Accepted version

Review

Impact of supplementation with amino acids or their metabolites on muscle

wasting in patients with critical illness or other muscle wasting illness: A

systematic review

Liesl Wandrag a*, Stephen J Brett b, Gary Frost a, Mary Hickson a.

a Nutrition and Dietetic Research Group, Department of Investigative Medicine, Imperial College London, UK

b Centre for Peri-operative Medicine and Critical Care Research, Imperial College Healthcare NHS Trust, London UK

Keywords:

Critical illness, muscle wasting, branched-chain amino acid, essential amino acid supplement,

leucine, Beta-hydroxy-beta-methylbutyrate

*Correspondence: Liesl Wandrag, Charing Cross Hospital, Department of Nutrition & Dietetics, Laboratory Block 13th

Floor, Fulham Palace Rd, W6 8RF, London, UK.

Tel: +44 (0) 20 331 17844

Fax: +44 (0) 20 331 11440

Email: l.wandrag@imperial.ac.uk

Statement of Authorship

All authors have made substantial contributions to the concept and design of the systematic review,

with grading of the studies and interpretation of data, drafting of the article, revising it critically and

all have approved the final version for submission to the journal.

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Abstract

Background: Muscle wasting during critical illness impairs recovery. Dietary strategies to

minimise wasting include nutritional supplements, in particular, essential amino-acids. The aim of

this paper is to review the evidence on enteral supplementation with amino acids or their

metabolites in the critically ill and in muscle wasting illness with similarities to critical illness, to

assess whether this intervention could limit muscle wasting in vulnerable patient groups.

Methods: Citation databases including MEDLINE, Web of Knowledge, EMBASE, the meta-

register of controlled trials and the Cochrane Collaboration library were searched for articles from

1950 - 2013. Search terms included ‘critical illness’, ‘‘muscle wasting’, ‘amino acid

supplementation’,’ chronic obstructive pulmonary disease’, ‘chronic heart failure’, ‘sarcopenia’ and

‘disuse atrophy’. Reviews, observational studies, sport nutrition, intravenous supplementation and

studies in children were excluded.

Results: One hundred and eighty studies were assessed for eligibility, one hundred and fifty eight

were excluded. Twenty two studies were graded according to standardised criteria using the

GRADE methodology, four in critical care populations, and 18 from other clinically relevant areas.

Methodologies, interventions and outcome measures used were highly heterogeneous and meta-

analysis was not appropriate.

Conclusions: Methodology and quality of studies were too varied to draw any firm conclusion.

Dietary manipulation with leucine enriched essential amino acids (EAA), beta-hydroxy-beta-

methylbutyrate and creatine warrant further investigation in critical care; EAA has demonstrated

improvements in body composition and nutritional status in other groups with muscle wasting

illness. High quality research is required in critical care before treatment recommendations can be

made.

Introduction

Muscle wasting on the Intensive Care Unit (ICU) is of considerable concern with patients losing

between 1-2% of lean body mass per day (Gamrin et al. 1997;Plank and Hill 2000), resulting in

delayed recovery, and increased mortality and morbidity (Bienvenu et al. 2012;Herridge et al.

2011). Physical disability has been identified up to five years after ICU admission due to muscle

weakness (Herridge 2011). Muscle weakness acquired during critical illness has been independently

associated with increased hospital mortality (Ali et al. 2008). Muscle wasting is thought to

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contribute significantly to the muscle weakness experienced (Herridge 2011). Muscle wasting is a

multi-factorial phenomenon with the following factors contributing: the neuro-endocrine response

to trauma and critical illness; infection, sepsis and cytokine release (Soeters and Grimble 2009);

corticosteroid use; critical illness myopathy; nerve and neuromuscular junction changes;

polyneuropathy; disuse atrophy (prolonged sedation and paralysis) and inadequate nutrition

(Anzueto 1999;Griffiths and Hall 2010). Even with intense nutritional support muscle wasting still

occurs (Reid et al. 2004).

Preservation of muscle mass is clearly crucial for patient rehabilitation (NICE, 2009) however the

exact mechanisms involved with muscle wasting are still disputed and optimal nutritional support

on ICU has yet to be defined.

Various strategies to miniminse muscle wasting have been explored, Propranolol (a beta-blocker)

and Oxandrolone (a testosterone analogue) increased lean body mass and protein synthesis in

children after severe burns (Herndon et al. 2001;Herndon et al. 2012;Jeschke et al. 2007). Many

strategies have however failed to show direct benefit and some have proven to worsen outcome e.g.

increased patient mortality observed with growth hormone administration in critically ill patients

(Takala et al. 1999). Studies looking at early mobilisation of ventilated patients have demonstrated

feasibility although prospective studies will be required to evaluate the impact of such stategies on

muscle mass (Kress 2009). Electrical muscle stimulation (EMS) has shown initial promise in terms

of preservation of muscle mass (Gerovasili et al. 2009) however larger prospective studies will be

required to evaluate the exact role of EMS in muscle strength preservation (Parry et al. 2013).

Similarly numerous protein and amino acid preparations have been studied over the past decades

with various levels of success. Branched-chain amino acids (BCAA; valine, leucine, isoleucine)

were studied in the ICU population with some promise but studies often failed to address

background nutrition. Furthermore the ratio of BCAA used (valine: leucine: isoleucine of 1:1:1)

now appear sub-optimal (Cynober and Harris 2006). Importantly, leucine is known to promote

muscle protein synthesis by stimulating the mammalian target of rapamycin (mTOR) signalling

pathway involved with protein synthesis (Rennie et al. 2006). Studies using leucine

supplementation demonstrate muscle protein synthesis in elderly subjects (Anthony et al. 2001)

however although muscle protein synthesis was observed in young subjects this did not translate to

an overall gain in protein balance (Glynn et al. 2010). For populations with muscle wasting

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disorders, such as cachexia, beta-hydroxy-beta-methylbutyrate (HMB), a metabolite of leucine, has

shown some promise in restoring lean body mass (Clark et al. 2000;May et al. 2002).

Few amino acid supplementation studies exist in critical care, therefore conditions with similarities

to critical care have been assessed - these include studies in patients with chronic obstructive

pulmonary disease (COPD), chronic heart failure (CHF), age-related muscle wasting and bed rest

studies. Whilst COPD and CHF as chronic diseases clearly differ from critical illness as an acute

insult, many similarities in the pathophysiology of muscle wasting exist: muscle wasting during

critical illness may be initiated by systemic inflammation as suggested by raised C-reactive protein

(CRP), interleukin-6 (IL-6) and tumour necrosis factor alpha (TNF-α) levels (Callahan and Supinski

2009;McCarthy and Esser 2010;Winkelman 2010) whilst raised CRP, IL-6 and TNF-α levels have

also been reported in COPD patients with muscle wasting (Agusti et al. 2004;Jagoe and Engelen

2003;Yende et al. 2006) whilst systemic inflammation of both acute and chronic nature has been

described in COPD patients (Oudijk et al. 2003). Oxidative stress with raised levels of nitric oxide

and reactive oxygen species may contribute to muscle breakdown during critical illness (Dodd et al.

2010;McCarthy 2010) as well as in COPD patients (Oudijk 2003). Tissue hypoxia may also

contribute, particularly as the nuclear factor kappa-B (NF-κB) catabolic pathway may be activated

during hypoxia (de et al. 2011;McCarthy 2010), this pathway has also been found to be activated in

COPD patients with muscle wasting (Agusti 2004;Ladner et al. 2003;Oudijk 2003). Skeletal muscle

apoptosis or programmed cell death has been observed in COPD and CHF patients (Agusti et al.

2002) as well as in the critically ill (Attaix et al. 2005;Jespersen et al. 2011). Muscle fibre atrophy

and fibre type change have additionally been reported in the critically ill (Puthucheary et al. 2013),

in COPD patients (Gosker et al. 2003) and in CHF patients (Vescovo et al. 1996). Quantitatively

muscle wasting in the critically ill may be more accelerated, between 1-2% per day (Puthucheary

2013;Reid 2004), whilst reported as 20-40% during the disease process for COPD patients (Jagoe

2003).

Age-related muscle wasting, or sarcopenia, also share similarities to muscle wasting in critical

illness, least of all as the elderly are increasingly represented within critical care but also as

sarcopenia is characterised by diminished anabolic signals, promotion of catabolic signals, such as

pro-inflammatory cytokines, as well as the presence of oxidative stress and mitochondrial

dysfunction (Carmeli et al. 2002;Kamel 2003;Morley and Baumgartner 2004;Roubenoff and

Hughes 2000).

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Lastly, although the muscle wasting experienced during bed rest or immobility cannot directly be

compared to that of muscle wasting during critical illness, immobility during critical illness may

contribute to muscle wasting (Ferrando et al. 2006) and bed rest studies may provide a model to

study disuse atrophy as cortisol has been infused into healthy subjects undergoing bed rest to

simulate a stress response (Paddon-Jones et al. 2005).

Understanding which amino acid component might play a role in ameliorating muscle wasting

during critical illness may contribute to the successful rehabilitation of these patients. The purpose

of this paper is to review the literature focusing on enteral supplementation with amino acids and

amino acid metabolites in the critically ill or other muscle wasting illness with similarities to critical

illness, to assess whether this intervention can limit muscle wasting in vulnerable patient groups.

Methods

A literature search was conducted using Pubmed, MEDLINE, Web of Knowledge (Web of Science,

Biosis Citation Index and Journal Citation Reports), EMBASE, the meta-register of controlled trials

and the Cochrane Collaboration library database. Keywords used in the search were ‘critical

illness’, ‘critically ill patient’, ‘intensive care’, ‘chronic obstructive pulmonary disease’, ‘chronic

heart failure’, ‘sarcopenia’, ‘bed rest’, ‘disuse atrophy’, ‘muscle wasting’, ‘amino acid

supplementation’, ‘amino acid metabolite’, ‘enteral nutritional supplementation’, ‘muscle mass’,

‘lean mass’, ‘muscle breakdown’ and ‘muscle building’. Reference lists were hand searched. There

were no restrictions for language of publication provided an English abstract was available.

Databases were searched from 1950 to 1 December 2013. Indices of current clinical trials were also

accessed to identify current trials: Clinical trials (www.clinicaltrials.gov); current controlled trials

(www.controlled-trials.com) and the meta-register of clinical trials

(www.controlled-trials.com/mrct); INVOLVE via www.invo.org.uk/database and www.nihr.ac.uk.

Inclusion criteria: Interventional studies using enteral supplementation with amino acid or amino

acid metabolites in the adult critically ill, in patients with chronic obstructive pulmonary disease,

chronic heart failure, age-related muscle wasting (sarcopenia) or disuse atrophy (bed rest studies)

were included. Studies were included if the aim of the study was to improve nitrogen balance, lean

body mass and muscle function or strength with amino acid or metabolite supplementation.

Exclusion criteria: Papers were excluded if they were review papers; observational studies;

interventional studies focusing on multiple nutritional supplementation (e.g. combination products

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containing fish oils, anti-oxidants and amino acids); studies where amino acids were used for

immunonutrition purposes (i.e. glutamine); sport nutrition studies; intravenous supplementation

studies; studies in children; studies in young subjects not undergoing bed rest, studies including

cancer cachexia; liver disease; renal disease and HIV/AIDS. Molecular and mechanistic discussions

fall outside of the scope of this review.

Interventions: Enteral supplementation with amino acid and amino acid metabolites were included:

Leucine and beta-hydroxy-beta-methylbutyrate (HMB - a leucine metabolite), and mixed

preparations of amino acids, branched-chain amino acids or essential amino acids.

Outcome measures: These varied enormously and were placed into seven broad categories:

1. ICU/hospital outcomes: Length of stay; mortality and infectious outcomes including

antibiotic use.

2. Functional and strength testing: Step, walk and stair tests; lung function tests; activities of

daily living (ADL) scores; grip strength; quadriceps twitch force and muscle function tests.

3. Body composition: Dual energy x-ray absorptiometry (DEXA); weight, body mass index

(BMI); bioelectrical impedance analysis (BIA); computed tomography (CT); air

displacement techniques; mid arm circumference (MAC); triceps skinfold (TSF) and muscle

biopsies.

4. Nutritional: Mini nutritional assessment (MNA); resting energy expenditure (REE) via

indirect calorimetry; food intake charts and enteral feed tolerance.

5. Nitrogen balance/protein turnover: Plasma amino acids; protein turnover studies; urinary

studies including urinary nitrogen and 3-methyl histidine (3-MH).

6. Blood markers/inflammation: Inflammatory markers and cytokines; glucose; insulin.

7. Quality of Life (QoL): Mood questionnaires; depression scores; SF-36 and a respiratory

symptom questionnaire.

Quality Assessment: Studies were initially assessed by the lead investigator (LW) following the

GRADE methodology (Atkins et al. 2004) and subsequently consensus on inclusion, exclusion and

classification was achieved amongst the whole study group. Briefly, studies were initially graded

according to whether they were blinded, controlled, whether a placebo product was used and

whether they were adequately powered. Randomised controlled trials were given a high grade and

non-randomised studies a moderate or low grade. Subsequently, studies were marked down on

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limitations of methodology or risk of bias. If a study title contained the words ‘randomised study’

but randomisation was not described adequately in the methodology, the study was downgraded to

moderate.

Synthesis of results: Results were described using the ‘PICO’ framework (Population,

intervention, comparator, outcome), quality assessment of individual studies was performed by

using GRADE criteria. Results for ICU studies are presented in Table 1 and for non-ICU studies in

Table 2. Meta-analysis was not performed due to the heterogeneous populations, interventions and

outcome measures studied. Risk of bias was assessed per GRADE criteria based on study

randomisation, blinding and use of control group and placebo. The PRISMA 2009 checklist was

used to guide the design of this systematic review (Moher et al. 2009).

Results

One hundred and eighty articles were assessed for eligibility, 158 were excluded with reason.

Twenty two studies were included and graded. Four ICU studies included two studies in trauma

patients, one in general ICU patients and one in critically ill COPD patients. The 18 non-ICU

studies included 3 studies in patients with COPD, 1 study in chronic heart failure patients, 9 studies

in elderly subjects with sarcopenia, and 5 studies in subjects undergoing bed rest.

The process of study selection is shown in Figure 1.

Iden

tifica

tion

Scre

enin

gEl

igib

ility

Incl

uded

628 records identified through database searching

415 records after duplicates removed

415 records screened 235 excluded

180 full-text articles assessed for eligibility

158 excluded with reason

22 studies included and graded

4 ICU studies 18 non-ICU studies

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Figure 1: Process of study selection in accordance with PRISMA 2009 guidelines

ICU Studies

No ICU studies were graded as high quality. One study was classed as moderate quality (Kuhls et

al. 2007) and three as low quality (Hsieh et al. 2006;Jensen et al. 1996;Mansoor et al. 2007) . A

summary of each study is shown in Table 1.

Moderate Quality ICU Study

Kuhls et al studied 100 trauma ICU patients comparing HMB supplementation, HMB with

additional arginine and glutamine (HMB+) or an iso-caloric, iso-nitrogenous placebo product

(Kuhls 2007). The aim of this study was to assess whether HMB alone or HMB in combination with

arginine and glutamine had any effect on nitrogen balance and muscle breakdown. The HMB

supplement contained 3g HMB, HMB+ contained an additional 14g L-arginine and 14g L-

glutamine. The findings indicate that nitrogen balance improved with the intervention, the mean

nitrogen balance was -9, -10.9 and -6.5g/day for the control, HMB+, and HMB groups respectively

(p=0.05). Interestingly the HMB+ group appeared to remain in a more negative nitrogen balance

throughout the study (p<0.05), suggesting a possible detriment to added arginine or glutamine. The

3-MH and urinary nitrogen results combined suggest that supplementation did not affect muscle

breakdown. There were no significant differences between groups for mortality, 28 day antibiotic

use, inflammatory markers, hospital and ICU length of stay, ventilator days and number of new

infections. This study followed a randomised, double-blinded and placebo-controlled design,

however, the study did have some important limitations.

Although a sample size of N=100 was reported, only data for N=72 patients were analysed, leading

to a potential type II error. Twenty- eight subjects were excluded from statistical analysis as they

did not meet 50% compliance for supplementation. Additionally 3-MH was used as a sole marker

for muscle proteolysis despite some of its known limitations as a biomarker (i.e. not a sensitive

marker of specific tissue pools), whilst pre-albumin was used as a surrogate for protein synthesis.

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Despite the limitations HMB supplementation appeared to improve nitrogen balance in severely

injured trauma patients, and the addition of arginine and/or glutamine appears to neutralise any

benefit of HMB alone.

Low quality ICU studies

Twenty eight ICU patients were given EAA or non-EAA supplements over 10 days with the aim of

comparing plasma amino acid differences and nitrogen balance (Jensen 1996). The authors found

no clinical benefit with enteral glutamine-enriched EAA supplementation (no difference found in

immune function, plasma amino acid levels or in nitrogen balance).

Mansoor et al performed a study in 12 trauma ICU patients and 8 healthy controls (Mansoor 2007)

where supplementation with threonine, serine, cysteine and aspartic acid was assessed to determine

effects on protein metabolism. This unusual mixture of amino acids was selected based on the

authors’ previous work demonstrating a demand for these particular amino acids in septic rats

(reduced weight loss, muscle catabolism and an enhanced recovery) (Breuille et al. 2006). No

significant differences between the two ICU groups were found. This study was limited mainly by

the small sample size, lack of statistical power and short interventional period.

The final ICU study included 34 ventilated COPD patients given 3g/d HMB supplementation over 7

days to examine effects on inflammation, protein metabolism and pulmonary function (Hsieh 2006).

The authors report anti-catabolic effects however protein metabolism was not adequately assessed

in this study. This study was furthermore limited by small sample size with lack of statistical power

analysis, a short interventional period, non-randomised design and lack of placebo product. Thus,

although this data again points to the potential benefits of HMB, the design flaws limit the value of

the data.

Overall the data from ICU studies are very limited, but suggest that HMB may improve nitrogen

balance, although this improvement was marginal. Nevertheless, ICU is an extremely challenging

environment in which to carry out research and so an exploration of other clinically relevant areas

may be warranted.

Non-ICU Studies

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Eighteen studies were identified in non-ICU settings. No studies were graded high quality, three

were of moderate quality (Brooks et al. 2008;Fuld et al. 2005;Rondanelli et al. 2011) and fifteen

studies of low quality (Aquilani et al. 2008; Borsheim et al. 2008;Casperson et al. 2012;Dal Negro

et al. 2010;Dillon et al. 2009;Engelen et al. 2007;Ferrando et al. 2010;Freyssenet et al.

1996;Katsanos et al. 2005;Paddon-Jones et al. 2003;Paddon-Jones et al. 2004;Rieu et al. 2006;Stein

et al. 1999;Verhoeven et al. 2009;Volpi et al. 2003). A summary of each study is shown in Table 2.

Chronic Obstructive Pulmonary Disease

One moderate quality study and two low quality studies were included. In a moderate quality study

in 38 COPD patients undergoing pulmonary rehabilitation the use of 5.7g oral creatine monohydrate

(a metabolite of arginine, glycine, and methionine which is widely used in the exercise arena to

support muscle anabolism) was assessed on muscle function, strength, lung function and quality of

life (Fuld 2005). Creatine increased fat-free mass, lower limb strength and endurance in the post

loading and post rehabilitation phase. Limitations of this study include small sample size, high

drop-out rate (39% for the placebo group and 22% for creatine), and no assessments of nutrient

intake nor baseline nutritional status. Thus, creatine may offer an option for supplementation in ICU

patients that could be tested.

Muscle wasting is commonly observed in COPD patients, contributing to poor physical function.

Dal Negro et al assessed whether 4g EAA supplementation would improve body composition,

muscle metabolism, physical activity and health status in 32 COPD outpatients with sarcopenia (Dal

Negro 2010). Body weight and fat-free mass increased in the supplemented group (Table 2)

however p-values were not reported. The study is limited by small sample size and that dietary

intakes were not assessed. Nevertheless, interventions aimed at ameliorating muscle wasting in

COPD patients with sarcopenia may offer some suggestion of what intervention might minimise

muscle wasting during critical illness.

A soy test meal with additional BCAA was studied to assess differences in protein metabolism

between healthy elderly subjects and COPD patients (Engelen 2007). Whole body protein synthesis

increased in the group consuming soy + BCAA compared to soy feeding alone (p<0.05). This study

received a low grade as the sample size was small, the study period short and the study was non-

randomised, although a crossover design was followed.

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Chronic Heart Failure

One low quality study in CHF patients was included. Aquilani et al examined whether diet

combined with 4g EAA supplementation had a positive effect on nutritional and metabolic status of

chronic heart failure patients (Aquilani 2008). These patients are another group where muscle

wasting is common and related to hypoxia or systemic inflammation; in this study patients were

specifically selected for depleted muscle mass. An increase in weight and BMI was found in the

EAA group compared to the control group (p<0.01). Limitations to this study include small sample

size, lack of blinding, no placebo product used, the control group had higher baseline body weight,

and body composition was measured by anthropometric measures alone. This study used a higher

ratio of leucine within the BCAA composition (Table 2), which may require further investigation

when formulating future supplements to prevent muscle wasting.

Elderly

One moderate quality study and eight low quality studies were included. Rondanelli et al explored

whether 4g EAA supplementation twice daily would improve nutritional status, muscle function,

activity levels and health related quality of life in elderly institutionalised patients potentially

suffering from sarcopenia (Rondanelli 2011). Results showed that the EAA mixture improved

nutritional status, physical performance, muscle function and levels of depression significantly. This

was a well conducted study however sample size was small. Sarcopenia, although not identical to

ICU related muscle wasting, shares some similarities due to deranged anabolic signals and presence

of pro-inflammatory cytokines (Ershler and Keller 2000). Clearly this differs from the dramatic

muscle wasting observed in ICU but positive results in this group may offer clues to a useful

supplement in critical illness.

Verhoeven et al studied the effect of 2.5g leucine supplementation over 3 months on muscle mass

and strength in 29 healthy elderly men (Verhoeven 2009). No differences were observed after 3

months of supplementation for any of the outcome measures. This adds useful data showing that

leucine supplementation alone does not increase muscle mass or strength and may be most

beneficial when given in addition to the other branched-chain amino acids, but not as a sole

supplement (Cynober 2006).

Elderly females with sarcopenia were given a 7.5g EAA supplement over a three month period to

assess anabolic response (Dillon 2009). Muscle fractional synthesis rate and lean body mass were

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both significantly higher after three months of EAA supplementation compared to the placebo

(Table 2). This study was limited by small sample size however again shows that a BCAA

supplement with a higher ratio of leucine seems to have an effect on muscle synthesis. This study is

relevant to the ICU population because sarcopenia involves alterations in the mechanisms of muscle

breakdown and synthesis.

Volpi et al studied 14 healthy elderly subjects, 6 were given an 18g EAA supplement in 10 minute

boluses over 3 hours and 8 subjects were given 40g of EAA and non-EAA. Although both groups

demonstrated increased protein synthesis no difference was reported between the two groups (Volpi

2003).

Freyssenet et al looked at the combined effect of a BCAA supplement and endurance exercise on

skeletal muscle structural characteristics in elderly men with sarcopenia (Freyssenet 1996). The

supplement contained a high leucine component of 1:8:1 for the BCAA ratio. The number of

capillaries per fibre was the only significant change observed after the 6 week endurance exercise

programme, this was thought to be due to endurance exercise, not the BCAA supplementation. This

study was limited by small sample size and unclear randomisation.

Rieu et al studied subjects over 5h to assess the effect of a complete meal with or without additional

leucine on muscle protein synthesis and whole body protein metabolism in 20 healthy elderly men

(Rieu 2006). An increase in muscle fractional synthesis rate by 55% was observed in the leucine

group after 5 hours (p<0.05) with no change in whole body protein turnover observed. Limitations

to this study include a non-randomised design, small sample size and short study period.

Borsheim et al studied twelve elderly glucose intolerant subjects to assess whether 11g EAA +

arginine taken twice daily over 16 weeks would increase lean body mass, strength and functional

capacity (Borsheim 2008). Significant improvements in lean body mass (p=0.038), leg strength

(p<0.001), walking speed (p=0.002) and functional assessment (p=0.007) were seen after 16 weeks

on EAA + arginine. The leucine content in this supplement was high, with BCAA ratio of 1:5:1

(val: leu: iso). This study has several limitations: a non-randomised design, lack of blinding, lack of

a control group and a small sample size. Elderly glucose intolerant participants again cannot be

directly compared to ICU patients, however the anabolic effect of this supplement in insulin-

resistant participants may hint at strategies that could be explored in insulin- resistant ICU patients.

Casperson et al used 12g leucine supplementation to assess the effects on muscle protein synthesis

and mTOR signalling in elderly subjects (Casperson 2012). Two weeks of leucine supplementation

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significantly activated the mTOR signalling pathway (Day 1: 1.03 versus Day 15: 1.23

phosphorylated/total, p=0.01), demonstrating nutrient anabolic signalling, as well as increasing post

absorptive muscle protein synthesis (Day 1: 0.063 versus Day 15: 0.074%/h, p=0.004). Limitations

to this study include a non-randomised, single-blinded design, small sample size and no use of a

placebo product. The elderly are increasingly represented in the ICU population, interventions

shown to be effective in the elderly could potentially be explored in an ICU population.

Katsanos et al assessed whether elderly and young subjects responded differently to a bolus of 6.7g

EAA in terms of leg muscle protein synthesis (Katsanos 2005;Katsanos et al. 2006). The

investigators report lower muscle protein synthesis in the elderly than in the young (Table 2). The

methodology has several limitations: It was non-randomised, not blinded and included a small

sample size. Examining whether young and elderly participants differ in anabolic response to EAA

supplementation could broaden the applicability of the supplement use, particularly if this

intervention is explored in a heterogeneous ICU population.

Disuse atrophy

One moderate quality study and four low quality studies were included. Disuse atrophy studies are

pertinent to this topic since critically ill patients experience long periods of immobility, and despite

the fact that catabolism is far more severe and rapid in critical illness, this group may offer insights

into suitable supplement to test. In the moderate quality study Brooks et al compared the effects of a

daily 15g EAA supplement (alone, before or after resistance exercise) in healthy men undergoing a

28 day bed rest period to see if this combined treatment would affect muscle strength and body

composition (Brooks 2008). Results showed that the EAA supplementation along with resistance

training provided greater protection against muscle and strength loss during 28 days of bed rest. The

EAA-only group showed no protection against muscle or strength loss suggesting resistance

training is a critical part of the effective strategy (Table 2). Limitations of this study include small

sample size and no comparator group receiving resistance training only.

Ferrando et al examined whether 15g EAA in addition to a standard diet during 10 days of bed rest

would maintain muscle mass and function in elderly subjects (Ferrando 2010). The authors

concluded that muscle function may be preserved with EAA supplementation during periods of

compulsory bed rest (Table 2). Limitations to this study include a non-randomised design, single

blinding and small sample size. Disuse atrophy from bed rest studies again cannot be directly

13

compared to muscle wasting following critical illness, however these types of studies may provide

some clue as to potential interventions to minimise muscle wasting during critical illness.

In a further bed rest study 49.5g/day EAA with carbohydrate was examined for anabolic stimulus

during 28 days of bed rest (Paddon-Jones 2004). Muscle wasting appeared to be minimised with

this EAA supplement however strength was not fully preserved. This study was limited by small

sample size and an inadequate description of randomisation. This data supports that theory that the

ratio of BCAA should include higher levels of leucine compared to the other amino acids.

Stein et al conducted a bed rest study to assess whether 30mmol/d supplemental BCAA during 7

days of bed rest prevented nitrogen loss (Stein 1999). The authors observed higher nitrogen

retention in the BCAA group than in control subjects (62.5 8.0 mg/kg/d BCAA group versus 21.6

14.7 mg/kg/d controls, p<0.03). Limitations include small sample size, high dropout rate, a short

study period, exclusion of faecal and insensible losses from nitrogen balance calculations and the

lack of robust protein turnover work.

Paddon-Jones et al studied 12 healthy subjects (Paddon-Jones 2003) to examine whether a 27h

cortisol infusion (to simulate a stress response) changed muscle protein metabolism after a bolus

ingestion of 15g of EAA (Table 2). Anabolic response to EAA ingestion was increased during acute

hypercortisolemia. Whether this response will be observed after actual trauma would require further

investigation in a larger, prospective RCT with a much longer interventional period. This simulated

stress response again cannot directly be compared to ICU related muscle wasting, however these

studies may provide some pointers as to potential interventions to minimise muscle wasting

following critical illness.

The moderate quality studies suggest that improvements in nutritional status, body composition,

physical performance and muscle function with EAA and creatine supplementation are possible in

patient groups relevant to ICU muscle wasting, but considerable work is required to elucidate the

dose and BCAA ratio of EAA supplementation, more leucine seemed to be more beneficial. Two

studies indicate that resistance training is required along with EAA supplementation to protect

against muscle wasting. An important caveat is that the subjects in these studies will not have been

experiencing the degree of infective or inflammatory challenge commonly exhibited by ICU

patients.

14

Low quality studies suggests that EAA and BCAA supplements in COPD patients may increase

protein synthesis, body weight and lean body mass. In the elderly EAA may improve muscle

protein synthesis, muscle fractional synthesis rate, lean body mass and function and in participants

undergoing bed rest EAA may maintain weight and fractional synthesis rate whereas strength is

unlikely to be preserved. Leucine supplementation stimulated the mTOR pathway and increased

muscle protein synthesis in the elderly however leucine alone does not appear to protect against

muscle wasting.

Discussion

The evidence from ICU studies suggests that HMB supplementation may be the only amino acid

supplement tested to date that could lead to improvement in nitrogen balance in severely injured

trauma patients. However, since the pool of data is extremely limited further exploration of other

EAA ratio’s and doses is justified. Non-ICU studies suggest that EAA mixtures may well have a

beneficial effect on lean body mass, but the data for function and strength are less clear. Leucine, in

particular, appears to be the critical amino acid but the amount of leucine required in the

supplement remains to be elucidated, along with the formulation of the mixture of other amino acids

accompanying the leucine. This conclusion is also supported by the lower quality studies. As

alluded to earlier, the other diseases with similarities to ICU are used as they may inform the design

of future supplementation studies in ICU however the comparisons are provisional in nature.

The ratio of BCAA also appears to be important and higher ratios of leucine appear to be more

effective. Some of the studies reviewed used BCAA ratios as high as 1:5:1 and 1:8:1 as additional

leucine is thought to promote muscle protein synthesis (Anthony 2001;Breen and Phillips

2011;Crozier et al. 2005;Dickinson et al. 2011), however the results did not always support this

theory. Leucine, although known to promote muscle protein synthesis by stimulating the mTOR

signalling pathway, does not necessarily demonstrate a protein synthetic response in healthy

subjects. In the 3 month supplementation study authors commented on healthy elderly subjects’

mean daily protein intake (1g/kg/day) and how this exceeded that of the recommended daily protein

intake for the elderly (0.85g/kg/day). This may have led to already maximised muscle protein

synthesis in this group (Verhoeven 2009). Equally, young healthy subjects do not appear to

demonstrate net protein synthesis as the elderly do following leucine supplementation (Glynn

2010). Data on the potential muscle building capacity of leucine enriched essential amino acid

15

supplementation (BCAA ratio of 1:2:1 or higher), HMB and also creatine are still the most

intriguing of all the data presented.

Strengths of this review include the systematic approach followed, grading of each study to set

criteria and the comprehensive list of studies provided that could be used to integrate current

knowledge or to plan future interventional studies.

Since the data do not provide suitable evidence, no firm conclusions or treatment recommendations

can be made for supplementation in the critically ill. Future studies need to be adequately powered,

conducted in more homogeneous patient sub-groups in ICU, use clearly defined interventions which

control for baseline nutrition as well as providing supplementary EAA, and which are also provided

for long enough to assess clinical outcomes, as well as acute anabolic response or metabolic change.

Yet trying to influence clinical outcomes such as length of stay or mortality requires very large

sample sizes.

One on-going enteral amino acid supplementation study has been identified on the clinical trials

register in survivors of critical illness (study number: NCT01063738). This study is looking at EAA

supplementation during rehabilitation post critical illness, aiming to recruit 180 subjects and

examine the effect of both diet and physical rehabilitation in a randomised, single-blinded design.

Results from this study will add to current evidence.

Limitations

This review is limited by the published data available, in particular it was not possible to conduct

meta-analyses due to the heterogeneity of the published ICU studies. Many publications suffered

from poor methodological quality and/or reporting, particularly the process of randomisation, and

this fact means there is a risk of bias in this review.

In addition, bed rest studies and studies in the elderly, in patients with COPD and in patients with

CHF may not present the best models to study ICU related muscle wasting as the muscle wasting in

the critically ill is clearly so multi-faceted. However, these groups were included since the data may

offer pointers to the most promising interventions to investigate. Clearly more high quality research

in ICU patients is needed to identify whether nutritional supplementation can limit muscle wasting

and improve clinical outcomes.

16

Conclusion

Dietary manipulation with leucine enriched EAA, HMB or creatine warrant further investigation in

the critically ill and may offer a supportive strategy towards minimising muscle mass loss in these

vulnerable patients. Variability in quality and methodology of current evidence does not allow for

formulation of any clinical recommendations. Studies in other muscle wasting illnesses with

similarities to critical illness suggest that EAA mixtures may have a beneficial effect on lean body

mass, however the data for function and strength are less clear.

Acknowledgements

The authors acknowledge support from the Department of Health via the National Institute of

Health Research who supported this work through the provision of a Clinical Doctoral Research

Fellowship for LW. The research was further supported by the National Institute for Health

Research (NIHR) Biomedical Research Centre based at Imperial College Healthcare NHS Trust and

Imperial College London. The views expressed are those of the authors and not necessarily those of

the NHS, the NIHR or the Department of Health.

Conflict of interest and statement of authorship

The authors declare that there are no conflicts of interest. All authors have made substantial

contributions to the concept and design of the systematic review, with grading of the studies and

interpretation of data, drafting of the article, revising it critically and all have approved the final

version for submission to the journal.

17

References

Agusti, A., Morla, M., Sauleda, J., Saus, C., & Busquets, X. 2004. NF-kappaB activation and iNOS upregulation in skeletal muscle of patients with COPD and low body weight. Thorax, 59, (6) 483-487.

Agusti, A.G., Sauleda, J., Miralles, C., Gomez, C., Togores, B., Sala, E., Batle, S., & Busquets, X. 2002. Skeletal muscle apoptosis and weight loss in chronic obstructive pulmonary disease. Am.J.Respir.Crit Care Med., 166, (4) 485-489.

Ali, N.A., O'Brien, J.M., Jr., Hoffmann, S.P., Phillips, G., Garland, A., Finley, J.C., Almoosa, K., Hejal, R., Wolf, K.M., Lemeshow, S., Connors, A.F., Jr., & Marsh, C.B. 2008. Acquired weakness, handgrip strength, and mortality in critically ill patients. Am.J.Respir.Crit Care Med., 178, (3) 261-268.

Anthony, J.C., Anthony, T.G., Kimball, S.R., & Jefferson, L.S. 2001. Signaling pathways involved in translational control of protein synthesis in skeletal muscle by leucine. J.Nutr., 131, (3) 856S-860S.

Anzueto, A. 1999. Muscle dysfunction in the intensive care unit. Clin.Chest Med., 20, (2) 435-452.

Aquilani, R., Opasich, C., Gualco, A., Verri, M., Testa, A., Pasini, E., Viglio, S., Iadarola, P., Pastoris, O., Dossena, M., & Boschi, F. 2008. Adequate energy-protein intake is not enough to improve nutritional and metabolic status in muscle-depleted patients with chronic heart failure. Eur.J.Heart Fail., 10, (11) 1127-1135.

Atkins, D., Best, D., Briss, P.A., Eccles, M., Falck-Ytter, Y., Flottorp, S., Guyatt, G.H., Harbour, R.T., Haugh, M.C., Henry, D., Hill, S., Jaeschke, R., Leng, G., Liberati, A., Magrini, N., Mason, J., Middleton, P., Mrukowicz, J., O'Connell, D., Oxman, A.D., Phillips, B., Schunemann, H.J., Edejer, T.T., Varonen, H., Vist, G.E., Williams, J.W., Jr., & Zaza, S. 2004. Grading quality of evidence and strength of recommendations. BMJ, 328, (7454) 1490.

Attaix, D., Mosoni, L., Dardevet, D., Combaret, L., Mirand, P.P., & Grizard, J. 2005. Altered responses in skeletal muscle protein turnover during aging in anabolic and catabolic periods. Int.J.Biochem.Cell Biol., 37, (10) 1962-1973.

Bienvenu, O.J., Colantuoni, E., Mendez-Tellez, P.A., Dinglas, V.D., Shanholtz, C., Husain, N., Dennison, C.R., Herridge, M.S., Pronovost, P.J., & Needham, D.M. 2012. Depressive symptoms and impaired physical function after acute lung injury: a 2-year longitudinal study. Am.J.Respir.Crit Care Med., 185, (5) 517-524.

Borsheim, E., Bui, Q.U., Tissier, S., Kobayashi, H., Ferrando, A.A., & Wolfe, R.R. 2008. Effect of amino acid supplementation on muscle mass, strength and physical function in elderly. Clin.Nutr., 27, (2) 189-195.

Breen, L. & Phillips, S.M. 2011. Skeletal muscle protein metabolism in the elderly: Interventions to counteract the 'anabolic resistance' of ageing. Nutr.Metab (Lond), 8, 68.

18

Breuille, D., Bechereau, F., Buffiere, C., Denis, P., Pouyet, C., & Obled, C. 2006. Beneficial effect of amino acid supplementation, especially cysteine, on body nitrogen economy in septic rats. Clin.Nutr., 25, (4) 634-642.

Brooks, N., Cloutier, G.J., Cadena, S.M., Layne, J.E., Nelsen, C.A., Freed, A.M., Roubenoff, R., & Castaneda-Sceppa, C. 2008. Resistance training and timed essential amino acids protect against the loss of muscle mass and strength during 28 days of bed rest and energy deficit. J.Appl.Physiol, 105, (1) 241-248.

Callahan, L.A. & Supinski, G.S. 2009. Sepsis-induced myopathy. Crit Care Med., 37, (10 Suppl) S354-S367.

Carmeli, E., Coleman, R., & Reznick, A.Z. 2002. The biochemistry of aging muscle. Exp.Gerontol., 37, (4) 477-489.

Casperson, S.L., Sheffield-Moore, M., Hewlings, S.J., & Paddon-Jones, D. 2012. Leucine supplementation chronically improves muscle protein synthesis in older adults consuming the RDA for protein. Clin.Nutr., 31, (4) 512-519.

Clark, R.H., Feleke, G., Din, M., Yasmin, T., Singh, G., Khan, F.A., & Rathmacher, J.A. 2000. Nutritional treatment for acquired immunodeficiency virus-associated wasting using beta-hydroxy beta-methylbutyrate, glutamine, and arginine: a randomized, double-blind, placebo-controlled study. JPEN J.Parenter.Enteral Nutr., 24, (3) 133-139.

Crozier, S.J., Kimball, S.R., Emmert, S.W., Anthony, J.C., & Jefferson, L.S. 2005. Oral leucine administration stimulates protein synthesis in rat skeletal muscle. J.Nutr., 135, (3) 376-382.

Cynober, L. & Harris, R.A. 2006. Symposium on branched-chain amino acids: conference summary. J.Nutr., 136, (1 Suppl) 333S-336S.

Dal Negro, R.W., Aquilani, R., Bertacco, S., Boschi, F., Micheletto, C., & Tognella, S. 2010. Comprehensive effects of supplemented essential amino acids in patients with severe COPD and sarcopenia. Monaldi Arch.Chest Dis., 73, (1) 25-33.

de, T.C., Costes, F., Langen, R.C., Pison, C., & Gosker, H.R. 2011. Hypoxia and muscle maintenance regulation: implications for chronic respiratory disease. Curr.Opin.Clin.Nutr.Metab Care, 14, (6) 548-553.

Dickinson, J.M., Fry, C.S., Drummond, M.J., Gundermann, D.M., Walker, D.K., Glynn, E.L., Timmerman, K.L., Dhanani, S., Volpi, E., & Rasmussen, B.B. 2011. Mammalian target of rapamycin complex 1 activation is required for the stimulation of human skeletal muscle protein synthesis by essential amino acids. J.Nutr., 141, (5) 856-862.

Dillon, E.L., Sheffield-Moore, M., Paddon-Jones, D., Gilkison, C., Sanford, A.P., Casperson, S.L., Jiang, J., Chinkes, D.L., & Urban, R.J. 2009. Amino acid supplementation increases lean body mass, basal muscle protein synthesis, and insulin-like growth factor-I expression in older women. J.Clin.Endocrinol.Metab, 94, (5) 1630-1637.

19

Dodd, S.L., Gagnon, B.J., Senf, S.M., Hain, B.A., & Judge, A.R. 2010. Ros-mediated activation of NF-kappaB and Foxo during muscle disuse. Muscle Nerve, 41, (1) 110-113 available from: PM:19813194

Engelen, M.P., Rutten, E.P., De Castro, C.L., Wouters, E.F., Schols, A.M., & Deutz, N.E. 2007. Supplementation of soy protein with branched-chain amino acids alters protein metabolism in healthy elderly and even more in patients with chronic obstructive pulmonary disease. Am.J.Clin.Nutr., 85, (2) 431-439.

Ershler, W.B. & Keller, E.T. 2000. Age-associated increased interleukin-6 gene expression, late-life diseases, and frailty. Annu.Rev.Med., 51, 245-270.

Ferrando, A.A., Paddon-Jones, D., Hays, N.P., Kortebein, P., Ronsen, O., Williams, R.H., McComb, A., Symons, T.B., Wolfe, R.R., & Evans, W. 2010. EAA supplementation to increase nitrogen intake improves muscle function during bed rest in the elderly. Clin.Nutr., 29, (1) 18-23.

Ferrando, A.A., Paddon-Jones, D., & Wolfe, R.R. 2006. Bed rest and myopathies. Curr.Opin.Clin.Nutr.Metab Care, 9, (4) 410-415.

Freyssenet, D., Berthon, P., Denis, C., Barthelemy, J.C., Guezennec, C.Y., & Chatard, J.C. 1996. Effect of a 6-week endurance training programme and branched-chain amino acid supplementation on histomorphometric characteristics of aged human muscle. Arch.Physiol Biochem., 104, (2) 157-162.

Fuld, J.P., Kilduff, L.P., Neder, J.A., Pitsiladis, Y., Lean, M.E., Ward, S.A., & Cotton, M.M. 2005. Creatine supplementation during pulmonary rehabilitation in chronic obstructive pulmonary disease. Thorax, 60, (7) 531-537.

Gamrin, L., Andersson, K., Hultman, E., Nilsson, E., Essen, P., & Wernerman, J. 1997. Longitudinal changes of biochemical parameters in muscle during critical illness. Metabolism, 46, (7) 756-762.

Gerovasili, V., Stefanidis, K., Vitzilaios, K., Karatzanos, E., Politis, P., Koroneos, A., Chatzimichail, A., Routsi, C., Roussos, C., & Nanas, S. 2009. Electrical muscle stimulation preserves the muscle mass of critically ill patients: a randomized study. Crit Care, 13, (5) R161.

Glynn, E.L., Fry, C.S., Drummond, M.J., Timmerman, K.L., Dhanani, S., Volpi, E., & Rasmussen, B.B. 2010. Excess leucine intake enhances muscle anabolic signaling but not net protein anabolism in young men and women. J.Nutr., 140, (11) 1970-1976.

Gosker, H.R., Kubat, B., Schaart, G., van der Vusse, G.J., Wouters, E.F., & Schols, A.M. 2003. Myopathological features in skeletal muscle of patients with chronic obstructive pulmonary disease. Eur.Respir.J., 22, (2) 280-285.

Griffiths, R.D. & Hall, J.B. 2010. Intensive care unit-acquired weakness. Crit Care Med., 38, (3) 779-787.

Herndon, D.N., Hart, D.W., Wolf, S.E., Chinkes, D.L., & Wolfe, R.R. 2001. Reversal of catabolism by beta-blockade after severe burns. N.Engl.J.Med., 345, (17) 1223-1229.

20

Herndon, D.N., Rodriguez, N.A., Diaz, E.C., Hegde, S., Jennings, K., Mlcak, R.P., Suri, J.S., Lee, J.O., Williams, F.N., Meyer, W., Suman, O.E., Barrow, R.E., Jeschke, M.G., & Finnerty, C.C. 2012. Long-term propranolol use in severely burned pediatric patients: a randomized controlled study. Ann.Surg., 256, (3) 402-411.

Herridge, M.S., Tansey, C.M., Matte, A., Tomlinson, G., az-Granados, N., Cooper, A., Guest, C.B., Mazer, C.D., Mehta, S., Stewart, T.E., Kudlow, P., Cook, D., Slutsky, A.S., & Cheung, A.M. 2011. Functional disability 5 years after acute respiratory distress syndrome. N.Engl.J.Med., 364, (14) 1293-1304.

Hsieh, L.C., Chien, S.L., Huang, M.S., Tseng, H.F., & Chang, C.K. 2006. Anti-inflammatory and anticatabolic effects of short-term beta-hydroxy-beta-methylbutyrate supplementation on chronic obstructive pulmonary disease patients in intensive care unit. Asia Pac.J.Clin.Nutr., 15, (4) 544-550.

Jagoe, R.T. & Engelen, M.P. 2003. Muscle wasting and changes in muscle protein metabolism in chronic obstructive pulmonary disease. Eur.Respir.J.Suppl, 46, 52s-63s.

Jensen, G.L., Miller, R.H., Talabiska, D.G., Fish, J., & Gianferante, L. 1996. A double-blind, prospective, randomized study of glutamine-enriched compared with standard peptide-based feeding in critically ill patients. Am.J.Clin.Nutr., 64, (4) 615-621.

Jeschke, M.G., Finnerty, C.C., Suman, O.E., Kulp, G., Mlcak, R.P., & Herndon, D.N. 2007. The effect of oxandrolone on the endocrinologic, inflammatory, and hypermetabolic responses during the acute phase postburn. Ann.Surg., 246, (3) 351-360.

Jespersen, J.G., Nedergaard, A., Reitelseder, S., Mikkelsen, U.R., Dideriksen, K.J., Agergaard, J., Kreiner, F., Pott, F.C., Schjerling, P., & Kjaer, M. 2011. Activated protein synthesis and suppressed protein breakdown signaling in skeletal muscle of critically ill patients. PLoS.One., 6, (3) e18090.

Kamel, H.K. 2003. Sarcopenia and aging. Nutr.Rev., 61, (5 Pt 1) 157-167.

Katsanos, C.S., Kobayashi, H., Sheffield-Moore, M., Aarsland, A., & Wolfe, R.R. 2005. Aging is associated with diminished accretion of muscle proteins after the ingestion of a small bolus of essential amino acids. Am.J.Clin.Nutr., 82, (5) 1065-1073.

Katsanos, C.S., Kobayashi, H., Sheffield-Moore, M., Aarsland, A., & Wolfe, R.R. 2006. A high proportion of leucine is required for optimal stimulation of the rate of muscle protein synthesis by essential amino acids in the elderly. Am.J.Physiol Endocrinol.Metab, 291, (2) E381-E387.

Kazi, A.A., Pruznak, A.M., Frost, R.A., & Lang, C.H. 2011. Sepsis-induced alterations in protein-protein interactions within mTOR complex 1 and the modulating effect of leucine on muscle protein synthesis. Shock, 35, (2) 117-125.

Kress, J.P. 2009. Clinical trials of early mobilization of critically ill patients. Crit Care Med., 37, (10 Suppl) S442-S447.

21

Kuhls, D.A., Rathmacher, J.A., Musngi, M.D., Frisch, D.A., Nielson, J., Barber, A., MacIntyre, A.D., Coates, J.E., & Fildes, J.J. 2007. Beta-hydroxy-beta-methylbutyrate supplementation in critically ill trauma patients. J.Trauma, 62, (1) 125-131.

Ladner, K.J., Caligiuri, M.A., & Guttridge, D.C. 2003. Tumor necrosis factor-regulated biphasic activation of NF-kappa B is required for cytokine-induced loss of skeletal muscle gene products. J.Biol.Chem., 278, (4) 2294-2303.

Mansoor, O., Breuille, D., Bechereau, F., Buffiere, C., Pouyet, C., Beaufrere, B., Vuichoud, J., Van't Of, M., & Obled, C. 2007. Effect of an enteral diet supplemented with a specific blend of amino acid on plasma and muscle protein synthesis in ICU patients. Clin.Nutr., 26, (1) 30-40.

May, P.E., Barber, A., D'Olimpio, J.T., Hourihane, A., & Abumrad, N.N. 2002. Reversal of cancer-related wasting using oral supplementation with a combination of beta-hydroxy-beta-methylbutyrate, arginine, and glutamine. Am.J.Surg., 183, (4) 471-479.

McCarthy, J.J. & Esser, K.A. 2010. Anabolic and catabolic pathways regulating skeletal muscle mass7. Curr.Opin.Clin.Nutr.Metab Care, 13, (3) 230-235.

Moher D, Liberati A, Tetzlaff J, Altman DG, The PRISMA Group (2009). Preferred Reporting Items for Systematic Reviews and Meta-Analyses: The PRISMA Statement. PLoS Med 6(6): e1000097.

Morley, J.E. & Baumgartner, R.N. 2004. Cytokine-related aging process. J.Gerontol.A Biol.Sci.Med.Sci., 59, (9) M924-M929.

National Institute for Health and Care Excellence (NICE), 2009. Rehabilitation after critical illness, (CG83); http://guidance.nice.org.uk/CG83.

Oudijk, E.J., Lammers, J.W., & Koenderman, L. 2003. Systemic inflammation in chronic obstructive pulmonary disease. Eur.Respir.J.Suppl, 46, 5s-13s.

Paddon-Jones, D., Sheffield-Moore, M., Creson, D.L., Sanford, A.P., Wolf, S.E., Wolfe, R.R., & Ferrando, A.A. 2003. Hypercortisolemia alters muscle protein anabolism following ingestion of essential amino acids. Am.J.Physiol Endocrinol.Metab, 284, (5) E946-E953.

Paddon-Jones, D., Sheffield-Moore, M., Urban, R.J., Aarsland, A., Wolfe, R.R., & Ferrando, A.A. 2005. The catabolic effects of prolonged inactivity and acute hypercortisolemia are offset by dietary supplementation. J.Clin.Endocrinol.Metab, 90, (3) 1453-1459.

Paddon-Jones, D., Sheffield-Moore, M., Urban, R.J., Sanford, A.P., Aarsland, A., Wolfe, R.R., & Ferrando, A.A. 2004. Essential amino acid and carbohydrate supplementation ameliorates muscle protein loss in humans during 28 days bedrest. J.Clin.Endocrinol.Metab, 89, (9) 4351-4358.

22

Parry, S.M., Berney, S., Granger, C.L., Koopman, R., El-Ansary, D., & Denehy, L. 2013. Electrical muscle stimulation in the intensive care setting: a systematic review. Crit Care Med., 41, (10) 2406-2418.

Plank, L.D. & Hill, G.L. 2000. Similarity of changes in body composition in intensive care patients following severe sepsis or major blunt injury. Ann.N.Y.Acad.Sci., 904, 592-602.

Puthucheary, Z.A., Rawal, J., McPhail, M., Connolly, B., Ratnayake, G., Chan, P., Hopkinson, N.S., Padhke, R., Dew, T., Sidhu, P.S., Velloso, C., Seymour, J., Agley, C.C., Selby, A., Limb, M., Edwards, L.M., Smith, K., Rowlerson, A., Rennie, M.J., Moxham, J., Harridge, S.D., Hart, N., & Montgomery, H.E. 2013. Acute skeletal muscle wasting in critical illness. JAMA, 310, (15) 1591-1600.

Reid, C.L., Campbell, I.T., & Little, R.A. 2004. Muscle wasting and energy balance in critical illness. Clin.Nutr., 23, (2) 273-280.

Rennie, M.J., Bohe, J., Smith, K., Wackerhage, H., & Greenhaff, P. 2006. Branched-chain amino acids as fuels and anabolic signals in human muscle. J.Nutr., 136, (1 Suppl) 264S-268S.

Rieu, I., Balage, M., Sornet, C., Giraudet, C., Pujos, E., Grizard, J., Mosoni, L., & Dardevet, D. 2006. Leucine supplementation improves muscle protein synthesis in elderly men independently of hyperaminoacidaemia. J.Physiol, 575, (Pt 1) 305-315.

Rondanelli, M., Opizzi, A., Antoniello, N., Boschi, F., Iadarola, P., Pasini, E., Aquilani, R., & Dioguardi, F.S. 2011. Effect of essential amino acid supplementation on quality of life, amino acid profile and strength in institutionalized elderly patients. Clin.Nutr., 30, (5) 571-577.

Roubenoff, R. & Hughes, V.A. 2000. Sarcopenia: current concepts. J.Gerontol.A Biol.Sci.Med.Sci., 55, (12) M716-M724.

Soeters, P.B. & Grimble, R.F. 2009. Dangers, and benefits of the cytokine mediated response to injury and infection. Clin.Nutr., 28, (6) 583-596.

Stein, T.P., Schluter, M.D., Leskiw, M.J., & Boden, G. 1999. Attenuation of the protein wasting associated with bed rest by branched-chain amino acids. Nutrition, 15, (9) 656-660.

Takala, J., Ruokonen, E., Webster, N.R., Nielsen, M.S., Zandstra, D.F., Vundelinckx, G., & Hinds, C.J. 1999. Increased mortality associated with growth hormone treatment in critically ill adults. N.Engl.J.Med., 341, (11) 785-792.

Verhoeven, S., Vanschoonbeek, K., Verdijk, L.B., Koopman, R., Wodzig, W.K., Dendale, P., & van Loon, L.J. 2009. Long-term leucine supplementation does not increase muscle mass or strength in healthy elderly men. Am.J.Clin.Nutr., 89, (5) 1468-1475.

Vescovo, G., Serafini, F., Facchin, L., Tenderini, P., Carraro, U., Dalla, L.L., Catani, C., & Ambrosio, G.B. 1996. Specific changes in skeletal muscle myosin heavy chain

23

composition in cardiac failure: differences compared with disuse atrophy as assessed on microbiopsies by high resolution electrophoresis. Heart, 76, (4) 337-343.

Volpi, E., Kobayashi, H., Sheffield-Moore, M., Mittendorfer, B., & Wolfe, R.R. 2003. Essential amino acids are primarily responsible for the amino acid stimulation of muscle protein anabolism in healthy elderly adults. Am.J.Clin.Nutr., 78, (2) 250-258.

Winkelman, C. 2010. The role of inflammation in ICU-acquired weakness. Crit Care, 14, (4) 186.

Yende, S., Waterer, G.W., Tolley, E.A., Newman, A.B., Bauer, D.C., Taaffe, D.R., Jensen, R., Crapo, R., Rubin, S., Nevitt, M., Simonsick, E.M., Satterfield, S., Harris, T., & Kritchevsky, S.B. 2006. Inflammatory markers are associated with ventilatory limitation and muscle dysfunction in obstructive lung disease in well functioning elderly subjects. Thorax, 61, (1) 10-16.

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Table Legend

Table 1: Amino acid supplementation – ICU studies

Table 2: Amino acid supplementation – Non-ICU studies

25