containing-dipeptides to show a
possible impact on the in-vivo translation.
Our data indicate on a possible role of nutrition related AGEs on in-vivo aging by
incorporation into newly synthesized proteins within translation.
81
(23) Two putative selenium binding proteins as modulators of C.
elegans stress response and life span
Karl Köhnlein1, Nadine Urban1, Holger Steinbrenner1, Christoph Kaether2, Lars-Oliver
Klotz1
1Institute of Nutrition, Department of Nutrigenomics, Friedrich-Schiller-Universität Jena, Germany
2Leibniz Institut für Alternsforschung-Fritz Lipmann Institut, Jena, Germany
Selenium-binding proteins do not contain selenium in the form of selenocysteine or
selenomethionine but as inorganic selenium bound to the protein. The C. elegans
genome encodes only one selenocysteine-containing selenoprotein, TrxR-1. However,
at least two ORFs encode putative selenium-binding proteins, CeSELENBP1 and
CeSELENBP2. These are 54% and 36% homologous to human selenium-binding
protein-1 (SELENBP1), respectively, and both contain a cysteine residue hypothesized
to bind selenite as in SELENBP1.
Considering the role of selenium in antioxidant defense, we hypothesized that
CeSELENBP1 and CeSELENBP2 may modulate the response of C. elegans to
oxidative stress. Unexpectedly, life-long knock-down of either of the genes significantly
increased life span and stress resistance to the redox cycler paraquat.
DAF-16 and SKN-1 are key players in the C. elegans stress response that are known to
mediate many of the published life-extending effects of chemical compounds,
phytochemicals and nutrition regimens. However, RNAi against CeSELENBP1 and
CeSELENBP2 significantly increased life span of the above mutants to an extent similar
to that found in wild type worms exposed to RNAi targeting either of the putative
selenium binding proteins. This suggests that neither DAF-16 nor SKN-1 are involved in
life span extension elicited by knock-down of CeSELENBP1 and CeSELENBP2.
82
(24) A genetically encoded indicator of methionine oxidation
Kuldyushev N.A., Schönherr R., Heinemann S.H.
Center for Molecular Biomedicine, Department of Biophysics, Friedrich Schiller University Jena and Jena University Hospital, Hans-Knöll-Str. 2, 07745 Jena, Germany nikita.kuldyushev@med.uni-jena.de, stefan.h.heinemann@uni-jena.de
Post-translation modifications are essential in the fine-tuning of protein signaling
networks. During aging, the levels of some post-translational modifications are
changing, thereby affecting the functions of modified proteins. Methionine oxidation
to methionine sulfoxide (Met(O)) is a post-translational modification that can appear
unspecifically during an exposure of methionine to oxidative stress, such as
occurring in aged organisms and in a number of degenerative diseases and
pathologies. The importance of methionine oxidation is further emphasized by the
fact that virtually all organisms harbor a set of enzymes – methionine sulfoxide
reductases (MSRs) – that specifically reduce free and protein-based methionine
sulfoxide back to methionine.
Here we introduce a genetically encoded fluorescent sensor for methionine
oxidation. Variants based on green fluorescent protein (GFP) were designed to
provide a ratiometric fluorescence signal reporting on the degree of methionine
oxidation. Such fluorescent proteins were produced in a bacterial expression
system and their properties were studied under in vitro conditions in a cuvette.
Experimental oxidation with H2O2, chloramine T, or hypochloric acid resulted in
robust changes in the fluorescent ratio F400/F470. Incubation with recombinantly
produced MSRs indicated that the sensor is not a substrate for the enzymes. Upon
expression in mammalian cells (HEK293T), the sensor produced ratiometric
fluorescence signals suited for single-cell photometry and life-cell imaging to
monitor intracellular methionine oxidation events. This study will contribute to a
better understanding of how oxidative modification levels change during aging and
disease states.
83
(25) Potential of glycated beta-Amyloid as an early diagnostic marker
for Alzheimer's disease
Anne Kummer1, Alexander Navarrete Santos2, Hendrik Treede1, Andreas Simm1,2
1 Clinic for Heart Surgery, Martin-Luther-University Halle-Wittenberg, Halle (Saale)
2 Center for Medical Basic Research, Martin-Luther University Halle-Wittenberg, Halle (Saale)
Alzheimer’s disease (AD) is an age-related, neurodegenerative disorder that poses a
growing challenge especially due to the ongoing demographic transition. Furthermore,
AD is not curable and medication is only capable of delaying or slowing the disease
progression. Hence, an early diagnosis is of utmost importance.
One patho-mechanism of Diabetes, a major risk factor for AD, is the carbohydrate-
induced glycation of proteins, forming long-lasting and non-reversible advanced
glycation end products (AGEs). Beta-Amyloid (Abeta), a polypeptide implicated in AD,
exhibits increased cytotoxicity in its glycated form. Thus, this project aims to clarify
whether glycated, soluble Abeta is a suitable early marker for the diagnosis of AD.
For this purpose, an existing Abeta-based, flow cytometric assay was modified to detect
glycated forms of the soluble polypeptide in cerebrospinal fluid (CSF). In parallel, a
recombinant standard was established to also facilitate the quantification of the modified
Abeta via flow cytometry. In principle, the carboxymethyl-lysine containing, recombinant
standard of Abeta can be detected by this assay. Currently, the reproducibility is
assessed for this assay and alternative detection methods of glycated Abeta are
considered.
Furthermore, immunoprecipitation of Abeta and subsequent mass spectrometric
analyses will be utilized to identify the glycated Abeta-forms predominantly present in
CSF of AD-patients in comparison to healthy controls.
The ultimate goal, however, is to establish a diagnostic assay for the early diagnosis of
Alzheimer’s based on the obtained results.
84
(26) Assessment of murine gut motility under stress conditions
Johanna Langner
Gut failure and the development of a megacolon emerging from impaired gut motility are
significant pathological factors in severe sepsis. Furthermore, imbalance of the gut
microbiome, such as excessive C. difficile growth following antibiotics treatment, can
cause life-threatening complications. In this project molecular mechanisms underlying
gut dysmotility, which can lead to gut failure and even death, are to be investigated.
Gut motility strongly relies on rhythmic depolarization of the smooth muscle cells
(SMCs), which at least in part are triggered by specialized SMCs with pacemaking
function: Interstitial Cells of Cajal (ICC-My). Besides measuring the mechanical force
exerted by the smooth muscle layer, the membrane potential (Vm), the intracellular Ca2+
concentration ([Ca2+]i), or the intracellular redox potential are potential parameters for
the characterization of gut motile activity. To measure fluctuations of these parameters
in SMCs and ICC-My, genetically encoded fluorescence sensors reporting on Vm,
[Ca2+]i, or the formation of disulfide bonds are developed and examined.
Electrical rhythm generation requires the concerted activity of ion channels, which may
be relevant targets for stress factors affecting gut motility. Here we concentrate on K+
and Cl– conductances that are important regulators of the electrical activity in ICC-My.
In particular we examine how the Ca2+-activated Cl– channel TMEM16A operates in
ICC-My under physiological conditions and after exposure to oxidative stress (e.g.
reactive species, heme, Fe2+) and low molecular weight compounds which are released
from the microbiome under control and disease conditions.
85
(27) The Gid/CTLH Ubiquitin Ligase Complex: A Novel Cilia-
Associated Regulator of Autophagy
Huaize Liu1,6, Jie Ding1, Pablo Villavicencio-Lorini2, Peter Walentek4, Gerd Hause5,
Thomas Hollemann1, Thorsten Pfirrmann1,6,7
1 Martin-Luther University Halle-Wittenberg, Institute of Physiological Chemistry, Halle, Germany
2 Martin-Luther University Halle-Wittenberg, Institute of Human Genetics, Halle, Germany
4 University of California at Berkeley, Division of Genetics, Genomics and Development, Centre for Integrative
Genomics, Berkeley, USA 6 These authors contributed equally to the work
7 Address correspondence to: Thorsten.pfirrmann@medizin.uni-halle.de
Keywords: CTLH complex / ubiquitin/ ciliogenesis/ ciliopathies/ Xenopus laevis/ autophagy/ ageing
Pharmacological or genetic manipulations that increase life span in model organisms
often stimulate autophagy by the specific inhibition of the mTOR pathway. Cilia are
highly evolutionary conserved organelles involved in embryonic development, sensing
of extracellular milieu, and a variety of signal transduction pathways. Recent studies
propose a dual-interplay between cilia function and autophagy that regulates metabolic
homeostasis, e.g. elongated primary cilia have a reduced mTOR activity and an
increased autophagic flux. Here, we describe the function of an evolutionary conserved,
cilia associated ubiquitin ligase complex as a novel modulator of mTOR signaling and
autophagy. In Saccharomyces cerevisiae, the Gid/CTLH complex regulates the
metabolic switch from gluconeogenesis to glycolysis by targeting key enzymes of
gluconeogenesis for proteasomal degradation. However, its function and substrates in
vertebrates remains enigmatic. We provide evidence that the Gid/CTLH complex has a
function during the formation of both motile and primary cilia. The subunit Rmnd5a co-
localizes with the basal body of primary cilia in NIH3T3 cells, and Rmnd5a loss of
function leads to the elongation of primary cilia. Interestingly, this correlates with an
increase in autophagic flux and attenuated mTOR activity. Together, our data suggests
an intimate function of cilia-associated regulation of autophagy and metabolic
homeostasis that most likely is mediated by an unknown substrate of the Gid/CTLH
complex. We hypothesize that the primary cilium is a nutrient sensor that regulates
metabolic homeostasis and nutrient signaling via the Gid/CTLH complex. Thus the
Gid/CTLH complex emerges as a promising new target to pharmacological alter mTOR
activity and its known effects on longevity.
86
(28) Age related glycation hotspots in plant proteome
Tatiana Bilova,1,2 Tatiana Mamontova,1,3 Gagan Paudel,1,4 Dominic Brauch,4 Svetlana Milrud,2,3
Rico Schmidt,5 Elena Tarakhovskaya,2 Nikita Shilyaev,3 Galina Smolikova,2 Alain Tissier,6
Thomas Vogt,6 Claudia Birkemeyer,4 Andrea Sinz,5 Wolfgang Brandt,1 Ludger A. Wessjohann,1
and Andrej Frolov,1
1Department of Bioorganic Chemistry, Leibniz Institute of Plant Biochemistry (Halle/Saale),
2Departments of Plant
Physiology and Biochemistry and 3
Biochemistry, St. Petersburg State University (Russia), 4Faculty of Chemistry and
Mineralogy, Universität Leipzig,
5Institute of Pharmacy, Martin-Luther Universität Halle-Wittenberg and
6Department
of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry (Halle/Saale).
Advanced glycation is referred to as a non-enzymatic reaction of proteins with α-dicarbonyl
intermediates of sugar autoxidation, glycoxidation and lipid peroxidation. It is a factor of protein
damage resulting in accumulation of advanced glycation end products (AGEs) in mammalian
tissues. In this context, AGEs are well-known markers of ageing. In comparison to mammals,
plants are characterized with higher tissue contents of reactive carbohydrates and increased
rates of oxidative processes. This might result in enhanced carbonyl stress, and, hence, higher,
in comparison to mammals, levels of protein glycation. Indeed, recently, rich constitutive plant
glycation patterns were characterized by our group [1]. However the susceptibility of plant
proteins to age-related glycation was not addressed so far. Therefore, here we consider this
problem in the context of possible mechanisms underlying age-related glycation using 7-12-
week-old A. thaliana plants and methods of biochemistry, molecular biology, metabolomics and
proteomics. Although qualitative glycation patterns were relatively stable in time, multiple
proteins demonstrated strong increase of glycation (up to 28 fold) by specific sites, i.e. glycation
hotspots were observed [2]. Many up-glycated proteins were involved in protein metabolism and
transcriptional regulation. Four of them contained AGE-modifications in their functional domains
that could affect protein functionality. The identified glycation hotspots dominated with arginine-
derived hydroimidazolones, originating from glyoxal (Glarg) and methylglyoxal (MG-H). Analysis
of glycation-specific sequence motifs revealed ionic (K, R, E, D) and non-ionic (L,V, A, P)
residues in close proximity to the glycation hotspots. Homology modeling of glycated proteins
revealed glutamyl and aspartyl residues in close proximity to the hotspots (at the distance less
than 5Å), that might indicate their enhancement effect on AGE formation.
References
1. Bilova T, Lukasheva E, Brauch D, Greifenhagen U, Paudel G, Tarakhovskaya E, Frolova N, Mittasch J, Balcke GU, Tissier A, Osmolovskaya N, Vogt T, Wessjohann LA, Birkemeyer C, Milkowski C, Frolov A (2016). A Snapshot of the Plant Glycated Proteome: Structural, Functional and Mechanistic Aspects. J. Biol. Chem., 291(14):7621-36.
2. Bilova T., Paudel G., Shilyaev N., Schmidt R., Brauch D., Tarakhovskaya E., Milrud S., Smolikova G., Tissier A.,
Vogt T., Sinz A., Brandt W., Birkemeyer C., Wessjohann L. A., Frolov A. (2017). Global Proteomic Analysis of
Advanced Glycation End Products in the Arabidopsis Proteome Provides Evidence for Age-related Glycation Hotspots. J. Biol. Chem., 2017 (First Published on June 13, 2017, accepted jbc.M117.794537. doi:10.1074/jbc.M117.794537)
87
(29) Advanced Glycation End Products, oxidative stress and Glyoxalase expression in breast cancer
Norbert Naß*1, Ludwig Andreas1, Atanas Ignatov2, Johannes Haybäck1 and Thomas
Kalinski1
1: Otto von Guericke University Magdeburg, Department of Pathology, Leipziger Str. 44, D-39120 Magdeburg, Germany; 2: Otto von Guericke University Magdeburg, Department of Obstetrics and Gynecology, Gerhart-Hauptmann Str. 35, 39108 Magdeburg, Germany * Contact: Norbert.nass@med.ovgu.de
Proteins modified by Advanced Glycation End products (AGEs) are well documented
biomarkers for ageing and several degenerative diseases. AGE-modification has
significant effects on the stability of the extracellular matrix thus contributing to age-
related stiffening of organs. Signaling molecules and enzymes can be inactivated by
glycation and AGEs are recognized by the receptor for AGEs (RAGE), which leads to
an activation of the NF-κB transcription factor. Due to metabolic changes, especially the
Warburg effect and oxidative stress, tumors often accumulate significant amounts of
AGE-modified proteins. Thus, cancer cells need to detoxify glycating aldehydes by
means of glyoxalases, namely glyoxalase-1 (GLO1) and DJ-1 (PARK7).
We here investigated the contribution of the AGE-system to breast cancer biology and
prognosis. The accumulation of the AGE carboxymethyl-lysine (CML) was
retrospectively investigated by immunohistochemistry in a cohort of about 200 breast
cancer patients. High CML-accumulation correlated with estrogen receptor expression
and age and was related to unfavorable prognosis in estrogen receptor negative
tumors. In cell culture experiments, GLO1 turned out to be important to confer α-oxo-
aldehyde resistance. Also, the toxicity of α-oxo-aldehydes correlated with adaption of
estrogen receptor positive cell lines to the selective estrogen receptor mediator
tamoxifen and such cells showed an altered regulation of NF-kB signaling.
Altogether these data suggest an important role of the AGE-system for breast cancer
biology. Especially glyoxalases might represent effective targets for the treatment of this
disease.
88
(30) Dicarbonyls induce senescence of human vascular endothelial
cells
Alexander Navarrete Santos§, Kathleen Jacobs#, Andreas Simm#§, Nicole Glaubitz#;
Rüdiger Horstkorte*, Britt Hofmann#
§Centre for Medical Basic Research of the Martin-Luther-University Halle-Wittenberg, Germany
#Department of Cardiac Surgery, University Hospital Halle, Germany
*Institute for Physiological Chemistry, Martin-Luther-University Halle-Wittenberg, Germany
Rationale: Glyoxal (GO) and Methylglyoxal (MGO) are two dicarbonyls involved in the
formation of advanced glycation end products (AGEs). Endothelial cells in the vessels
are in constant contact with circulating AGEs and dicarbonyls. With this project, we
aimed to elucidate the effect of GO and MGO on primary human vascular endothelial
cells (HVECs).
Methods: Graft material from patients with coronary heart disease was used as HVECs
source. HVECs were treated with different concentrations of GO and MGO. -
Galactosidase related senescence activity and cell morphology were analyzed. AGEs
as well as p21 protein expression, glyoxalase-I expression and oxidative stress were
detected.
Results: We here provide evidences that GO and MGO induce senescence in primary
HVECs. Mechanistically GO and MGO induce senescence by increasing the ROS
production, the expression of p21, the accumulation of AGEs and the arrest of HVECs
in the G2 cell cycle phase. Aminoguanidine- a dicarbonyl scavenger- abrogated the
effect of GO and MGO.
Conclusion: Our data are relevant as they suggest that in diseases with elevated
dicarbonyl concentrations, deleterious effects on the endothelium and the development
of vascular dysfunction must be expected. On the other hand, treatment of patients with
dicarbonyl scavenger could prevent this.
89
(31) Age and diet affect the intestinal crypt proteome
Nadja Gebert1, Joanna Kirkpatrick1, Jerome Korzelius1, Henri Jasper1,2 and
Alessandro Ori1
1 Leibniz Institute on Aging - Fritz Lipmann Institute (FLI), Jena, Germany
2 Buck Institute for Research on Aging, Novato, USA
The small intestine is responsible for nutrient sensing and absorption, and it is one of
the most important interfaces between the environment and our body. During
aging, deregulation of intestinal stem cell (ISC) activity leads to loss of epithelial barrier
function, food malabsorption and dysbiosis of commensal bacteria.
In order to investigate both cell intrinsic and extrinsic factors influencing ISC aging, we
isolated crypts and stem cells from the small intestine of mice and D.
melanogaster midgut. We compared proteomic profiles of tissues and cells from
different age groups using state-of-the-art mass spectrometry. We found proteome
signatures in the intestinal epithelium, which indicate that aging affects metabolic
networks, stem cell proliferation, and epithelial immune responses. Of note, some of
these aging-associated alterations (e.g., immune modulatory responses) are reverted
by dietary restriction (DR), a health span extending intervention conserved across
species. In particular, we found that both aging and DR influence the expression of a
key enzyme in ketone body metabolism, and demonstrate that perturbation of its activity
as well as the exogenous supplementation of ketone bodies can influence ISC
regeneration in organoid cultures. Our data demonstrate how aging and dietary
intervention can modulate metabolic networks and influence stem cell activity by altering
the concentration of metabolites in the niche.
90
(32) Integrative analysis of age related changes in the transcriptome of
Caenorhabditis elegans
Tsimafei Padvitski, Adam Antebi, Andreas Beyer
To obtain insight into age-related changes of multicellular organism, we studied global
transcriptome of C. elegans under normal aging using RNAseq time-series experiment.
The data was obtained in RNAseq experiment on synchronized culture of wild-type
C.elegans sampled at day 0, 7, 14 and day 21 (all stages but the first are
postreproductive). Then we performed a gene-level differential expression analysis and
a subsequent functional annotation. The results support the theory that ageing is a
highly stochastic process. At the same time, they also show that specific processes and
pathways are more vulnerable to the stochastic changes.
Next, we used time-series clustering to obtain gene sets that represent transcriptome
modules with distinct temporal dynamics in aging. Finally, we questioned if these
transcriptome modules can be controlled by specific transcription factors. To answer
this question, we integrated public Chip-seq data from modENCODE with the results of
the time-series clustering using a simple statistical framework. We identified a
consistent set of TFs, all have been previously described as implicated in aging.
Interestingly, the same set of TFs can presumably control behavior of genes with
different dynamics during aging.
To summarize, we characterized in details age-related changes in the global
transcriptome of C. elegans and showed that integration of gene expression with Chip-
seq data allows the prediction of TFs that are capable of modulating the lifespan of C.
elegans.
91
(33) High risk of malnutrition is associated with low muscle mass in
older hospitalized patients - a prospective cohort study
Vincent D. Pierik BSc1, Dr. Carel G.M. Meskers2, Jeanine M. Van Ancum MSc1,3, Siger
T. Numans BSc1, Sjors Verlaan MSc1,4, Kira Scheerman MSc5, Dr. Roeliene C.
Kruizinga1, Prof. Dr. Andrea Maier3,6 *
1 Department of Internal Medicine, Section of Gerontology and Geriatrics, VU University Medical Center, Amsterdam, The
Netherlands
2 Department of Rehabilitation Medicine, VU University Medical Center, Amsterdam, The Netherlands
3 Department of Human Movement Sciences, MOVE Research Institute Amsterdam, VU University, Amsterdam, The
Netherlands
4 Nutricia Research, Nutricia Advanced Medical Nutrition, Utrecht, The Netherlands
5 Division I, VU University Medical Center, Amsterdam, The Netherlands
6 Department of Medicine and Aged Care, Royal Melbourne Hospital, University of Melbourne, Melbourne, Australia
Background: Malnutrition, low muscle strength and muscle mass are highly prevalent in older
hospitalized patients and associated with adverse outcomes. Malnutrition may be a risk factor for
developing low muscle mass. We aimed to investigate the association between the risk of
malnutrition and 1) muscle strength and muscle mass at admission and 2) the change of muscle
strength and muscle mass during hospitalization in older patients.
Methods: The EMPOWER study included 378 patients aged seventy years or older who were
acutely or electively admitted to four different wards of an academic teaching hospital in
Amsterdam. Patients were grouped into low risk of malnutrition and high risk of malnutrition
based on the Short Nutritional Assessment Questionnaire (SNAQ) score and were assessed for
hand grip strength and muscle mass using hand held dynamometry respectively bioelectrical
impedance analysis (BIA) within 48 hours after admission and at day seven, or earlier at the day
of discharge. Muscle mass was expressed as skeletal muscle mass, appendicular lean mass, fat
free mass and the skeletal muscle index.
Results: The mean age of the patients was 79.7 years (SD 6.39), 48.9 % were female. At
admission, being at high risk of malnutrition was significantly associated with lower muscle
mass (Odds Ratio, 95 % CI, 0.90, 0.85-0.96), but not with muscle strength. Muscle strength and
muscle mass did not change significantly during hospitalization in both groups.
Cconclusion: In older hospitalized patients, a high risk of malnutrition is associated with lower
muscle mass at admission, but not with muscle strength nor with change of either muscle
strength or muscle mass during hospitalization.
92
(34) Attempts to remove AGEs from human plasma using polysterene-
divinylbenzene polymer adsorbent
Veronika Piskovatska, Alexander Navarrete Santos, Andreas Simm
Martin-Luther-University Halle-Wittenberg, University Clinic for Cardiac Surgery
Background: Advanced glycation end products (AGEs), forming in vivo under
conditions when dicarbonyl compounds react with proteins, are shown to have a
significant role in ageing-associated diseases. Pharmacological agents, reducing AGEs
levels in plasma by inhibiting certain stages of glycation, demonstrated clinical benefits
in elderly patients with different conditions (metformin, aspirin, ACEi).
The aim of the project was to evaluate the binding capacity of CytoSorb adsorber,
currently approved for reduction of extremely elevated cytokine levels in intensive care
units patients. Adsorber efficiently binds proteins within 5-60 kDa range in
concentration-dependent manner.
Materials and methods: AGE-modified human plasma was put through centrifuge
columns, packed with CytoSorb material in buffer. Efficiency of AGEs binding was
compared in CytoSorb column in parallel with immobilized boronic acid resin column,
which is described to specifically bind AGEs. Glycated proteins from plasma samples
before and after treatments, washouts and elutions were analyzed by immunoblot.
Autofluorescence of AGEs at different wavelengths was evaluated via fluorescent plate-
reader.
Results: CytoSorb columns efficiently binds CML, pentosidine, MGH1, CEL and AGE-1
(glucose-modified) glycated proteins, qualitative immunoblot results demonstrated that
polymer-based column removes AGEs from plasma in comparable extent as the
boronic acid procedure. Column treatment decreased the autofluorescence of plasma
samples at 360/440 nm, corresponding to reduction of fluorescent AGEs. Repeated
plasma purification with polymer-filled column also increased the extent of AGEs
removal from the modified plasma samples.
Conclusion: the extent of binding, as well as specificity towards different kinds of AGEs
are yet to be identified, as well as specificity of the selected adsorbent system towards
variable protein posttranslational modifications of clinical relevance.
93
(35) Individual variance in resting metabolic rate and energy balance
in geriatric outpatients with mobility problems
Esmee M. Reijnierse1, Sjors Verlaan2, Marijke C. Trappenburg2,3, Carel G.M.
Meskers4,5, Marian A.E. de van der Schueren6,7, Andrea B. Maier1,5
Affiliations 1Department of Medicine and Aged Care, The Royal Melbourne Hospital, The University of Melbourne, Australia
2Department of Internal Medicine, Section of Gerontology and Geriatrics, VU University Medical Center, Amsterdam,
The Netherlands 3Department of Internal Medicine, Amstelland Hospital, Amstelveen, The Netherlands
4Department of Rehabilitation Medicine, VU University Medical Center, Amsterdam, The Netherlands
5Department of Human Movement Sciences, MOVE Research Institute Amsterdam, Vrije Universiteit, Amsterdam,
The Netherlands 6Department of Internal Medicine, Section of Nutrition and Dietetics, VU University Medical Center, Amsterdam, The
Netherlands 7Department of Nutrition and Health, Faculty of Health and Social Studies, HAN University of Applied Sciences
Nijmegen, The Netherlands
Rationale: In clinical practice, optimization of physical and nutritional intervention (“tailored
therapy”) requires patient specific data on energy expenditure and nutritional intake tailored to
the individual requirements; these data are generally lacking. This study aimed to explore the
nutritional needs of geriatric outpatients regarding energy expenditure and energy balance.
Methods: This inception cohort of older adults referred to the geriatric outpatient clinic of the
Center of Geriatric Medicine Amsterdam, VU University Medical Center included 26 outpatients.
Geriatric outpatients underwent standardized phenotyping based on regular care and additional
nutritional (indirect calorimetry to measure resting metabolic rate (RMR), three day food diary)
and physical activity assessments (accelerometry). Next to indirect calorimetry, RMR was
estimated using the equation by the World Health Organization based on age, sex, body weight
and height. Total energy expenditure (TEE) was calculated by RMR times the physical activity
level. Energy balance was defined as TEE versus the energy intake.
Results: A high individual variance was found in both RMR and energy balance. Mean
difference between the actual measured RMR and the estimated RMR was 234 kcal/day on a
population level. On the individual level, estimated RMR underestimated the actual measured
RMR by more than 10% in more than half of the outpatients. Mean difference between TEE and
energy intake was 429 kcal/day. Half of the outpatients had an energy consumption lower than
their needs by more than 10%.
Conclusions: RMR should be measured objectively due to the high underestimation of
estimating RMR. In geriatric outpatients, it is important to individually quantify the nutritional
needs.
94
(36) Lack of knowledge and availabiliy of diagnostic equipment hinder
the diagnosis of sarcopenia and its management
Esmee M. Reijnierse1, Marian A.E. de van der Schueren2,3, Marijke C. Trappenburg4,5,
Marjan Doves6, Carel G.M. Meskers7,8, Andrea B. Maier1,8
Affiliations 1Department of Medicine and Aged Care, University of Melbourne, Melbourne, Australia
2Department of Internal Medicine, Section of Nutrition and Dietetics, VU University Medical Center, Amsterdam, The
Netherlands 3Department of Nutrition and Health, Faculty of Health and Social Studies, HAN University of Applied Sciences,
Nijmegen, The Netherlands 4Department of Internal Medicine, Section of Gerontology and Geriatrics, VU University Medical Center, Amsterdam,
The Netherlands 5Department of Internal Medicine, Amstelland Hospital, Amstelveen, The Netherlands
6Institute of Human Movement Studies, Faculty of Health Care, University of Applied Sciences Utrecht, Utrecht, The
Netherlands 7Department of Rehabilitation Medicine, VU University Medical Center, The Netherlands
8Department of Human Movement Sciences, MOVE Research Institute, Vrije Universiteit, Amsterdam, The
Netherlands
Rationale: Sarcopenia is an emerging clinical challenge in an ageing population and is associated
with serious negative health outcomes. This study aimed to assess the current state of knowledge
about the concept of sarcopenia, diagnostic strategy and management in a cohort of Dutch
healthcare professionals (physicians, physiotherapists, dietitians and others) attending a lecture
cycle on sarcopenia.
Methods: Healthcare professionals (n=223) were asked to complete a questionnaire before, directly
after and five months after attending a lecture cycle on the pathophysiology of sarcopenia,
diagnostic strategy and management of sarcopenia, i.e. interventions and collaboration.
Results: Before attendance, 69.7% of healthcare professionals stated to know the concept of
sarcopenia and 21.4% indicated to know how to diagnose sarcopenia. Handgrip strength was the
most frequently used objective diagnostic measure (33.9%). Five months after attendance, reported
use of diagnostic tests was increased, i.e. handgrip strength up to 67.4%, gait speed up to 72.1%
and muscle mass up to 20.9%. Bottlenecks during implementation of the diagnostic strategy were
experienced by 67.1%; lack of awareness, acquisition of equipment and time constraints to perform
the diagnostic measures were reported most often. Before attendance, 36.4% stated not to consult a
physiotherapists or exercise therapists (PT/ET) or dietitian for sarcopenia interventions, 10.5%
consulted a PT/ET, 32.7% a dietitian and 20.5% both a PT/ET and dietitian. Five months after
attendance, these percentages were 28.3%, 21.7%, 30.0% and 20.0% respectively. A lack of
collaboration was experienced by 36.8%.
Conclusion: The concept of sarcopenia is familiar to most healthcare professionals but application
in practice is hampered, mostly by lack of knowledge, availability of equipment, time constraints and
lack of collaboration.
95
(37) Assessment of maximal handgrip strength: how many attempts
are needed?
Esmee M. Reijnierse1, Nynke de Jong2, Marijke C. Trappenburg2,3, Gerard J. Blauw4,5,
Gillian Butler-Browne6, Helena Gapeyeva7, Jean-Yves Hogrel6, Jamie S. McPhee8
Marco V. Narici9, Sarianna Sipilä10, Lauri Stenroth11, Rob C. van Lummel12,13, Mirjam
Pijnappels13, Carel G.M. Meskers13,14, Andrea B. Maier1,13
Affiliations 1Department of Medicine and Aged Care, University of Melbourne, Melbourne, Australia 2Department of Internal Medicine, Section of Gerontology and Geriatrics, VU University Medical Center, Amsterdam, The Netherlands 3Department of Internal Medicine, Amstelland Hospital, Amstelveen, The Netherlands 4Department of Gerontology and Geriatrics, Leiden University Medical Center, Leiden, The Netherlands 5Department of Geriatrics, Bronovo Hospital, The Hague, Netherlands, 6UPMC UM 76, INSERM U 974, CNRS 7215, Institute de Myologie, Paris, France 7Institute of Sport Sciences and Physiotherapy, University of Tartu, Tartu, Estonia 8School of Healthcare Science, John Dalton Building, Manchester Metropolitan University, Manchester, United Kingdom 9Division of Medical Sciences and Graduate Entry Medicine, MRC-ARUk Centre of Excellence for Musculoskeletal Ageing Research, University of Nottingham, Royal Derby Hospital Centre, Derby, United Kingdom 10Gerontology Research Centre and Department of Health Sciences, University of Jyväskylä, Jyväskylä, Finland 11Department of Biology of Physical Activity, University of Jyväskylä, Jyväskylä, Finland 12McRoberts BV, The Hague, The Netherlands 13Department of Human Movement Sciences, MOVE Research Institute, Vrije Universiteit, The Netherlands 14Department of Rehabilitation Medicine, VU University Medical Center, Amsterdam, The Netherlands
Rationale: Handgrip strength (HGS) is used to identify individuals with low muscle strength
(dynapenia). The influence of the number of attempts on maximal HGS is not yet known
and may differ depending on age and health status. This study aimed to assess how many
attempts of HGS are required to obtain maximal HGS.
Methods: Three cohorts (939 individuals) differing in age and health status were included.
HGS was assessed three times and explored as continuous and dichotomous variable.
Paired t-test, intraclass correlation coefficients (ICC) and Bland-Altman analysis were used
to test reproducibility of HGS. The number of individuals with misclassified dynapenia at
attempts 1 and 2 with respect to attempt 3 were assessed.
Results: Results showed the same pattern in all three cohorts. Maximal HGS at attempts 1
and 2 was higher than at attempt 3 on population level (P < 0.001 for all three cohorts). ICC
values between all attempts were above 0.8, indicating moderate to high reproducibility.
Bland-Altman analysis showed that 41.0 to 58.9% of individuals had the highest HGS at
attempt 2 and 12.4 to 37.2% at attempt 3. The percentage of individuals with a maximal
HGS above the gender-specific cut-off value at attempt 3 compared with attempts 1 and 2
ranged from 0 to 50.0%, with a higher percentage of misclassification in middle-aged and
older populations.
Conclusion: Maximal HGS is dependent on the number of attempts, independent of age
and health status. To assess maximal HGS, at least three attempts are needed if HGS is
considered to be a continuous variable. If HGS is considered as a discrete variable to
assess dynapenia, two attempts are sufficient to assess dynapenia in younger populations.
Misclassification should be taken into account in middle-aged and older populations.
96
(38) Is Being Malnourished According to the ESPEN Definition for
Malnutrition Associated With Clinically Relevant Outcome Measures
in Geriatric Outpatients?
N.M. van Rijssen RD, MSc, A.G.M. Rojer BSc, M.C. Trappenburg MD, PhD, E.M.
Reijnierse MSc, C.G.M. Meskers MD, PhD, A.B. Maier MD, PhD, M.A.E. de van der
Schueren RD, PhD Submitted
Background and aim: A body of evidence is supporting the association between (the risk
of) malnutrition in relation to physical performance, muscle strength, risk for depression and
cognitive status in geriatric outpatients. Associations between being malnourished
according to the newly proposed ESPEN definition for malnutrition and clinically relevant
outcome measures of the aforementioned variables have not been confirmed yet.
Therefore, the aim of this study was to examine the association between being
malnourished according to the ESPEN definition and clinically relevant outcome measures
in geriatric outpatients.
Methods: Associations between malnutrition and handgrip strength (HGS, kg), Short
Physical Performance Battery (SPPB-score, points), Timed Up and Go test (TUG,
seconds), and Hospital Anxiety and Depression Scale (HADS depression score, points),
were analysed using linear regression. History of falls (falls, yes/no) and a low score on the
Mini Mental-State Examination (MMSE-score ≤24 points) were analysed using logistic
regression. All analyses were adjusted for age and gender.
Results: A total of 185 geriatric outpatients (60% women) with a mean age of 82 (±7.3)
years were included. Being malnourished (8.2%) according to the ESPEN definition was
significantly associated with a lower HGS (-3.38 kg, p=0.031), lower SPPB-score (-1.8
point, p=0.025), higher TUG-time (1.35 times higher time, p=0.020) and higher HADS
depression score (2.03 times higher score, p=0.007). Being malnourished tended towards
an association with falls (OR 3.84, p=0.087). No significant association was found with low
MMSE-score (OR 2.61, p=0.110).
Conclusion: This study is the first to confirm the association between being malnourished,
defined by the ESPEN definition and clinically relevant outcome measures in geriatric
outpatients.
Keywords: malnutrition, nutrition, physical performance, muscle strength, depression,
aged.
97
(39) Instrumented assessment of physical activity is associated with
muscle function but not with muscle mass in a general population
A.G.M. Rojer BSc, E.M. Reijnierse MSc, M.C. Trappenburg MD, PhD, R.C. van Lummel MSc, M.
Niessen PhD, K.S. van Schooten PhD, M. Pijnappels PhD, C.G.M. Meskers MD, PhD, A.B. Maier
MD, PhD
Accepted, 26th of june 2017, Journal of Aging and Health.
Objectives: Self-reported physical activity has shown to affect muscle-related
parameters. As self-report is likely biased, this study aimed to assess the association
between instrumented physical activity (I-PA) and muscle-related parameters in a
general population.
Methods: Included were 156 young-to-middle-aged and 80 older community-dwelling
adults. Seven days of trunk accelerometry (Dynaport MoveMonitor, McRoberts B.V.)
quantified daily physical activity (i.e. active/inactive duration, number and mean duration
of active/inactive periods, number of steps per day). Muscle-related parameters
included muscle mass, handgrip strength and gait speed.
Results: I-PA was associated with handgrip strength in young-to-middle-aged adults
and with gait speed in older adults. I-PA was not associated with muscle mass in either
age-group.
Discussion: The association between I-PA and muscle-related parameters was age
dependent. The lack of an association between I-PA and muscle mass indicates the
relevance of muscle function rather than muscle mass.
Keywords: Motor activity; Sarcopenia; Muscle strength; Aged; Activity monitoring;
Physical performance
98
(40) Role of glycation on natural killer cell function
Philip Rosenstock1, Kaya Bork1, Vinayaga Srinivasan Gnanapragassam1, Franziska
Frank1, Rüdiger Horstkorte1
1Institute for Physiological Chemistry, Martin-Luther-University Halle-Wittenberg, Halle (Saale), Germany
Natural killer cells (NK cells) are cells of the innate immune system and provide a
major defence against tumor cells and virus infected cells. They are able to kill
modified or infected cells without the need for antigen recognition and can
produce different cytokines to modulate the function of other immune cells. The
activity of NK cells is regulated by activating and inhibitory receptors. In general
NK cells can be divided into two subpopulation based on their expression of
CD56. CD56 is also known as the neural cell adhesion molecule (NCAM) and is a
polysialylated molecule. CD56bright NK cells with a high CD56 expression are
predominantly cytokine producing cells whereas the CD56dim NK cells with a
lower CD56 expression are more cytotoxic. During human aging, which is
associated with an increasing amount of advanced glycation end products
(AGEs) the number, phenotype and function of NK cells is changed. We
investigated the effect of aging on human natural killer cells using the human NK
cell line NK-92 and studied how glycation and sialylation affects the function of
these cells. In our experiments treatment of NK-92 cells with methylglyoxal lead
to glycation and formation of advanced glycation end products. Using a flow
cytometry based cytotoxicity assay we could show that this glycation inhibits the
cytotoxic activity of NK-92 cells.
Presenting author’s email address: philip.rosenstock@uk-halle.de
99
(41) Role of ascorbic acid in protein glycation and PC12
adhesion/neurite outgrowth
Jonas Scheffler1, Kaya Bork1, Vinayaga Srinivasan Gnanapragassam1, Franziska
Frank1, Andreas Simm2, Rüdiger Horstkorte1
1 Institute for Physiological Chemistry, Halle (Saale), Germany
2 Clinic for Heart Surgery, Halle (Saale), Germany
Ascorbic acid is not only known as an important cofactor for multiple enzymatic
pathways including the synthesis of collagen or stresshormones, but also as a major
antioxidant, reducing intra- and extracellular oxidative stress and scavenging free
radicals. However, recent studies suggest an involvement of ascorbic acid and its
degradation products in protein glycation and generation of advanced glycation
endproducts (AGEs). AGEs play an important role in many age-related diseases,
including atherosclerosis, cataract, renal insufficiency and Alzheimer’s disease. Thus,
long-lived proteins like collagens as well as the cells themselves are possibly altered in
stability and function after exposure to high amounts of ascorbic acid, which could lead
to decreased attachment and differentiation. We could show a negative effect of
ascorbic acid-induced glycation on the neurite outgrowth of neuron-like PC12 cells.
Since these cells can adhere on extracellular matrix proteins, we also investigated the
impact of collagen IV-glycation on adhesion and neurite outgrowth.
Presenting author’s email address: scheffler995@gmail.com
100
(42) Neural stem and progenitor cells for the in vitro 2D- and 3D-
analysis of human neurogenesis
Jovita Schiller, Matthias Jung, Anne Puls and Dan Rujescu
Neurodegenerative and -developmental diseases such as ASD and schizophrenia are
often associated with genetic variations. Single nucleotide polymorphisms and copy
number variations presumably affect the neural development, resulting in extensive
cognitive impairments that may have a great impact on cell functions. Using the
example of schizophrenia, several risk genes are associated with the disease and some
of them are involved in neural development, neurotransmission as well as the immune
system. Reprogramming of human somatic cells enables the generation of induced
pluripotent stem cells (iPSCs) bearing the genetic information. The neural differentiation
of iPSCs might elucidate the impact of genetic variations and enable the improvement
of antipsychotic drug treatment by sub-classifying patients at a molecular level.
Therefore, patient-specific B cell-derived iPSCs were differentiated into neuroepithelial
rosettes containing self-renewing cells, which were cultured as free-floating
neurospheres to improve cell proliferation. Neurosphere single cells were maintained as
stable populations of neural stem cells (NSCs), which were further differentiated into
functional progenitor and mature neural cells when cultured in an adherent culture
system. Transcript and protein analysis as well as double immunostaining of specific
markers such as SOX2, MSI-1, NEUROG3, and NESTIN confirmed the successful
generation of NSCs as well as more mature neural cells (GFAP, O4, TUBB3, STX, and
NEUN). Patch-clamp recordings verified the presence of different neuron subtypes.
Alternatively, differentiation of neurospheres in a 3D culture system generated cerebral
organoids within 4 weeks, mimicking different brain regions. The regional identity was
confirmed through the expression of telencephalic and hippocampal markers (FOXG1,
OTX1, PROX1, and AUTS2). The presence of early neural cell fates was analyzed
through transcript and protein expression of specific markers as well. Different subtypes
such as GABAergic (GABAAR), glutamatergic (vGLUT1), and motor neurons (HB9)
were also detectable. Together, these results confirmed the efficient neural
differentiation of human iPSCs using both culture systems, enabling functional studies
of healthy and diseased human cortical development concerning the impact of specific
genetic variations.
101
(43) PTTG1/securin as a quantitative trait locus candidate gene
controlling progenitor cell survival and organismal lifespan
Desiree Schuetz1, Andreas Brown1, Kalpana J. Nattamai2, Hartmut Geiger1,2
1 Institute of Molecular Medicine, Ulm University, Ulm, Germany
2 EHCB, CCHMC, Cincinnati, USA
Prior work identified a positive correlation between the percentage of hematopoietic
progenitor cells (HPCs), defined as cobblestone-area forming cells (CAFC) day 7, killed
by the genotoxic agent hydroxyurea (HU) and median lifespan in eight inbred mouse
strains. These data suggest a common underlying molecular mechanism for both traits
(response of HPCs to HU and lifespan). Given that HU kills proliferating cells, this
finding was interpreted to mean that the fraction of proliferating HSC/HPCs was
significantly higher in long-lived C57BL/6 (B6) versus short-lived DBA/2 mice. Using
recombinant inbred mice derived from the B6 and DBA/2 mouse strains we mapped a
quantitative trait locus (QTL) on chromosome 11 that is responsible for the HU-killing
phenotype. The locus was found to overlap with a locus that is linked to lifespan, further
emphasizing a likely common mechanism. With this knowledge, congenic mice were
generated that have a B6 background and the specific locus on chromosome 11
replaced by the corresponding segment of DBA/2 mice, named line A, or vice versa for
DBA/2 mice, named line K. Transfer of the loci was verified by single-nucleotide
polymorphism (SNP) analyses. Using the CAFC approach, we could show that HPCs
from DBA/2 and line A mice exhibit higher sensitivity to HU, which confirms that the
locus on chromosome 11 indeed determines the sensitivity of HPCs to HU. We also
analyzed the cell cycle dynamics of HPCs from B6, DBA/2 as well as the congenic
strains. Surprisingly the HPCs of the four mouse strains do not differ in cell cycle
dynamics, which reveals that the distinct response of HPCs to HU is not correlated to
HPCs proliferation. Pituitary tumor-transforming gene 1 (PTTG1, also known as securin)
has been identified as a candidate gene within the locus on chromosome 11, which is
primarily involved in the regulation of sister chromatid separation during cell division.
PTTG1 shows high expression in progenitor cells from DBA/2 and line A mice.
Experiments are underway to verify PTTG1 as the quantitative trait gene in this locus
and to reveal the underlying molecular mechanisms that determine the distinct response
of HPCs to HU treatment.
102
(44) High concentrations of methylglyoxal and glyoxal epigenetically
modify histrone H3 in human trophoblast cells in vitro
Tom Seeling1, Katarzyna J. Grybel1, S. Mareike Pendzialek1, Maria Schindler1, Axel
Imhof2, Ignasi Forné2 and Anne Navarrete Santos1
1Department of Anatomy and Cell Biology, Martin Luther University Halle-Wittenberg,
Faculty of Medicine, Halle (Saale), Germany 2Department of Medical Biology, Ludwig Maximilians University Munich, Faculty of
Medicine, Munich, Germany
The Developmental Origins of Health and Disease (DOHaD) hypothesis states, that
long-term health and onset of ageing-associated diseases are already set during fetal
development. Particularly, the maternal metabolism influences the embryonic uterine
environment with consequences for trophoblast and placental development in early
pregnancy. In western society the first-birth age for women increases, leading in part to
higher risk of suffering from metabolic diseases like diabetes mellitus. Diabetes mellitus
is characterized by high concentrations of glucose and its metabolites like methylglyoxal
(MGO) and glyoxal (GO).
The aim of this study was to analyze the influence of a diabetic environment associated
with high levels of MGO and GO on histone protein modifications and histone-modifying
enzymes in placental cells.
Histone modifications were examined by LC-MS/MS. Therefore, the human trophoblast
cell line AC-1M88 was cultured with 100 µM MGO and GO simultaneously for 72 hours.
The exposition to MGO and GO changed the methylation pattern on several lysines,
especially on K9 and K27. Higher methylation was also detected at K79 after exposition
to MGO and GO, which is a marker for aging cells. Additionally expression of
methyltransferases and demethylase were examined on transcriptional level.
Our data show that trophoblast cells challenged by aging factors (MGO and GO),
changed epigenetic background to support survival through alterations in histone
modifications. These changes may promote metabolic programming with consequences
in later life.
This work was supported by the German Research Council (DFG RTG ProMoAge 2155), the FP7-
EpiHealthNet (N°317146), COST Action EpiConcept FA 1201 and SALAAM BM1308.
103
(45) Lysine acetylation patterns in T-Cells in the context of ageing
Georgiana Toma, Dagmar Quandt and Barbara Seliger
Institute of Medical Immunology, Martin-Luther University Halle, Germany
Ageing is a complex process affecting a wide range of physiological functions, including
the immune system, for which the age-related alterations are called
immunosenescence. One factor contributing to this process is T cell senescence.
Posttranslational modifications (PTM) of proteins are one of the key mechanisms
involved in aging. The most abundant PTMs are phosphorylation and acetylation.
Protein Acetylation is regulated by 2 types of enzymes: acetyltranferases and
deacetylases (initially named histone deacetylases – HDAC). HDACs are grouped into
four classes, the third being Sirtuins, and they have various substrates beyond histones.
Sirtuins, in particular Sirtuin 1, 3 and 6 have been associated to ageing. Increasing the
functionality of sirtuin-1 has been proven to prolong lifespan in certain model systems.
How T cell senescence is regulated by alterations of acetylation of key transcription
factors as well as cellular proteins is currently unknown.
Therefore our aim is to identify age-related alterations within the acetylation pattern of
different T cell subsets. This analysis will include acetyltransferase as well as HDAC
expression studies on mRNA and at the protein level. In addition we will analyse the
enzymatic activity of those enzymes in T cells.
Our current strategy is to determine differences in the protein acetylation patterns of
CD8+ and CD4+ T cells sorted from PBMCs obtained from young (< 30 years old) and
older (> 60 years old) healthy blood donors. To visualize the acetylation patterns, we
treat the cells with trichostatin a (TSA) – a HDAC class I and II inhibitor and
nicotinamide (NAM) – an inhibitor of HDAC class III (SIRT). Preliminary results show
more dramatic alterations in T cell acetylation patterns upon TSA treatment. That would
speak for a more prominent role of HDAC I and II as compared to sirtuins. Differentially
acetylatetd candidate proteins will be identified by mass spectrometry and functional T
cell studies will be performed.
Results of our study will potentially help to find new ways to improve and modulate T
cell immunity of the elderly.
104
(46) Epigenetic regulation of gene transcription by non-enzymatic
protein modification
Arina Urazova1, Kristin Wächter1, Patrick Winterhalter1, Wolfgang Hoehenwarter2,
Jochen Balbach3, Andreas Simm1
1University Hospital Halle, Clinic for Heart Surgery, Halle (Saale)
2Leibniz Institute of Plant Biochemistry, Halle (Saale)
3Institute of Physics, Halle (Saale)
E-mail: arina.urazova@uk-halle.de
Formation of Advanced Glycated End Products (AGEs) is a result of glycation, which is
non-enzymatic reaction between reducing sugars, or other α-dicarbonyl compounds,
and amino groups of proteins, lipids and nucleic acids. AGEs accumulate during aging
and have been implicated in the pathophysiology of numerous age-related diseases.
Glycation of nuclear proteins at lysins or arginins may alter their function and may have
an impact on epigenetic regulation of gene transcription.
Glycated nuclear proteins from young and replicative senescent cells, as well as from
normal and tumor cells, are going to be compared, and identified modified proteins will
be further analyzed.
Nuclei from human embryonic kidney cells 293A (HEK293A) and human kidney
carcinoma cell line Caki-2 were isolated and AGE-modified nuclear proteins were
compared via western blot using antibody against carboxymethyl-lysine (CML),
hydroimidazolone MG-H1 and argpyrimidine. Different pattern of modified proteins in
HEK293A and Caki-2 cell lines was observed. LC-MS/MS analysis revealed that in
HEK293A cells 7 out of 3424 identified proteins are modified, while in Caki-2 cells
among 2701 proteins 11 are modified. Two of them, Histone H4 and Myb-binding
protein 1A, were identified in both cell lines. Amount of modified peptides (12 out of
32127 identified peptides in HEK293A cells and 14 out of 23528 in Caki-2 cells) is too
low for reproducible quantification of differences in these modifications.
Immunoprecipitation was used to enrich samples with modified proteins prior to
analysis, enrichment steps are elucidated and optimized.
Identified AGE-modification of lysins/arginins within nuclear proteins indicate on a
possibility to modify the epigenetic regulation of cells.
105
(47) Only Body Mass Index prredicts metabolic syndrome in older
people
Jeanine M. Van Ancum MD a, Nini H. Jonkman PhD a, Natasja M. van Schoor PhD b,
Emily Tressel Bsc a, Carel G.M. Meskers MD, PhD c, Mirjam Pijnappels PhD a, Andrea
B. Maier MD, PhD a,d
a Department of Human Movement Sciences, Vrije Universiteit Amsterdam, Amsterdam Movement Sciences, The
Netherlands. b
Amsterdam Public Health Research Institute, Department of Epidemiology and Biostatistics, VU University Medical Center, Amsterdam, The Netherlands
c Department of Rehabilitation Medicine, VU University Medical Center, Amsterdam Movement Sciences, the
Netherlands. d
Department of Medicine and Aged Care, Royal Melbourne Hospital, University of Melbourne, Melbourne, Victoria,
Australia.
Background: Demographics, lifestyle, clinical measures and physical performance
have individually been shown to be associated with metabolic syndrome (MetS).
However, the combination of these variables has not yet been investigated in a large
population of older people. We aimed to identify independent determinants of MetS
prevalence, as well as predictors of MetS at three year follow-up in older people.
Methods: Demographics, lifestyle, clinical measures and physical performance, along
with the prevalence and incidence of MetS over three years were collected within the
Longitudinal Aging Study Amsterdam including individuals aged 55 years and older.
MetS was defined according to the National Cholesterol Education Program Adult
Treatment Panel III (NCEP-ATPIII). Stepwise backward selection in logistic regression
analyses was used to identify independent determinants of MetS prevalence in a cross-
sectional analysis (n=1292) and predictors of MetS incidence at three year follow-up
(n=218).
Results: MetS was prevalent in 37.1% individuals (n=479), and MetS incidence over
three years was 30.3% (n=66). In the cross-sectional analysis, heart disease, artery
disease, diabetes, alcohol consumption and Body Mass Index (BMI) were significantly
associated with MetS. In the longitudinal analysis, only BMI was an independent
predictor of MetS.
Conclusion: BMI predicts the development of MetS over three years, providing a first
step towards an easy-to-assess screening tool for healthcare in older people.
106
(48) Muscle strength and muscle mass in older patients during
hospitalization: the EMPOWER study
Jeanine M. Van Ancum MD a, Kira Scheerman MSc
b, Vincent D. Pierik BSc
b, Siger T.
Numans BSc b, Sjors Verlaan MSc
b, Hanne E. Smeenk MD
c, Monique Slee-Valentijn MD
d,
Roeliene C. Kruizinga MD, PhD b, Carel G.M. Meskers MD, PhD
e, Andrea B. Maier MD,
PhD a,f
a Department of Human Movement Sciences, Vrije Universiteit Amsterdam, Amsterdam Movement Sciences, The
Netherlands b Department of Internal Medicine, Section of Gerontology and Geriatrics, VU University Medical Center, Amsterdam
Movement Sciences, the Netherlands. c GERION, Department of General Practice and Elderly Care Medicine, VU University Medical Center, Amsterdam,
the Netherlands. d Center of Excellence in Geriatric Rehabilitation, Cordaan, Amsterdam, the Netherlands.
e Department of Rehabilitation Medicine, VU University Medical Center, Amsterdam Movement Sciences, The
Netherlands f Department of Medicine and Aged Care, Royal Melbourne Hospital, University of Melbourne, Melbourne, Victoria, Australia.
Background: Low muscle strength and muscle mass are associated with an increased length
of hospital stay and higher mortality rate in inpatients. To what extent hospitalization affects
muscle strength and muscle mass is unclear. We aimed to assess muscle strength and muscle
mass at admission and during hospitalization in older patients and their relation with being at
risk of four geriatric conditions: delirium, falls, malnutrition and functional disability.
Methods: The EMPOWER inception cohort included patients aged 70 years and older,
admitted to four wards of the VU university medical center in the Netherlands between April and
December 2015. At admission, patients were screened for being at risk of four geriatric
conditions. Both at admission and at discharge, muscle strength and muscle mass were
assessed.
Results: A total of 373 patients (mean age, SD: 79.6, 6.38 years) were included at admission of
which 224 patients (mean age, SD: 80.1, 6.32 years) were left for analysis at discharge. At
admission, lower muscle strength in both female and male patients and low muscle mass in
male patients were associated with being at risk of a higher cumulative number of geriatric
conditions. Muscle strength increased during hospitalization, but no change in muscle mass
was observed. Changes in muscle measures were not associated with being at risk of geriatric
conditions.
Conclusion: Older patients with lower muscle strength and muscle mass at admission were at
risk of a higher cumulative number of geriatric conditions. However, being at risk of geriatric
conditions did not forecast further decrease in muscle strength and muscle mass during
hospitalization.
107
(49) Muscle strength and muscle mass are associated with pre- and
posthospitalization falls in older male patients
Jeanine M. Van Ancum MD a, Mirjam Pijnappels PhD a, Nini H. Jonkman PhD a, Kira
Scheerman MSc b, Sjors Verlaan MSc b, Carel G.M. Meskers MD, PhD c, Andrea B.
Maier MD, PhD a,d
a Department of Human Movement Sciences, Vrije Universiteit Amsterdam, Amsterdam Movement Sciences, The
Netherlands. b
Department of Internal Medicine, Section of Gerontology and Geriatrics, VU University Medical Center, Amsterdam Movement Sciences, the Netherlands. c Department of Rehabilitation Medicine, VU University Medical Center, Amsterdam Movement Sciences, the
Netherlands. d
Department of Medicine and Aged Care, Royal Melbourne Hospital, University of Melbourne, Melbourne, Victoria, Australia.
Introduction: Low muscle strength and muscle mass are associated with falls in
community-dwelling older adults, but it is unknown if these muscle measures are also
associated with falls in older inpatients. This study aimed to investigate the association
between muscle measures and pre- and post-hospitalization falls in older inpatients.
Methods: An inception cohort of inpatients aged 70 years and older admitted to an
academic teaching hospital were included in this study, from April to December 2015.
Handgrip strength (HGS) and muscle mass were measured at admission using
dynamometry and bioelectrical impedance analysis. Pre-hospitalization falls were assessed
at admission with a questionnaire and dichotomized as at least one fall in six months before
admission. Post-hospitalization falls were assessed by telephone interview three months
after discharge and dichotomized as at least one fall. Associations were analyzed with
logistic regression analysis, stratified for gender.
Results: The study included 378 inpatients (mean age, SD: 79.7, 6.4). Pre-hospitalization,
50% of female and 41% of male patients experienced at least one fall. Post-hospitalization,
18% of female and 23% of male patients experienced at least one fall. Lower HGS was
associated with both pre- and post-hospitalization falls, and lower muscle mass was
associated with post-hospitalization falls in male, but not in female patients.
Conclusion: These findings highlight the likely involvement of muscle strength and muscle
mass in the occurrence of pre- and post-hospitalization falls in males, indicating a possible
target for interventions in older male patients with low HGS and muscle mass upon
admission to the hospital.
108
(50) Mitochondria preconditioning to promote healthy aging in
C.elegans
Natascia Ventura
The nematode C.elegans is a powerful model organism for aging studies. Hormesis or
preconditioning is a phenomenon by which the body benefits from various kinds of
moderate stress, which at high doses are harmful. A growing body of evidence indicates
that different forms of preconditioning could be exploited as a strategy to protect against
degenerative disorders in human and to extend lifespan in C. elegans.
Mitochondria play a central role in the aging process and hormetic-like effects on
C. elegans lifespan have been observed following different degrees of mitochondrial
stress [1, 2]. In the past years we have shown that either genetic or pharmacological
interventions reducing mitochondrial activity can promote C. elegans healthspan [3, 4].
Moreover we have identified different evolutionarily conserved, cytoprotective
responses causally involved in mitochondrial stress control of longevity such as
autophagy or transcription factors (p53 and Hif1) [2, 5]. Thanks to a recently established
phenotypebased screening platform [6] we are currently trying to identify possible
dietary components promoting healthspan through mitochondrial preconditioning.
1. Rea, S.L.*, Ventura N.*, and Johnson T.E. Relationship between mitochondrial electron transport chain dysfunction, development, and life extension in Caenorhabditis elegans. PLoS Biol, 2007. 5(10): p. e259. 2. Ventura, N., et al., p53/CEP-1 increases or decreases lifespan, depending on level of mitochondrial bioenergetic stress. Aging Cell, 2009. 8(4): p. 380-93. 3. Maglioni, S., et al., Mitochondrial stress extends lifespan in C. elegans through neuronal hormesis. Exp Gerontol, 2014. 56: p. 89-98. 4. Schiavi, A., et al., Autophagy induction extends lifespan and reduces lipid content in response to frataxin silencing in C. elegans. Exp Gerontol, 2013. 48(2): p. 191-201. 5. Schiavi, A., et al., Iron-Starvation-Induced Mitophagy Mediates Lifespan Extension upon Mitochondrial Stress in C. elegans. Curr Biol, 2015. 25(14): p. 1810-22. 6. Maglioni, S., et al., An automated phenotype-based microscopy screen to identify pro-longevity interventions
109
(51) Isolation and characterization of adipose-derived stem cells
(ASCs) from young and old female rabbits
Christin Volk1, Alexander Navarrete Santos2, Matthias Jung3, Anne Navarrete Santos1,
and Juliane-Susanne Jung1
1 Department of Anatomy and Cell Biology,
2 Centre for Medical Basic Research,
3 Department of Psychiatry, Psychotherapy, Psychosomatic Medicine, all: Martin Luther University Halle-Wittenberg,
Halle (Saale), Germany
The UN estimates that the number of people older than 65 years will grow from 7% in 2000 to 16%
in 2050 worldwide. Therefore, research on aging mechanisms and aged-related disease is one
major issue of our ageing world. Adult stem cells such as mesenchymal stem cells are affected by
molecular and cellular aging mechanisms. Mesenchymal stem cells have the capability to self-renew
and can differentiate into multiple cell types of the mesoderm germ layer. Bone marrow-derived stem
cells represent a well-established model, but donor material is limited and requires invasive
methods. Adipose tissue is an alternative source of mesenchymal stem cells due to facile harvesting
techniques. Therefore adipose-derived stem cells (ASCs) represent a promising tool for studying the
biological characteristics of tissue regeneration and age-related diseases in detail.
In the present study, ASCs were isolated from visceral adipose tissue of young (16 weeks) and old
female rabbits (>108 weeks). ASCs were prepared from tissue by mincing and dissociation with
collagenase A. ASCs were cultured under appropriate cell culture conditions until passage 3. Cells
were analyzed morphologically by microscopy at day 7, 10, 15, and 21. Furthermore, flow cytometry
and transcript analysis were used for molecular characterization of ASCs and differentiated early
adipocytes.
After 10 days of culturing, in vitro ASCs showed the typical morphology of mesenchymal stem cells.
Most of the cells were positive for the ASCs-associated cluster of differentiation (CD) 105 (94.3%)
and CD73 (51.2%) and negative for CD14 (0.0%), demonstrating their mesenchymal origin. Isolated
ASCs expressed mesenchymal stem cell markers including Myc, Klf4, Chd1, Rest and Kat6A, while
the pluripotency-associated genes like Nanog, Oct4 and Sox2 were not expressed. Adipogenic
differentiation of the ASCs (passage 3) was induced by dexamethasone (1.0 μM final), 3-isobutyl-1-
methylxanthine (IBMX - 0.5 mM final), indomethacin (0.2 mM final) and insulin (10.0 μM final)
enabling the analysis of aging during fat cell generation and regeneration. The adipogenic
differentiation efficiency was low, with 4.3% for ASCs from young rabbits. In a next step the ASC
differentiation procedure will be optimized to increase adipogenic differentiation.
Together, we successfully established rabbit ASC cultures representing an in vitro model for the
analysis of stem cell aging mechanisms.
110
(52) Bakery products as functional food
Kristin Wächter1, Alexander Navarrete Santos2, Veronika Somoza3, Hendrik Treede1,
Andreas Simm1, 2,
1 Clinic for Heart Surgery, Martin-Luther-University Halle-Wittenberg, Halle (Saale)
2 Center for medical basic research, Martin-Luther University Halle-Wittenberg, Halle (Saale)
3 Research Platform Molecular Food Science, University of Vienna, Vienna, Austria
Advanced glycation end products (AGEs) are the result of a non-enzymatic reaction of
proteins with reactive carbohydrates. Heat-processed food like bread contains high
amounts of AGEs. Numerous previous studies suggest an antioxidative and
cardioprotective potential of extracts from the bread crust (BCE). The activation of NF-
κB signaling pathway by BCE is well understood. However it is largely unknown whether
NRF2, the master regulator of oxidative stress resistance in mammalian cells, is
affected by BCE. Furthermore to date there is only little knowledge concerning the
bioactive compounds, which accounts for the antioxidant BCE-properties.
Understanding the molecular mechanism of BCE action is an important prerequisite to
establish BCE as a functional food product. Therefore, we developed a cell culture
model to elucidate the effect of different bread crust extracts on NF-κB as well as NRF2
activation in an endothelial reporter cell line (EAhy926 dual plasmid, NF-κB–GFP,
NRF2-mCherry). An increased nuclear translocation of NF-κB and NRF2 in these cells
results in elevated GFP and mCherry expression, which are analyzed by flow cytometry.
Our data showed for the first time, that soluble extracts from bread crust are capable to
stimulate not only NF-κB but also NRF2 signaling pathways in endothelial cells in a
dosage- and time-dependent manner. In a second approach, we uncovered that AGE
modified BSA is able to activate both transcription factors in comparison to unmodified
BSA in endothelial cells. This tempts us to speculate, that especially AGE-modified
proteins present in BCE could mediate the antioxidative potential. In addition, by size
exclusion experiments we found that peptides smaller than 3 kDa are sufficient for
NRF2 and NF-κB activation.
In continuative studies we want to optimize and enhance the action of BCE by changing
the baking conditions and ingredients. Furthermore, the results will be confirmed by
feeding studies in mice.
111
(53) Development of a tBHP-induced senescence model to study
molecular mechanisms of extrinsic skin aging
Sophia Wedel1, Maria Cavinato1, Pidder Jansen-Dürr1
1 Institute for Biomedical Aging Research, Innsbruck University, Innsbruck, Austria
Skin aging is influenced by intrinsic and environmental factors. Epidemiological studies
and many research groups have shown that sunlight, tobacco smoke and air pollution
can prematurely induce aging in skin. Still, the mechanisms that underlie the changes of
the skin’s integrity and appearance upon exposition to these extrinsic factors are
scarcely understood. Therefore, it is of great importance to develop new model that help
us to improve our understanding of this matter.
We have established a model for studying the mechanisms of extrinsic skin aging by
treating human dermal fibroblasts and reconstructed human skin equivalents with eight
sublethal doses of the organic peroxide tert-butyl-hydroperoxide (tBHP). The oxidative
stress injury and depletion of antioxidant defense resulting from tBHP treatment are
very similar to and consequently able to mimic the effects of tobacco smoke and
particulate matter on skin.
We found that tBHP treatment increased intracellular reactive oxygen species (ROS)
levels and subsequently induced DNA damage response in HFFs. Additionally, we were
able to show that tBHP induced changes in morphology and led to proliferative arrest by
triggering regulation of proteins that are important for cell cycle progression and stress
response including p53, p21cip/waf and pRB, p16INK4A in the fibroblast monolayer.
Furthermore, more than 90% of treated HFFs were positive for the activity of
senescence-associated β-galactosidase nine days after the tBHP administration
started. At this point in time cell death occurred infrequently. In human skin equivalents
treatment with tBHP increased thickness of stratum corneum and the abundance of
pyknotic nuclei in the epidermis, which can also be detected upon UV irradiation.
We have developed a new model system for extrinsic skin aging, employing treatment
with tBHP and inducing a senescent phenotype in fibroblasts. Additionally, we were able
to collect preliminary on skin equivalents. This new experimental setup allows us to do
further research on mechanisms of extrinsic skin aging and ultimately helps to improve
existing and map out novel anti-skin aging strategies.
112
(54) Determination of AGE modifications: To detect or not to detect,
that is the question
Patrick Rainer Winterhalter, Andreas Simm
BACKGROUND: In human age-related disorders, the in vivo formation of advanced
glycation end products (AGEs) is an important and well-known pathogenic event. The
generation of AGEs under pathological conditions is an unpreventable process via the
slow, non-enzymatic formation of amino acids with omnipresent metabolites of the
glycolysis as carbohydrates or dicarbonyls. However, the detection of AGEs is still
problematic and sometimes non-reproducible. Reasons might be the high variety of
possible modifications that are often not identified yet. In addition, the low abundance
and the insufficient identification tools hamper the description of AGEs and associated
dysfunctions.
OBJECTIVE: To determine the enrichment and detection conditions for AGE
modifications, in particular several mono- and polyclonal anti-AGE antibodies were
tested by protein immunoblot.
METHODS: Anti-AGE antibodies from different sources were analyzed utilizing non- or
exogenously AGE-modified model proteins as well as crude AKR-2B cell line lysate.
Thereby, the affinity and specificity for AGEs in general and for specific AGEs as well as
the test conditions were analyzed. A green fluorescent secondary antibody visualized
the protein immunoblots.
RESULTS: For monoclonal anti-AGE antibodies, the sensitivity of the specific AGE
detection is mainly limited. Further, an antigenic effect of surrounding amino acids is
assumed. In addition, the blocking solutions mainly modify the results. Polyclonal
antibodies have clear benefits regarding sensitivity and specificity.
CONCLUSION: A successful determination of AGEs mainly depends on the used
antibody and conditions.
113
(55) Glycation of Wnt inhibits canonical signaling
Zhennan Ye1, Sonnhild Mittag1, Andreas Simm2, Otmar Huber1
1Institute of Biochemistry II, Jena University Hospital, Friedrich Schiller University Jena, Nonnenplan 2-4, D-07743
Jena, Germany; 2Department of Cadiothoracic Surgery, University Hospital Halle, Ernst-Grube Str. 40, D-06120 Halle (Saale),
Germany
Signaling pathways involved in repair, regeneration and stem cell proliferation often
exhibit reduced activity during aging. Accumulation of advanced glycation endproducts
(AGEs) is a typical sign of aging tissues and organs. We here addressed if this
posttranslational modification affects Wnt/β-catenin signaling. Glyoxal (GO) or
methylglyoxal (MGO) treatment of Wnt3a-conditioned medium inducing glycation of
medium components reduces Topflash reporter gene activity. Control experiments
provide evidence that glycation of Wnt3a and not of other medium components are
responsible for this effect. In this respect we show a GO-induced molecular weight shift
of immunoprecipitated Wnt3a. In our hands it appears that AGEs per se can stimulate
canonical Wnt signaling, whereas glycation of Wnt has an inhibitory effect. Consistent
with lowered reporter gene activity, reduced transcription of endogenous target genes
and modulation of total β-catenin and phospho-β-catenin levels were observed. From
these results we speculate that the final effects of glycation depends on an interplay of
RAGE- and Wnt-mediated effects.
114
List of Speakers A – G Campisi, Judith Buck Institute for Research on Aging Novato, 8001 redwood Boulevard CA 94945, USA Lawrence Berkeley National Laboratory, Berkeley CA 94720, USA E-Mail: jcampisi@buckinstitute.org jcampisi@lbl.gov rabulencia@buckinstitute.org Tel.: +1 (0) 415 - 209 2043
Chondrogianni, Niki Institute of Biology, Medicinal Chemistry and Biotechnology 48 Vassileos Constantinou Ave. 116 35 Athens, GR E-mail: nikichon@eie.gr Tel.: +30 (0) 210-7273768 Fax: +30 (0) 210-7273677
de Grey, Aubrey Aubrey de Grey SENS Research Foundation 110 Pioneer Way, Suite J Mountain View, CA 94041, USA E-Mail: Aubrey@sens.org Tel: +1 650 938 6100
Eifert, Sandra Klinikum der Universität München Campus Grosshadern Herzchirurgische Klinik und Poliklinik Marchioninistraße 15 81377 München, D E-Mail: Sandra.Eifert@med.uni-muenchen.de Tel.: +49 (0) 89 – 4400 73458 Fax: +49 (0) 89 – 4400 78898
Emmert, Maximilian UniversitätsSital Zürich Universitäres Herzzentrum Zürich Rämistraße 11 8091 Zürich, CH E-Mail: Maximilian.Emmert@usz.ch Tel.: +41 (0) 44 – 255 9362
Franceschi, Claudio University of Bologna Department of Experimental, Diagnostic and Specialty Medicine DIMES Via Zamboni, 33 40126 Bologna BO, I E-Mail: stefano.salvioli@unibo.it Tel.:
Fülöp, Tamas Université de Sherbrooke Département de pédiatrie /Dienst d’immunologie-allergologie 2500, boul. Kanada J1K 2R1, CAN E-Mail: Tamas.Fulop@USherbrooke.ca Tel.: +1 (0) 819 – 821 8000
Gekle, Michael Julius-Bernstein-Institut für Physiologie Magdeburger Straße 6 D-06112 Halle (Saale) E-Mail: michael.gekle@medizin.uni-halle.de
Tel.: +49 (0) 345 – 557 1389
Fax: +49 (0) 345 – 557 4019
115
List of Speakers G - L
Gosch, Markus Universitätsprofessor der Paracelsus Medizinischen Privatuniversität für Geriatrie Chefarzt Med. Klinik 2 - Geriatrie Klinikum Nord Prof.-Ernst-Nathan-Straße 1, 90419 Nürnberg, D E-Mail: markus.gosch@klinikum-nuernberg.de Tel.: +49 (0) 911 398 - 2435 Fax: +49 (0) 911 398 - 2117
Ground, Ian Richard Department of Philosophy University of Hertfordshire Hatfield AL10 9AB, UK E-Mail: i.ground@herts.ac.uk
Grune, Tilman Deutsches Institut für Ernährungsforschung Potsdam-Rehbrücke Arthur-Scheunert-Allee 114-116 14558 Nuthetal, D Assistentin: Stephanie Wirth E-Mail: office.science@dife.de Tel. +49 (0) 33200 - 88-2416 Fax +49 (0) 33200 - 88-2555
Haendeler, Judith IUF – Leibniz-Institut für umwelt-medizinische Forschung gGmbH Auf’m Hennekamp 50 40225 Düsseldorf, D
E-Mail: juhae001@uni-duesseldorf.de
Tel.: +49 (0) 211 3389 291
Hoeijmakers, Jan Erasmus MC University Medical Center Rotterdam P.O. Box 2040, 3000 CA Rotterdam, NL
E-Mail: j.hoeijmakers@erasmusmc.nl Tel.: +31 (0) 10 – 7044844
Kirkland, James Robert and Arlene Kogod Center on Aging Minnesota-Rochester Mayo-Klinik 200 Erste St. SW Rochester, MN 55905, USA E-Mail: Kirkland.James@mayo.edu Kari Sczepanski Administrative Assistant Email: sczepanski.kari@mayo.edu Phone: +1 (0) 507 266-9151 Fax: +1 (0) 507 293-3853
Klöss, Thomas Martin-Luther-Universität Halle-Wittenberg Ärztlicher Direktor Ernst-Grube-Straße 30 06120 Halle (Saale), D
E-Mail : thomas.kloess@uk-halle.de Tel. : +49 (0) 345 – 557 4481
Lepperdinger, Günter Universität Salzburg Hellbrunnerstraße 34 5020 Salzburg, AT
E-Mail: guenter.lepperdinger@sbg.ac.at Tel.: +43 (0) 662 - 8044 5739 Fax: +43 (0) 662 - 8044 144
116
List of Speakers M - R
Maier, Andrea The University of Melbourne Melbourne Health The Royal Melbourne Hospital - City Campus, Level 6 North Grattan Street Parkville Victoria 3052, AUS E-Mail: Andrea.Maier@mh.org.au Tel.: +61 (0) 3 - 8387 2137 Fax: +61 (0) 3 - 8387 222
Martin, George M. University of Washington Department of Pathology Professor of Pathology Emeritus K-543 Health Sciences Building BOX 357470 1959 NE Pacific Street Seattle, WA 98195-7470, USA E-Mail: gmmartin@uw.edu Tel.: +1 (0) 206 - 543 5088 Fax: +1 (0) 206 - 685 8356
Mirelman, Anat Director- Laboratory of Early Markers of Neurodegeneration (LEMON) Department of Neurology Tel Aviv Sourasky Medical Center 6 Weizmann st, Tel Aviv 64239, IL
E-Mail: anatmi@tlvmc.gov.il Tel: +972 (0) 3 6973960 Fax: +972 (0) 3 6947514
Moskalev, Alexey Syktyvkar State University; Institute of Biology, Komi Science Center of the Ural Division of the Russian Academy of Science; MIPT (State University) 28 Kommunisticheskaja Str. 167982 Syktyvkar, RUS E-Mail: amoskalev@list.ru Tel.: +7 (0) 8212312894
Munoz-Canove, Pura ICREA and Pompeu Fabra University Department of Experimental and Health Sciences Dr. Aiguader, 88 08003 Barcelona, ES E-Mail: pura.munoz@upf.edu
Niemann, Bernd Klinik für Herz-, Kinderherz- und Gefäßchirurgie Universitätsklinikum Giessen und Marburg Standort Giessen Rudolf Buchheim Strasse 7 35392 Giessen, D E-Mail: Bernd.Niemann@chiru.med.uni-giessen.de
Tel.: +49 (0) 641 - 98556233 Fax: +49 (0) 641 - 98544309
Rheinsberg, Zoe
Universität Tübingen Institut für Ethik und Geschichte der Medizin Gartenstr. 47
72074 Tübingen, D E-Mail : zoe.rheinsberg@uni-tuebingen.de Tel. : +49 (0) 7071 – 2978032
Rudolph, Lenhard Fli Leibniz-Institut für Alternsforschung Fritz-Lipmann-Institut e.V Beutenbergstraße 11 07745 Jena, D
E-Mail: lenhard.rudolph@leibniz-fli.de
Tel.: +49 (0) 3641 – 656000 Sekretariat
117
List of Speakers S - T
Schnitzer-Ungefug, Jutta Deutsche Akademie der Naturforscher Leopoldina e. V. Nationale Akademie der Wissenschaften Jägerberg 1 06108 Halle (Saale) E-Mail: jutta.schnitzer-ungefug@leopoldina.org Tel.: 0345-47239912
Sebekova, Katarina Institute of Molecular Biomedicine Medical Faculty Comenius University Sasinkova 4 811 08 Bratislava Slovakia E-Mail: kata.sebekova@gmail.com Tel.: +421 (0) 2 59357 429 mobile:+421 (0) 907 794 093
Simm, Andreas Universitätsklinik und Poliklinik für Herzchirurgie Ernst-Grube-Str. 40 D-06120 Halle (Saale) E-Mail: andreas.simm@uk-halle.de Tel.: +49 (0) 345 – 557 2647 Fax: +49 (0) 345 – 557 7070
Treede, Hendrik
Universitätsklinik und Poliklinik für Herzchirurgie
Ernst-Grube-Straße 40 D-06120 Halle (Saale) E-Mail : hendrik.treede@uk-halle.de Tel. : +49 (0) 345 – 557 2720
118
Organizers
Treede, Hendrik Department of Cardiac Surgery Ernst-Grube-Straße 40 D-06120 Halle (Saale) E-Mail: hendrik.treede@uk-halle.de Tel.: +49 (0) 345 – 557 2720 Fax: +49 (0) 345 – 557 2782
Simm, Andreas Department of Cardiac Surgery Ernst-Grube-Str. 40 D-06120 Halle (Saale) E-Mail: andreas.simm@uk-halle.de Tel.: +49 (0) 345 – 557 2647 Fax: +49 (0) 345 – 557 7070
119
Involved Partners
German National Academy of Sciences Leopoldina Jägerberg 1 D-06108 Halle (Saale) E-Mail: leopoldina@leopoldina-halle.de Tel.: +49 (0) 345 - 47239 600 Fax: +49 (0) 345 - 47239 919 DGGG German Society of Gerontology and Geriatrics Geschäftsstelle Seumestr. 8 D-10245 Berlin E-Mail: gs@dggg-online.de Tel.: +49 (0) 30 - 52137 271 Fax: +49 (0) 30 - 52137 372
ProMoAge Martin-Luther-Universität Medizinische Fakultät Herzchirurgie Ernst-Grube-Str. 40 06120 Halle Telefon: +49 345/557-3041
IZAH Interdisciplinary Centre on Ageing Halle Ernst-Grube-Str. 40 D-06120 Halle E-Mail: andreas.simm@medizin.uni-halle.de Tel.: +49 (0) 345 - 557 2647 Fax: +49 (0) 345 - 557 7070
Friends and Sponsors of the Martin-Luther-University Halle-Wittenberg e.V. Universitätsplatz 10 D-06108 Halle E-Mail: ramona.mitsching@vff.uni-halle.de Tel.: +49 (0) 345 - 552 2912 Fax: +49 (0) 345 - 552 7076
120
Author Index Ale-Agha, N. 59, 60
Altschmied, Y. 59, 60, 62, 73
Amoah, A. 61
Andreas, L. 87
Antebi, A. 90
Aufenvenne, K. 62, 73
Balbach, J. 104
Baldensperger, T. 63, 80
Barthel, F. F. 75
Bertinat, R. 70
Beyer, A. 90
Bezold, V. 64
Bilova, T. 86
Birkemeyer, C. 86
Blauw, G. J. 95
Bork, K. 64, 68, 98, 99
Brandt, W. 86
Brauch, D. 86
Brown, A. 101
Butler-Browne, G. 95
Cabbia, A. 65
Campisi, J. 54
Cavinato, M. 66, 111
Chondrogianni, N. 22
Cifuentes, M. 70
de Grey, A. 38
de Jong, N. 95
de van der Schueren, M. A. E. 93, 94, 96
Demuth, H.-U. 18
Di Sanzo, S. 67
Ding, J. 85
Doves, M. 94
Eberlein, P. 75
Ehrhardt, T. 64, 68, 74, 79
Eifert, S. 32
Emini, R. 61
Emmert, M. 46
Ender, C. 69
Florian, C. 61
121
Forman, K. 70
Forné, I. 102
Franceschi, C. 8
Frank, F. 98, 99
Frolov, A. 86
Fülöp, T. 16
Gapeyeva, H. 95
Gebert, N. 89
Geiger, H. 61, 101
Giegling, I. 68, 74, 79
Glaubitz, N. 88
Glomb, M. A. 63, 76, 80
Gnanapragassam, V.S. 72, 98, 99
Gosch, M. 28
Goy, Ch. 60, 62, 73
Grabe, S. 71
Großmann, C. 78
Ground, I. R. 36
Grune, T. 20, 69
Grybel, K. J. 102
Haendeler, J. 30, 59, 60, 62, 73
Hartmann, C. 68, 74, 79
Hause, G. 85
Haybäck, J. 87
Heinemann, S. H. 82
Heiss, Ch. 62
Heller, R. 69
Henning, C. 76
Hermann, M. 66
Hoehenwarter, W. 104
Hoeijmakers, J. 56
Hofmann, B. 75, 88
Hogrel, J.-Y. 95
Hohmann, C. 76
Hollemann, Th. 85
Horstkorte, R. 64, 72, 75, 88, 98, 99
Huber, N. 77
Huber, O. 113
Hübschmann, R. 78
Hönicka, M. 61
Ignatov, A. 87
Imhof, A. 102
122
Jacobs, K. 88
Jakobs, Ph. 73
Jansen-Dürr, P. 66, 111
Jasper, H. 89
Jenewein, B. 66
Jonkman, N. H. 105, 107
Jung, J.-S. 71, 109
Jung, M. 64, 74, 79, 100, 109
Junk, A. 80
Kaether, Ch. 81
Kalinski, Th. 87
Keller, A. 61
Kirkland, J. 52
Kirkpatrick, J. 89
Klotz, L.-O. 81
Korzelius, J. 89
Koziel, R. 66
Kruizinga, R. C. 91
Kryeziu, N. 69
Kuldyushev, N. A. 82
Kummer, A. 80, 83
Köhnlein, K. 81
Langner, J. 84
Lepperdinger, G. 48
Liebold, A. 61
Liehr, K. 76
Liu, H. 85
Maier, A. 10, 91, 93, 94, 95, 96, 97, 105, 106, 107
Mamontova, T. 86
Martin, G. M. 34
Martinez, F. 70
McPhee, J. S. 95
Mece, O. 69
Meskers, C. G. M. 91, 93, 94, 95, 96, 97, 105, 106, 107
Milrud, S. 86
Mirelman, A. 14
Mittag, S. 113
Moskalev, A. 50
Munoz-Cánove, P. 44
Nagasundaram, M. 72
Narici, M. V. 95
123
Naß, N. 87
Nattamai, K. J. 101
Navarrete-Santos, Alex 71, 83, 88, 92, 109, 110
Navarrete-Santos, Anne 71, 102, 109
Niemann, B. 26
Niessen, M. 97
Nualart, F. 70
Numans, S. T. 91, 106
Ori, A. 89
Padvitski, T. 90
Pai, G. M. 69
Paudel, G. 86
Pendzialek, S. M. 102
Pfirrmann, Th. 85
Pierik, V. D. 91, 106
Pijnappels, M. 95, 97, 105, 107
Piskovatska, V. 92
Puls, A. 100
Quandt, D. 103
Reijnierse, E. M. 93, 94, 95, 96, 97
Rheinsberg, Z. 40
Rojer, A. G. M. 96, 97
Romani, N. 66
Rosenstock, Ph. 98
Rudolph, K. L. 42, 77
Ruhs, S. 78
Rujescu, D. 68, 74, 79, 100
Salazar, K. 70
Scheer, M. 72
Scheerman, K. 91, 106, 107
Scheffler, J. 99
Schiller, J. 79, 100
Schindler, M. 71, 102
Schmidt, R. 86
Schuetz, D. 101
Schönherr, R. 82
Sebekova, K. 24
Seeling, T. 71, 102
Seliger, B. 103
Shilyaev, N. 86
Simm, A. 12, 75, 80, 83, 88, 92, 99, 104, 110, 112, 113
124
Sinz, A. 86
Sipilä, S. 95
Smolikova, G. 86
Somoza, V. 110
Spannbrucker, T. 59
Steinbrenner, H. 81
Stenroth, L. 95
Tarakhovskaya, E. 86
Tissier, A. 86
Toma, G. 103
Trappenburg, C. 93, 94, 95, 96, 97
Treede, H. 75, 83, 110
Tressel, E. 105
Unfried, K. 59
Urazova, A. 104
Urban, N. 81
van Ancum, J. M. 91, 105, 106, 107
van Lummel, R. C. 95, 97
van Rijssen, N. M. 96
van Schoor, N. M. 105
van Schooten, K. S. 97
Ventura, N. 108
Verlaan, S. 91, 93, 106, 107
Villavicencio-Lorini, P. 85
Vogt, Th. 86
Volk, Ch. 109
von Ameln, F. 60
Walentek, P. 85
Wedel, S. 66, 111
Wessjohann, L. A. 86
Wienke, A. 75
Winterhalter, P. 80, 104, 112
Wächter, K. 104, 110
Ye, Z. 113
Zurek, M. 60
125
Thank’s to our Sponsors (in alphabetical order)
Bayer HealthCare 1,500 €
Biotronik Vertriebs GmbH & Co. KG 5,000 € Bristol-Myers Squibb 3.000 €
DFG up to 18,000 € Fumedica Medizintechnik GmbH 1,000 €
JR-herzchirurgische OP-Sets OHG 4,000 €
126
Map of Halle (Saale)
127
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