PLENARNESEKCIJE

PLENARY SESSIONS

Sekcija 1

BIOHEMIJSKIMARKERI

OBOLJENJA

Session 1

BIOCHEMICALMARKERS OF THE DISEASES

JMB 2008; 27 (2) 175

POTENCIJAL SR^ANIH MARKERA ZAUNAPRE\ENJE LE^ENJA PACIJENATA

C. Miler

Odeljenje interne medicine, Univerzitetska bolnica, Bazel, [vajcarska

B-tip natriuretskog peptida (BNP) jeste kvantita-tivni marker za sr~ane bolesti. Pokazalo se da upotre-ba BNP-a kod pacijenata sa dispneom unapre|ujele~enje i smanjuje tro{kove tretmana. Dodatne indi-kacije koje imaju potencijal da unaprede le~enje pa-cijenata uklju~uju pra}enje toka le~enja u slu~ajevi-ma akutnih i hroni~nih sr~anih bolesti, plu}ne embo-lije i koronarne arterijske bolesti.

NAPREDAK U LE^ENJU SR^ANIH BOLESNIKA ZAHVALJUJU]I

UPOTREBI SR^ANIH MARKERA

B. Hercig

Centar za razvoj kardiologije, Vizbaden, Nema~ka

Sr~ani markeri omogu}uju dijagnostikovanje ikarakterizaciju sr~anih oboljenja. Visokoosetljivi testza troponin specifi~an je za infarkt miokarda, dok jemieloperoksidaza mo}na alatka za rano otkrivanjenastanka plaka. Homocistein je povezan sa brojnimneurodegenerativnim bolestima, kardiovaskularnimdoga|ajima a naro~ito {logom. Na osnovu povi{enihkoncentracija u plazmi mo`e se predvideti ishod imogu se prepoznati grupe sa visokim rizikom kojimabi pomogle prevencija i terapija. Homocistein sepokazao kao dragocen marker za rano dijagnostiko-vanje deficita folata i vitamina B koji su uklju~eni uanemije i brojne hroni~ne bolesti. U ovom radu pred-stavljeni su inovativni marker MPO, dobro poznatimarker troponin i vi{estruko korisni marker homocis-tein, i obja{njen je njihov zna~aj u le~enju sr~anihbolesnika.

THE POTENTIAL OF CARDIAC MARKERS TO IMPROVE PATIENT MANAGEMENT

C. Müller

Departement of Internal Medicine, University Hospital Basel, Basel, Switzerland

B-type natriuretic peptide (BNP) is a quantita-tive marker for heart failure. The use of BNP in pa-tients with dyspnea has consistently shown to impro-ve patient management and reduce treatment cost.Additional indications with the potential to improvepatient management include treatment monitoring inacute and chronic heart failure, pulmonary embo-lism, and coronary artery disease.

IMPROVED CARDIAC PATIENTMANAGEMENT THROUGH CARDIAC

MARKERS

B. Herzig

Cardiac Area Business Development, Wiesbaden, Germany

Cardiac markers enable the diagnosis and char-acterization of cardiac diseases. While the highly sen-sitive troponin assay is specific for myocardial infarc-tion, myeloperoxidase is a powerful for early determi-nation of plaque formation. Homocysteine is inde-pendently associated with numerous neurodegenera-tive diseases, cardiovascular events and stroke in par-ticular. Elevated plasma concentrations predict out-come and help identify high-risk groups most likely tobenefit from prevention and therapy. Homocysteineturned out as a valuable marker for early diagnosesof folate and B-vitamin deficiencies that are involvedin anemias and numerous chronic diseases. This arti-cle presents the innovative marker MPO, the wellestablished marker troponin and the versatile anduseful marker homocysteine, and explains theirimportance in cardiac patient management.

UDK 577.1 : 61 ISSN 1452-8258

JMB 27: 175–178, 2008 Plenarne sekcije

Plenary sessions

BONE MARKERS – THEIR NATURE AND CLINICAL USE

M. Theis

Roche Diagnostics GmbH Sandhofer Strasse 116

68305 Mannheim, Germany

Bone remodeling units are the centerpiece ofbone metabolism. They are fueled by a synchronizedand well balanced interaction of osteoclasts andosteoblasts, the activity of which releases specificsubstances known as bone markers into the blood.Resorption markers result from osteoclastic activity,formation markers from osteoblastic activity, andturnover markers from both cell types. In clinicalpractice, bone markers are today widely used formonitoring of anti-resorptive therapy and patientcompliance. There is strong evidence that they arealso useful for risk assessment with respect to osteo-porosis, here complementing established imagingmethods. Other possible and partly not yet investi-gated indications include monitoring of side-effectsof certain therapeutic drugs and oncology. In partic-ular the combination of resorption and formationmarkers may open up a more differentiated insightinto the metabolic situation of a patient's bone. Theactivity of osteoclasts and osteoblasts is triggered andmodulated by numerous factors, some of which areof endocrine nature. Easily measurable in today's lab-oratory are for instance PTH, calcitonin and vitaminD. While calcitonin is not widely used in osteology,PTH and vitamin D define risk factors for an acceler-ated loss of bone and impaired mineralization ofosteoid with the related diseases of osteoporosis,ricketts and osteomalacia. Recent developments inlab diagnosis of bone diseases focus on rheumaticdiseases like rheumatoid arthritis, where anti-CCP isa much more specific marker than the commonrheuma factors.

ADVANCES IN THE GENETICBASIS OF ISCHEMIC STROKE

S. Stankovi}, N. Majki}-Singh

Institute for Medical Biochemistry, University School of Pharmacy and

Clinical Center of Serbia, Belgrade, Serbia

As one of the leading causes of death within boththe developed and developing world, stroke is a world-wide problem. About 80% of strokes are ischemic. It iscaused by multiple genetic factors, environmental fac-tors, and interactions among these factors. There is along list of candidate genes that have been studied fora possible association with ischemic stroke. Among themost widely investigated genes are those involved in

KO[TANI MARKERI– PRIRODA I KLINI^KA UPOTREBA

M. Teis

Roche Diagnostics GmbH Sandhofer Strasse 116

68305 Mannheim, Nema~ka

Jedinice za ko{tano remodelovanje su sredi{nata~ka metabolizma kostiju. Pokre}e ih sinhronizovanai uravnote`ena interakcija osteoklasta i osteoblasta,~ija aktivnost izaziva otpu{tanje u krv specifi~nih sup-stanci poznatih pod imenom ko{tani markeri. Markeriresorpcije su rezultat aktivnosti osteoklasta, markeriformacije rezultat su aktivnosti osteoblasta, a markeriprometa mogu nastati od obe vrste }elija. U klini~kojpraksi se ko{tani markeri danas koriste za pra}enjetoka antiresorptivne terapije i odgovora pacijenta naterapiju. Postoje i ~vrsti dokazi o tome da su korisni zaprocenu rizika kad je re~ o osteoporozi, gde su kom-plementarni ustanovljenim »imaging« metodama.Ostale mogu}e i donekle jo{ nedovoljno istra`eneindikacije uklju~uju pra}enje ne`eljenih efekata nekihlekova i onkologiju. Naro~ito kombinacija markeraresorpcije i formacije mo`e omogu}iti bolje diferenci-rani uvid u metaboli~ko stanje kosti pacijenta. Aktiv-nosti osteoklasta i osteoblasta pokre}u i moduli{ubrojni faktori, od kojih su neki endokrine prirode. Udana{njim laboratorijama se lako mogu izmeriti npr.PTH, kalcitonin i vitamin D. Kalcitonin se ne primen-juje mnogo u osteologiji, ali PTH i vitamin D defini{ufaktore rizika za ubrzani gubitak ko{tane mase i naru-{enu mineralizaciju osteoida, uz srodne bolesti osteo-porozu, rahitis i osteomalaciju. Novija dostignu}a izoblasti laboratorijske dijagnostike fokusirana su nareumatske bolesti poput reumatoidnog artritisa, gdeje anti-CCP mnogo specifi~niji marker od obi~nihreumatskih faktora.

NOVINE NA POLJU GENETIKE ISHEMIJSKOG MO@DANOG UDARA

S. Stankovi}, N. Majki}-Singh

Institut za medicinsku biochemiju, Farmaceutski fakultet i Klini~ki centar Srbije,

Beograd, Srbija

Kao jedan od vode}ih uzroka smrtnosti kako urazvijenim tako i u zemljama u razvoju, mo`dani udarje svetski problem. Oko 80% svih mo`danih udara ~iniishemijski mo`dani udar. U patogenezi mo`danog uda-ra va`nu ulogu imaju genetski faktor, faktori sredine injihova interakcija. Veliki broj gena-kandidata povezu-je se sa ishemijskim mo`danim udarom. Geni kandi-dati koji se ispituju u mo`danom udaru dele se u neko-

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liko grupa u zavisnosti od toga da li uti~u na hemo-stazu, inflamaciju, sintezu azot monoksida, metaboli-zam homocisteina i lipida, renin-angiotenzim-aldo-steron sistem. Kombinovanjem linkid`/asocijacionihstudija utvr|eno je da geni koji kodiraju PDE4D iALOX5AP doprinose riziku od dobijanja mo`danogudara. U daljem tekstu bi}e prikazani rezultati studijakoje su se bavile ispitivanjem ovih gena, a koji bi moglida imaju potencijalnu primenu u ranoj dijagnostici,prevenciji i le~enju pacijenata sa ishemijskom bole{}umozga.

haemostasis, inflammation, nitric oxide production,homocysteine and lipid metabolism, renin-angioten-sin-aldosterone system. Combined linkage/associationstudies have demonstrated that genes encodingPDE4D and ALOX5AP confer risk for stroke. We re-view the studies of these genes which may have poten-tial application on the early diagnosis, prevention andtreatment ischemic stroke patients.

JMB 2008; 27 (2) 177

Sekcija 2

PRIMENAINDIKATORAKVALITETA U MEDICINSKIM

LABORATORIJAMA

Session 2

APPLICATION OF QUALITYINDICATORS IN A MEDICAL LABORATORY

»IZVESNOST U NEIZVESNOM SVETU« – STAV JEDNOG KLINI^ARA

O SENZITIVNOSTI I PRECIZNOSTI

P. Tumej

Odeljenje za klini~ku biohemiju, Bolnica Ipsvi~, Safok, Ujedinjeno Kraljevstvo

Klini~ka praksa evoluira kao i istra`ivanja. Sli~notome, analiti~ke tehnologije bivaju unapre|ene svakegodine. S pravom ili ne, klini~ari sve vi{e pa`nje prida-ju takvim ispitivanjima na u{trb klini~ke istorije i pregle-da. Kako se ljudski vek produ`ava, u porastu je i preva-lenca dugotrajnih stanja kao {to su bolest {titne `lezde,kardiovaskularna oboljenja i maligniteti. Testovi zasno-vani na klini~koj biohemiji imaju va`nu ulogu u tretira-nju (skriningu, dijagnostikovanju, prognozi i pra}enju)takvih stanja. U Ujedinjenom Kraljevstvu je to postaloo~igledno 2004. godine, od kada u primarnoj nezipreko 100 od 550 klini~kih zaklju~aka zavisi od rezul-tata testova iz oblasti klini~ke biohemije. Rezultati semogu razlikovati izme|u testova usled odstupanja, pre-ciznosti, specifi~nosti i senzitivnosti testa. Do danas semalo zna~aja pridaje potencijalnom klini~kom efektupreciznosti. Ova prezentacija ispita}e efekte kojepreciznost testa mo`e imati na tretiranje va`nih dugo-trajnih stanja kao {to su bolest {titne `lezde, kardio-vaskularna oboljenja i maligniteti.

GRE[KE U LABORATORIJSKOJ MEDICINI I KAKO IH IZBE]I

O. Sonntag

Ortho-Clinical Diagnostics, Neckargemünd, Nema~ka

Medicinsko tuma~enje klini~kih laboratorijskihpodataka je komparativni proces odlu~ivanja u komse rezultat laboratorijskog testiranja pojedinca poredisa referentnim intervalom izvedenim od referentnepopulacije. ^ini se da ova re~enica opisuje jednos-tavnu situaciju. Bi}e predstavljen detaljniji prikaz otome {ta se de{ava sa uzorkom pacijenta i klini~kimuzorkom sa uvidom u laboratoriju. Bi}e prikazane ne-ke grani~ne vrednosti koje poti~u direktno sa uzorkai od pacijenta. Nedostatak znanja o kvalitetu uzorka i

»CERTAINTY IN AN UNCERTAIN WORLD« – A CLINICIAN’S VIEWPOINT

OF SENSITIVITY AND PRECISION

P. Twomey

Clinical Biochemistry, The Ipswich Hospital, Suffolk IP4 5PD, United Kingdom

Clinical practice is evolving as research evolvesfrom the bench to the bedside. Similarly, analyticaltechnologies are improving on an annual basis. Rightlyor wrongly, increased emphasis is now placed by clini-cians on such investigations to the detriment of clinicalhistory and examination. As people live longer, the pre-valence of long-term conditions such as thyroid disea-se, cardiovascular disease and malignancies is increas-ing. Clinical biochemistry assays play an important partin the management (screening, diagnosis, prognosisand monitoring) of such conditions. This is reflected inthe UK since 2004 by the primary care contract whereover 100 of the 550 clinical points depend on clinicalbiochemistry assay results. Inter-assay results may dif-fer due to bias, precision, assay specificity and assaysensitivity. To date, little emphasis has been placed onthe potential clinical effect of precision. This presenta-tion will explore the effect that assay precision can haveon the management of important long-term conditionssuch as thyroid disease, cardiovascular disease andmalignancies.

ERRORS IN LABORATORY MEDICINE AND WAYS TO AVOID THEM

O. Sonntag

Ortho-Clinical Diagnostics, Neckargemünd, Germany

The medical interpretation of clinical laboratorydata is a comparative decision-making process inwhich a laboratory test result for an individual is com-pared with a reference interval derived from a refer-ence population. This sentence seems to describe aneasy situation. A closer look to what happens to apatient and to a clinical sample will be discussed inview of the laboratory. One will learn about some lim-its arriving direct from the sample and the patient. Alack of knowledge about the quality of the sample

JMB 2008; 27 (2) 181

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JMB 27: 181–184, 2008 Plenarne sekcije

Plenary sessions

and the special issues with biological material createsa gap between the laboratory and the clinician. Tomeasure quality control material is important for theanalytical process. The major concerns in this fieldare the missing real sample and the commutability.We do quality control with artificial samples and hopethey will mimic real patient material. We will movemuch closer to the issues around the sample and itsquality. With a lot of examples from literature andown investigation one will learn more about:– use and misuse of reference intervals– statistical presentation and misleading interpreta-

tion,– stability of the analyte in the sample,– volume displacement effect,– lipemia, turbidity, hemolysis and icterus,– calibration,– special cases of drug interferences.

Finally, we have to better investigate the sampleand the preanalytical process to provide reliableresults. The consultation is an important tool thatopens a field of communication and will help us tobring the laboratory in a new view to the clinician. Amodern analytical tool – known as POCT – offers onlya short term solution and will not cover the pre- andpostanalytical part. Who takes care of the quality ofthe sample in this field? There is a strong need toshow the value of a laboratory that understands itscustomers. A rapid and reliable test result combinedwith a comment or a consultation will offer the valueof the laboratory’s work. Knowledge and sharing theknowledge will result in high quality laboratory workto provide safer patient care.

DILEMMAS AND CONTROVERSIES IN INTERPRETATION

OF LABORATORY RESULTS

D. Popovi}1, M. Popovi}2

1Centre for Clinical-Laboratory Diagnostics,2Clinic of ENT and MFS,

Clinical Centre of Montenegro, Podgorica, Montenegro

Concentration of many substances in blood is agood indicator of the physiologic state of a patient. Itis usual that results obtained represent the real con-centration of tested substances of a patient, that is,they represent his physiological state. The influence ofsome factors indicates that this assumption is notalways true. Mistakes owing to analytic factors arereduced to the least possible rate by using the qualitycontrol. Also, many nonanalytic factors can change theconcentration of one or more substances of the sam-ple, so the results obtained are not the indicator of thephysiological state of the patient. Results of clinical-biochemical determination are interpreted by compar-ing with the referential values, and so the conclusion is

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posebnim problemima u vezi sa biolo{kim materi-jalom stvaraju jaz izme|u laboratorije i klini~ara. Zaanaliti~ki proces bitno je izmeriti materijal za kontrolukvaliteta. Glavni problem u toj oblasti je nedostatakpravog uzorka i mogu}nosti razmene. Kontrolu kva-liteta vr{imo uz pomo} ve{ta~kih uzoraka u nadi da}e se oni pona{ati kao pravi materijal dobijen odpacijenta. Bi}e detaljnije obrazlo`ena pitanja u vezi sauzorkom i njegovim kvalitetom. Uz pomo} mno{tvaprimera iz literature i na osnovu sopstvenog istra`i-vanja, bi}e prikazana saznanja o:– upotrebi i pogre{noj upotrebi referentnih intervala,– statisti~koj prezentaciji i varljivom tuma~enju,– stabilnosti analitika u uzorku,– uticaju smanjenja zapremine,– lipemiji, zamu}enju, hemolizi i ikterusu,– kalibraciji,– posebnim slu~ajevima interferencije lekova.

Najzad, moraju se bolje ispitati uzorak i procespred analizu da bi se obezbedili pouzdani rezultati.Konsultacija je va`na alatka koja omogu}ava boljukomunikaciju i poma`e nam da kod klini~ara stvo-rimo potpuno novu sliku o laboratoriji. Moderna ana-liti~ka alatka – poznata pod nazivom POCT – nudisamo kratkoro~no re{enje i ne pokriva deo pre i posleanalize. Ko se brine o uzorku o kvalitetu uzorka u tojoblasti? Postoji velika potreba za pokazivanjem vred-nosti laboratorije koja razume svoje klijente. Brz ipouzdan rezultat testiranja u kombinaciji sa komen-tarom ili konsultacijom prikazuje vrednost laboratorij-skog rada. Znanje i distribucija znanja obezbe|ujubezbedniju negu pacijenata uz pomo} visokokva-litetnog laboratorijskog rada.

DILEME I KONTROVERZE U TUMA^ENJULABORATORIJSKIH NALAZA

D. Popovi}1, M. Popovi}2

1Centar za klini~ko-laboratorijsku dijagnostiku, 2Klinika za ENT i MFS, Klini~ki centar Crne Gore,

Podgorica, Crna Gora

Koncentracija mnogih supstanci u krvi je dobarodraz fiziolo{kog stanja pacijenta. Uobi~ajeno je dadobijeni rezultati predstavljaju stvarnu koncentracijuispitivane supstance kod pacijenta, odnosno da pred-stavljaju njegovo fiziolo{ko stanje. Uticaj nekoliko fak-tora ukazuje da ova pretpostavka nije uvijek ta~na.Gre{ke zbog analiti~kih faktora, svode se na najmanjumogu}u mjeru primjenom kontrole kvaliteta. Tako|e,mnogi neanaliti~ki faktori mogu mijenjati koncen-traciju jedne ili vi{e supstanci u uzorku, tako da dobi-jeni rezultati nijesu odraz fiziolo{kog stanja pacijenta.Rezultati klini~ko-biohemijskih odre|ivanja interpretira-ju se pore|enjem sa referentnim vrijednostima, pa se izaklju~ak donosi metodom pore|enja. Da bi ovaj pro-

ces mogao pravilno da se izvede potrebno je da pos-toje referentne vrijednosti za svaki odre|ivani pa-rametar. Cikli~ne varijacije, fizi~ki napor, stres i drugifaktori imaju zna~ajan uticaj na dobijene vrijednostianaliza. Pri tuma~enju rezultata ove specifi~nosti mora-ju biti uzete u obzir, jer }e u protivnom biti protu-ma~ene kao patolo{ke, {to navodi na pogre{an zaklju-~ak. Neadekvatna priprema pacijenta za odre|enuanalizu i nepo{tovanje pravila koja se odnose na pri-premu i analiziranje uzorka, mogu dovesti do drasti-~nog odstupanja rezultata od stvarne vrijednosti. Iz tihrazloga pri tuma~enju rezultata nastaju odre|ene di-leme i kontroverze.

STATUS RAZVOJA I IMPLEMENTACIJEAKREDITACIJE MEDICINSKIH

LABORATORIJA U SRBIJI

Lj. Gligi}

Akreditaciono telo Srbije, Beograd, Srbija

Objavljivanjem standarda SRPS ISO 15189:2008»Medicinske laboratorije: posebni zahtevi za kvalitet ikompetentnost« stvoreni su osnovni preduslovi za nje-govu primenu u Srbiji. Primena standarda ISO 15189danas je prihva}eni mehanizam za pobolj{anje kvalitetausluga medicinskih laboratorija u Evropi. Na taj na~insu harmonizovani do tada razli~iti pristupi pobolj{anjakvaliteta medicinskih laboratorija. Funkcionalna organi-zacija procesa akreditacije medicinskih laboratorija uve}ini evropskih zemalja sprovodi se kroz saradnjunacionalnih akreditacionih tela, medicinskih eksperatadelegiranih iz nau~nih dru{tava i resornih ministarstava.Takav tip saradnje pokazao se uspe{nim u Velikoj Brita-niji, Nema~koj, Ma|arskoj, Francuskoj, Hrvatskoj i dr.Na{e iskustvo u akreditaciji medicinskih laboratorijaprema standardu SRPS ISO/IEC 17025 (akreditovanoje pet laboratorija) i pozitivno iskustvo evropskih zemal-ja u procesu akreditacije su osnova za razvoj programaakreditacije medicinskih laboratorija u Srbiji. Prvi koraku tom nastojanju je formiranje komisije, sastavljene odeksperata iz razli~itih oblasti medicine, stru~njaka ATS-a i predstavnika nadle`nog ministarstva, i definisanjenjenih zadataka, kao {to su: izrada potrebne doku-mentacije, kvalifikacionih kriterijuma i programa obukeocenjiva~a, u~estvovanje u izradi {eme eksterne proce-ne kvaliteta preko me|ulaboratorijskih ispitivanja, pra-}enje rada evropskih organizacija za akreditaciju, orga-nizovanje zajedni~kih ocenjivanja sa doma}im i stranimocenjiva~ima i u~estvovanje u odlu~ivanju prilikom do-deljivanja akreditacije.

JMB 2008; 27 (2) 183

made by comparing method. In order to perform thisprocess properly, referential values for each specificparameter are necessary to exist. Cyclic variations,physical activity, stress, and other factors significantlyaffect on obtained result analyses. In interpretation ofresults, these specificities have to be considered, oth-erwise they will be interpreted as pathologic, whichleads to wrong conclusion. Inadequate preparation ofa patient for a certain analysis and disrespect of rulesreferring preparation and sample analyzing, can leadto drastic deviation of results from the real values.From those reasons there are certain dilemmas andcontroversies in the result interpretation.

STATUS OF DEVELOPMENT AND IMPLEMENTATION OF MEDICAL

LABORATORY ACCREDITATION IN SERBIA

Lj. Gligi}

Accreditation Board of Serbia, Belgrade, Serbia

Through the release of the standard SRPS ISO15189:2008 entitled »Medical Laboratories –ParticularRequirements for Quality and Competence«, condi-tions have been created for the accreditation of me-dical laboratories in Serbia. The application of the ISO15189 standard is an accepted mechanism for improv-ement of the quality of medical laboratory services inthe EU today. In that way, different approaches to thequality improvement of medical laboratories have beenharmonised. Functional organisation of the accre-ditation process of medical laboratories in most Euro-pean countries is mainly carried out in cooperation withnational accreditation bodies, medical experts appoint-ed by scientist associations and the Ministry of Health.This type of collaboration has proven successful in theUnited Kingdom, Germany, Hungary, France, Croatia,etc. Experiences of the Accreditation Board of Serbia(ABS) in the accreditation of medical laboratoriesaccording to the SRPS ISO/IEC 17025 standard (5laboratories have been accredited) and the positiveexperiences of the European countries in the accredi-tation process constitute the basis for the developmentof a program of medical laboratory accreditation inSerbia. The first step in this direction is the formationof a committee consisting of experts from differentfields of medicine, ABS experts and representatives ofthe competent ministry, and the definition of theirtasks, such as: preparation of nessesary documenta-tion, qualification criteria and training programs forassessors, participation in the development of the sche-me of external quality assessment through interlabora-tory testing, monitoring the work of the Europeanorganisations for accreditation, organisation of mutualassessments with national and international assessorsand participation in decision making during accredita-tion.

Sekcija 3

BIOHEMIJSKIMARKERI

OBOLJENJA

Session 3

BIOCHEMICALMARKERS OF THE DISEASES

BIOHEMIJSKI MARKERI ATEROSKLEROZE

M. \eri}, S. Koji}-Damjanov, V. ^abarkapa, N. Eremi}

Centar za laboratorijsku medicinu, Klini~ki centar Vojvodine, Novi Sad, Serbia

U ovom radu razmatrani su samo neki lipidniparametri i serumski markeri inflamacije u pogledunjihove prediktivne povezanosti s aterosklerotskombole{}u. Nastavlja se debata o zna~aju merenja razli-~itih lipida i lipoproteina, uklju~uju}i koncentracijuLDL ~estica i nivoe apolipoproteina. Tako|e, nisuuspostavljene preporuke za apolipoprotein (a) feno-tipizaciju i druge lipidne markere. Poslednjih godinapreporu~uje se simultano merenje nekoliko markerai izra~unavanje lipidnih indeksa kao {to su lipid tetra-da index (LTI), lipid pentada index (LPI) i aterogeniindeks plazme (AIP). Nekoliko cirkuli{u}ih markerainflamacije, npr. C-reaktivni protein, serumski fibrino-gen i povi{enje broja leukocita, dosledno su udru`enis aterosklerozom. Iako nema dokaza za korisnost me-renja C-reaktivnog proteina u {iroj zajednici, formi-rane su preporuke za njegovu upotrebu u dijagnosti-ci i le~enju koronarne bolesti srca. Neki proinflama-torni citokini, adhezioni molekuli i markeri leukocitneaktivacije su obe}avaju}i markeri, ali zaslu`uju daljaprospektivna ispitivanja. Pitanje koje zahteva odgovorje i da li su ti inflamatorni markeri direktno uklju~eniu patogeni proces.

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BIOCHEMICAL MARKERS OF ATHEROSCLEROSIS

M. \eri}, S. Koji}-Damjanov, V. ^abarkapa, N. Eremi}

Department of Laboratory Medicine, Clinical Center of Vojvodina, Novi Sad, Serbia

This paper is a brief review of some lipid param-eters and serum markers of inflammation in a view oftheir predictive relevance for the atherosclerotic dis-ease. A discourse on the importance of measuringdifferent lipids and lipoproteins, concentration of LDLparticles and apolipoprotein levels is still underway.Also, the recommendations for apolipoprotein (a),phenotypization and other lipid markers have not yetbeen established. In recent years the recommenda-tions imply simultaneous measuring of multiplemarkers and calculating the lipid index values such aslipid tetrad index (LTI), lipid pentad index (LPI) andatherogenic index of plasma (AIP). Several circula-ting markers of inflammation such as C-reactive pro-tein, serum fibrinogen and elevated leukocyte num-ber, are consistently associated with atherosclerosis.In spite of a lack of evidence on measuring the C-reactive protein in a wide population, the guidelinesfor its application in diagnostics and therapy of coro-nary heart disease were developed. Some proinflam-matory cytokines, adhesion molecules and markersof leukocyte activation are promising markers, requir-ing, however, more detailed prospective evaluation.The question to be elucidated is if these inflammato-ry markers are directly involved in the pathogenicprocess.

UDK 577.1 : 61 ISSN 1452-8258

JMB 27: 187–194, 2008 Plenarne sekcije

Plenary sessions

INFLAMMATORY AND APOPTOTIC MARKERS IN ISCHEMIC HEART

DISEASE PATIENTS

V. B. \or|evi}1, T. Risti}2, V. ]osi}2, P. Vlahovi}2,L. Zvezdanovi}2, G. \or|evi}3

1Institute of Biochemistry, Faculty of Medicine, Ni{, Serbia

2Centre for Medical Biochemistry, Clinical Centre, Ni{, Serbia

3Clinic for Neurology, Clinical Centre, Ni{, Serbia

Ischemic heart disease is the most frequent causeof cardiovascular morbidity and mortality. It isdeveleped on the bases of atherosclerosis which is cur-rently considered a chronic inflammatory disease. It isdocumented by an increase in inflammatory andimmune biomarkers, such as C-reactive protein, fib-rinogen, neopterin, leukocytes, lymphocytes and oth-ers, that are significantly changed in patients withunstable angina or acute myocardial infarction. CRP ismostly studied. Increased concentrations of CRP areassociated with a series of risk factors. CRP may predictrecurrent events and mortality independently of cardiactroponin levels, and it is also independent predictor ofcardiovascular event after adjustment for traditional riskfactors. Although CRP currently appears to be the mostpromising biological marker, there is still controversyregarding its use in clinical practice. Although bothnecrotic and apoptotic cell death are documented dur-ing atherogenesis the limited data are available aboutapoptotic markers in ischemic heart disease patients.Increasing evidence supports the existence of apoptot-ic death initiated by ligation of membrane-bound deathreceptors or by release of cytochrome c from mito-chondria, as well as their regulators in the heart. Thestudies of serum markers show that apoptotic processis disregulated in ischemic heart disease patients.Tumor necrosis factor-related apoptosis-inducing ligand(TRAIL) is present in stable atherosclerotic lesions, isincreased in vulnerable plaques, but its serum levels arereduced significantly in patients with unstable angina.Serum Fas concentrations are increased and FasL aredecreased in subjects at high cardiovascular risk. Theresults of our study show significant changes of serumFas, FasL, and Bcl-2 concentrations, and lymphocytecaspase-3 activity in different stages of ischemic heartdisease. For now, there is evidence that statins areeffective in the regulation of some apoptotic markers.The better understanding of the pathways of apoptosisand their regulation is promissing in yielding novel ther-apeutic targets for cardiovascular disease.

MARKERI INFLAMACIJE I APOPTOZE KOD BOLESNIKA SA ISHEMIJSKOM

BOLE[]U SRCA

V. B. \or|evi}1, T. Risti}2, V. ]osi}2, P. Vlahovi}2,L. Zvezdanovi}2, G. \or|evi}3

1Institut za biohemiju, Medicinski fakultet, Ni{, Srbjia

2Centar za medicinsku biohemiju, Klini~ki centar, Ni{, Srbija

3Klinika za neurologiju, Klini~ki centar, Ni{, Srbija

Ishemijska bolest srca je naj~e{}i uzrok kardio-vaskularnog morbiditeta i mortaliteta. Razvija se naterenu ateroskleroze koja se danas smatra hroni~nominflamatornom bole{}u. Inflamacija je dokazana pra}e-njem inflamatornih i imunih biomarkera kao {to su C-reaktivni protein (CRP), fibrinogen, neopterin, leukociti,limfociti i drugi koji se zna~ajno menjaju kod bolesnikasa nestabilnom anginom pektoris ili akutnim infarktommiokarda. Naj~e{}e ispitivan marker je CRP. Pove}anekoncentracije CRP su udru`ene sa brojnim rizik faktori-ma. Na osnovu vrednosti CRP mogu se predvidetirekurentni doga|aji i mortalitet nezavisno od vrednostikardija~nog troponina, a tako|e je nezavisan prediktorkardiovaskularnog doga|aja nakon korekcije tradi-cionalnih rizik faktora. Iako se CRP trenutno smatrabiolo{kim markerom koji najvi{e obe}ava, jo{ uvek pos-toje kontroverzije u vezi sa njegovim kori{}enjem uklini~koj praksi. Mada je dokazano da se i nekroza iapoptoza de{avaju u toku aterogeneze malo je dostup-nih podataka koji se odnose na markere apoptoze kodbolesnika sa ishemijskom bole{}u srca. Sve je vi{e infor-macija koje ukazuju da se u srcu apoptoza kao i njenaregulacija odvijaju, i da se apoptoza mo`e iniciratipreko membranskih receptora smrti, ali i osloba|anjemcitohroma c iz mitohondrija. Ispitivanje serumskihmarkera je pokazalo da je proces apoptoze poreme}enkod bolesnika sa ishemijskom bole{}u srca. TRAIL(tumor necrosis factor-related apoptosis-inducing lig-and) je prisutan u stabilnim aterosklerotskim lezijama,pove}an u vulnerabilnim plakovima, dok su njegove se-rumske vrednosti zna~ajno sni`ene kod bolesnika sanestabilnom anginom. Koncentracija serumskog Fas jepove}ana, a FasL sni`ena kod osoba sa visokimrizikom. Rezultati na{e studije su pokazali zna~ajnepromene koncentracija Fas, FasL i Bcl-2 u serumu, kaoi aktivnost kaspaze-3 u limfocitima u razli~itim sta-dijumima ishemijske bolesti srca. Za sada, postojepodaci o efektivnosti statina u regulaciji nekih markeraapoptoze. Bolje razumevanje puteva apoptoze i njihoveregulacije mo`e omogu}iti nov terapijski pristup kardio-vaskularnim bolestima.

188

IZOFORME APOPROTEINA(A)I KONCENTRACIJA LP(A) U PLAZMI

KOD ^LANOVA ISTE PORODICE

DD. Labudovi}, KN. Toseska-Trajkovska, SB.Alabakovska, JJ. Bogdanska, BB. Todorova

Centar za medicinsku i eksperimentalnu biohemiju,Medicinski fakultet, Skoplje, Makedonija

Apoprotein(a) je multikringel protein u kojem jeuo~ljiv genetski nasledan polimorfizam veli~ine. Huma-ni gen APO(a) nalazi se u telomeri~kom regionu hro-mozoma 6. Polimorfizam veli~ine apo(a) glavna jedeterminanta nivoa Lp(a). Cilj studije je da opi{e uticajveli~ine apo(a) na nivoe Lp(a) u plazmi tri porodice. K-EDTA je dobijen od svakog ~lana porodice posle celo-no}nog posta. Izoforme apo(a) odre|ene su uz pomo}3–15% SDS-PAGE-a, a zatim tehnikom »Western imu-noblot«. Nivoi Lp(a) kvantifikovani su imunonefelome-trijski. Svako dete nasledilo je jednu izoformu od majkea drugu od oca. Deca iz prve porodice nasledila su obeizoforme apo(a) od oba roditelja i kod njih su nivoiLp(a) bili sli~ni onima koji su izmereni kod njihovihroditelja. ]erke iz druge i tre}e porodice nasledile sudominantnu maj~inu izoformu S3 apo(a), kao i maj~invisok nivo Lp(s). Odredili smo nivoe Lp(a) u plazmi kod~lanove druge porodice: kod majke 32,3 mg/dl, oca<9,6 mg/dl i }erke 36 mg/dl, {to je vi{i nivo Lp(a)nego kod njene majke. Kod ~lanova tre}e porodice,otac je bio nosilac »nula« fenotipa, majka je imaladominantan S3 i izmerili smo veoma visok nivo Lp(a)(74,1 mg/dl) u njenoj krvi; obe }erke imale su ni`enivoe Lp(a) – 46,5 mg/dl i 44,6 mg/dl. Na osnovu tihnalaza zaklju~ili smo da se nasle|ivanje izoformi apo(a)odvija prema Mendeljejevom sistemu. Tako|e smootkrili generacijsko sni`avanje nivoa Lp(a) kod dece utre}oj porodici.

ISPITIVANJE IZOFORMI APOLIPOPROTEINA(A) I KONCENTRACIJE LIPOPROTEINA (A)

U KORONARNOJ BOLESTI

N. Bogavac-Stanojevi}1, Z. Jeli}-Ivanovi}1, V. Spasojevi}-Kalimanovska1, S. Spasi}1, L. Memon2, D. Kalimanovska-O{tri}3

1Institut za medicinsku biohemiju,Farmaceutski fakultet, Beograd

2Klini~ka laboratorija, Klini~ko-bolni~ki centar »Be`anijska kosa«, Beograd

3Institut za kardiovaskularne bolesti, Klini~ki centar Srbije, Beograd

Povi{ena koncentracija lipoproteina (a) Lp(a) iprisustvo malih izoformi apolipoproteina (a) apo(a) u

JMB 2008; 27 (2) 189

APOPROTEIN(A) ISOFORMS AND PLASMALP(A) CONCENTRATION AMONG MEMBERS

OF THE SAME FAMILY

DD. Labudovi}, KN. Toseska-Trajkovska, SB. Alabakovska, JJ. Bogdanska, BB. Todorova

Department of Medical and ExperimentalBiochemistry, Faculty of Medicine, Skopje, Macedonia

Apoprotein(a) is a multikringle protein whichshows a genetically inherited size polymorphism. Thehuman APO(a) gene is located at the telomeric regionof chromosome 6. Apo(a) size polymorphism is a majordeterminant of Lp(a) levels. The aim of this study is toreport the influence of apo(a) size on the plasma Lp(a)levels in 3 families. K-EDTA was obtained from everymember of the family after an over night fast. Apo(a)isoforms were determined by 3–15% SDS-PAGE fol-lowed by Western immunoblot technique. Plasma Lp(a)levels were quantified immuneonephelometrically.Every child inherited one isoform from its mother andthe other from its father. The children from the firstfamily inherited the apo(a) isoform each from both par-ents and had Lp(a) levels similar to those measured intheir parents. The daughters from the second and thirdfamily inherited the dominant mother s S3 apo(a) iso-form, and also mother's high Lp(a) levels, respectively.We have determined plasma Lp(a) levels among themembers of the second family: mother 32.3 mg/dl,father <9.6 mg/dl and daughter 36 mg/dl, which washigher than Lp(a) level determined in her mother.Among the members of the third family, the father wasnull phenotype carrier, mother had a dominant S3 andwe measured a very high Lp(a) level (74.1 mg/dl) inher blood; both her daughters had lower Lp(a) levels –46.5 mg/dl and 44.6 mg/dl respectively. On the basisod these findings we concluded that the inheritance ofapo(a) isoforms occurrs according to the Mendeleevsystem; also, we found a generation decrease of theLp(a) level in the children of the third family.

EXAMINATION OF APOLIPOPROTEIN (A) ISOFORMS AND LIPOPROTEIN (A) CONCENTRATION IN CORONARY

ARTERY DISEASE

N. Bogavac-Stanojevi}1, Z. Jeli}-Ivanovi}1, V. Spasojevi}-Kalimanovska1, S. Spasi}1, L. Memon2, D. Kalimanovska-O{tri}3

1Institute of Medical Biochemistry, University School of Pharmacy, Belgrade

2Laboratory of Biochemistry, Medical Centre »Be`anijska kosa«, Belgrade

3Institute of Cardiovascular Diseases, Clinical Centre of Serbia, Belgrade

Elevated lipoprotein (a) Lp(a) levels and smallerapolipoprotein (a) apo(a) isoforms play an important

role in atherosclerosis. There were no data regardingapo(a) isoforms within the Serbian population and coro-nary artery disease (CAD) patients. Therefore, the aimof the study was to examine apo(a) isoform distributionand the impact of apo(a) isoforms on Lp(a) concentra-tion and on the severity of CAD. Two hundred and sev-enty nine CAD patients and 280 healthy subjects wereenrolled in this study. Patients were categorized into 4groups based on the extent of CAD, assessed by coro-nary angiography. Group 1 included patients with <50% luminal narrowing on major vessels. Groups 2, 3and 4 included patients with luminal stenosis 50% in 1,2, or 3 major arteries. Apo(a) isoform analysis was per-formed using sodium dodecyl sulphate agarose gelelectrophoresis followed by immunoblotting. The plas-ma concentration of Lp(a) was measured usingimmunoturbidimetry. In both patients and controls thedistribution of Lp(a) concentrations was skewed towardsa lower level. In contrast, the distribution of apo(a) iso-forms in both populations was bimodal. Lp(a) concen-trations correlated negatively with apo(a) isoforms inboth controls (R=–0.592, P<0.001) and patients(R=–0.553, P<0.001). CAD patients compared tohealthy subjects, showed significantly higher levels ofLp(a) (106.1 mg/L vs. 167.5 mg/L, p<0.001). Apo(a)isoforms were significantly smaller in CAD patients,compared to healthy subjects (p<0.001). The differentnumber of stenosed vessels was related to increasedLp(a) concentrations, without (p=0.009) and withadjustment (p=0.001) for apo(a) isoforms. However,apo(a) isoforms were not significantly associated withthe extent of CAD. This study shows that apo(a) iso-forms are the primary factor associated with the varia-tion in Lp(a) concentration. Apo(a) isoforms were relat-ed to CAD, but not to the severity of CAD.

PON1 POLYMORPHISM IN PATIENTS WITH ANGIOGRAPHICALLY ASSESSED

CORONARY ARTERY DISEASE

J. Kotur-Stevuljevi}, S. Spasi}, A. Stefanovi}, V. Spasojevi}-Kalimanovska, Z. Jeli}-Ivanovi}

Institut of Medical Biochemistry, Faculty of Pharmacy, Belgrade

Traditional cardiovascular disease (CVD) risk-fac-tors such as hypercholesterolemia, age, sex, hyperten-sion, diabetes, sedentary way of life, smoking, could bediagnosed in 50% of CVD patients. Coronary heart dis-ease pathogenesis in the other 50% of patients, with-out traditional risk-factor profiles, could be explainedwith a predominance of small dense LDL (low denselipoprotein) particles, increased oxidative LDL concen-trations, increased levels of homocysteine, oxidativestress and inflammation influence. It is very importantto explain the protective role of HDL particles; it is in

cirkulaciji imaju zna~ajnu ulogu u procesu ateroskle-roze. Po{to ne postoje podaci o veli~ini apo(a) za zdra-vu populaciju Srbije, niti za pacijente sa koronarnomarterijskom bole{}u (KAB), cilj ovog istra`ivanja je da seispita distribucija veli~ina apo(a), kao i uticaj veli~ineapo(a) na koncentraciju Lp(a) i na pojavu te`eg oblikastenoze na koronarnim krvnim sudovima (KKS). Ustudiju je uklju~eno 279 pacijenata sa KAB i 280 zdra-vih osoba. Pacijenti su na osnovu rezultata koronarneangiografije svrstani u 4 grupe. Grupa 1 je formiranaod pacijenata sa stenozom < 50% na KKS. Grupe 2,3 i 4 su formirane od pacijenata sa stenozom 50% na1, 2 ili 3 KKS. Izoforme apo(a) su odre|ivane natrijumdodecil sulfat elektroforezom na agarozi i imunoblotin-gom. Koncentracija Lp(a) odre|ivana je imunoturbidi-metrijskom metodom. Pokazano je da je koncentracijaLp(a) u kontrolnoj i u grupi pacijenata pomerena kani`im vrednostima. Za razliku od koncentracije Lp(a)koja pokazuje log-normalnu distribuciju, distribucijaizoformi apo(a) je bimodalna u ispitivanim grupama.Dokazana je obrnuta korelacija izme|u koncentracijeLp(a) i veli~ine apo(a), kako u kontrolnoj grupi(R=–0,592; P<0,001), tako i u grupi pacijenata(R=–0,553; P<0,001). Koncentracija Lp(a) je vi{a(167,5 mg/L) u grupi pacijenata u odnosu na kontrol-nu grupu (106,1 mg/L, p<0,001), dok su izoformeapo(a) zna~ajno manje u grupi pacijenata u odnosu nakontrolnu grupu (p<0,001). Koncentracija Lp(a) rastesa pove}anjem broja KKS zahva}enih stenozom uslu~aju kada nije izvr{ena korekcija za veli~inu izoformiapo(a) (p=0,009), i nakon te korekcije (p=0,001).Me|utim, takva veza nije uo~ena za izoforme apo(a). Ustudiji je dokazano da izoforme apo(a) imaju najve}iuticaj na varijaciju u koncentraciji Lp(a). Male izoformeapo(a) su u vezi sa KAB, ali ne i sa te`im oblikomstenoze.

POLIMORFIZAM PARAOKSONAZE 1 KODPACIJENATA SA ANGIOGRAFSKI

DOKAZANOM KORONARNOM BOLE[]U

J. Kotur-Stevuljevi}, S. Spasi}, A. Stefanovi}, V. Spasojevi}-Kalimanovska, Z. Jeli}-Ivanovi}

Institut za medicinsku biohemiju, Farmaceutski fakultet, Beograd

Tradicionalni faktori rizika za razvoj kardiovasku-larnih (KVB) bolesti: hiperholesterolemija, starost, pol,hipertenzija, dijabetes, fizi~ka neaktivnost i pu{enje,mogu se na}i kod 50% osoba obolelih od kardiovasku-larnih bolesti. Patogeneza tih bolesti kod ostalih 50%obolelih osoba kod kojih se ne identifikuje nijedan odpomenutih faktora rizika obja{njava se prisustvom ma-lih, gustih LDL ~estica, pove}anom koncentracijomoksidovanog LDL-a, povi{enim nivoom homocisteina,uticajem inflamacije i oksidativnog stresa. Poznato jeda za{titna uloga HDL lipoproteinskih ~estica poti~e od

190

njihovog u~e{}a u reverznom transportu holesterola, alii prisustva mnogobrojnih antioksidativnih enzima kojisu sme{teni na njima, a posebno enzima paraokso-naza-1 (PON1). Mi smo ispitivali vezu izme|u statusaenzima PON1 (aktivnost enzima prema dva supstrata,paraoksonu i diazoksonu i fenotip za polo`aj 192 Q/R)i statusa pacijenata sa angiografski dokazanom koro-narnom arterijskom bole{}u. Ispitivali smo i status enz-ima PON1 kod pacijenata i kontrolne grupe u odnosuna oksidativno-stresni status koji smo procenjivali me-renjem koncentracija malondialdehida (MDA), nivoasuperoksidnog anjona (O

2

.–), kao i na osnovu aktiv-

nosti enzima superoksid-dizmutaze (SOD) i koncentra-cije sulfhidrilnih grupa. Distribucija PON1192 fenoti-pova (QQ, QR, RR) izme|u zdravih osoba i pacijenatasa kardiovaskularnim oboljenjem nije se statisti~ki zna-~ajno razlikovala. Rezultati ove studije ukazuju na zna-~ajno ni`u paraoksonaznu aktivnost enzima PON1 kodpacijenata u odnosu na kontrolnu grupu. Povi{enekoncentracije MDA i O

2

.–kao i sni`ena aktivnost SOD

ukazale su na to da bi jedan od uzroka sni`ene PON1aktivnosti mogao biti uznapredovali oksidativni streskod koronarnih bolesnika. Parametri oksidativnog stre-sa (MDA i O

2

.–) bili su u zna~ajnoj korelaciji sa mark-

erima inflamacije (fibrinogen i hsCRP) kod pacijenatasa kardiovaskularnom bole{}u, {to je ukazalo na to dase oksidativni stres razvija kao posledica hroni~ne ni-skostepene inflamacije i da se udru`eno delovanje oksi-dativnog stresa i inflamacije odra`ava na sni`avanjeaktivnosti PON1 enzima, a time i na smanjivanje za{tit-nih sposobnosti HDL ~estica.

PREPOZNAVANJE PACIJENATA SA VISOKIMRIZIKOM U CILJU PRIMARNE PREVENCIJE

ATEROSKLEROZE

D. Pap

Zavod za zdravstvenu za{titu studenata, Slu`ba laboratorijske dijagnostike, Novi Sad, Srbija

Vi{e od polovine iznenadnih smrti usled koronar-ne sr~ane bolesti nastaje kod osoba bez ranije prepoz-natih simptoma. Me|u brojnim novim kardiovaskular-nim faktorima rizika koji su ispitani su subklase lipopro-teina, faktori vezani za koagulaciju (faktor VIIc, plazmi-nogen aktivator inhibitor 1-PAI 1, Von Willebrandov fak-tor), faktori vezani za metabolizam (lipoprotein(a), ho-mocistein, insulin) i faktori vezani za inflamaciju – fibri-nogen, hsC-reaktivni protein, serumski amiloidni A pro-tein, interleukini, MPO, protein S 100B, holin CD 40L,asimetri~ni dimetilarginin-ADMA. Cilj rada je procenalipidnog statusa u studentskoj populaciji iz rizi~nih poro-dica i komparativna analiza lipidnog statusa studenata izporodica bez rizika. Pedeset studenata, starosti 23 godi-ne, iz rizi~nih porodica (KB, AIM, hipertenzija, gojaz-nost, pu{enje, dijabetes melitus, HLP) i 50 studenataiste starosne strukture, oba pola, iz porodica bez rizika,izabrano je za ovu studiju. U svim uzorcima su ura|ene

the well-known antiatherogenic function of this lipopro-tein conducted through its involvement in reverse cho-lesterol transport and also in many antiatherogenicenzymes located on HDL, especially paraoxonase-1(PON1). We have investigated the relationship bet-ween PON1 status [PON1 enzymatic activity towardstwo substrates, paraoxon and diazoxon and also phe-notype PON1192 Q/R, coming from the substitutionof amino acid glutamine (Q) with arginine (R)] and thedisease status of angiographycally assessed coronarydisease patients. We have also investigated the relationbetween PON1 status and oxidative stress status whichwas estimated by measuring malondialdehyde (MDA)concentration, superoxide anion (O

2

.–) level, and also

by superoxide dismutase (SOD) activity and total sul-phydril (SH) groups content. PON1192 phenotype dis-tribution (QQ, QR, RR) in the healthy control groupwas not significantly different from that in coronaryheart disease patients. Results of this study indicate sig-nificantly lower paraoxonase PON1 activity in CHD pa-tients compared to controls. Increased MDA and O

2

.–

levels as well as decreased SOD activity showed thatone of the reasons for lowered PON1 activity could beexaggerated oxidative stress. Oxidative stress parame-ters (MDA and O

2

.–) were in statistically significant,

strong correlation with inflammation markers (fibrino-gen and hsCRP) in CVD patients, which indicated thatoxidative stress is a consequence of chronic low-gradeinflammation and their common action influence onPON1 activity lowering and diminished HDL protectivecapability.

IDENTIFYING HIGH-RISK PATIENTS FOR THE PRIMARY PREVENTION OF

ATHEROSCLEROSIS

D. Pap

Student Health Protection Institute, Department of Laboratory Diagnostics, Novi Sad, Serbia

More than half of sudden deaths from coronaryheart disease occur in individuals with no prior symp-toms recognised. Among a number of new cardiovas-cular risk factors being evaluated are lipoprotein sub-species, excesssive plasma levels of homocysteine,abnormal blood coagulation characteristics and inflam-mation – MPO, hsCRP, protein S 100B, choline CD40L, asymmetric dimethylarginine-ADMA, etc. Theaim of the study is to estimate the lipid status in a stu-dent population from famillies at risk and to run a com-parative analysis of students from families without sucha risk. Fifty students from at-risk families (CHD, AMI,hypertension, obesity, smoking, diabetes mellitus, HLP)and 50 students from families without such a risk ofboth sexes were selected for the study. The followingdeterminations were made: total cholesterol (TCH),triglycerides (TG), HDL-c, low density lipoprotein cho-

JMB 2008; 27 (2) 191

192

slede}e analize: ukupni holesterol (UH), trigliceridi(TG), HDL-hol, lipoproteini male gustine (LDL-hol.), li-poproteini vrlo male gustine (VLDL-hol.), indeks ate-roskleroze (IA), i utvr|eni su faktori rizika (FR). Odre|enje i indeks telesne mase (BMI). Vrednosti UH bile suzna~ajno vi{e u rizi~noj grupi. Vrednosti LDL-hol, trigli-cerida, indeksa ateroskleroze i faktora rizika zna~ajno suvi{i u rizi~noj grupi (p<0,01). Vrednosti HDL-hol.zna~ajno su ni`e u rizi~noj grupi (p<0,01). Rezultati supokazali da izme|u BMI i UH, LDL-hol. i TG postoji sta-tisti~ki zna~ajna razlika (p<0,05), dok izme|u BMI iHDL-hol. nije utvr|ena takva razlika (p>0,05). Na ispi-tivane parametre u obe grupe nije uticao pol ispitanika.Dobijeni rezultati ukazuju na potrebu skrininga lipidnogstatusa u studentskoj populaciji iz rizi~nih porodica, pri-menu mera primarne prevencije kroz promenu na~ina`ivota, promociju zdravog na~ina `ivota, kao i modifiko-vanje faktora rizika u cilju spre~avanja progresijeateroskleroze a samim tim i koronarne arterijske bolesti,u pojedinim slu~ajevima i le~enje utvr|enih klini~kihslu~ajeva.

EFIKASNOST RAZLI^ITIH TERAPIJSKIH PROTOKOLA ZA SR^ANU INSUFICIJENCIJU

KOD PACIJENATA SA SR^ANOM INSUFICIJENCIJOM I POVI[ENIM

NIVOOM NATRIURETSKIH PEPTIDA

R. Kova~evi}, M. Miri}

Institut za kardiovaskularne bolesti »Dedinje«,Biohemijska laboratorija,

Centar za kardiovaskularna istra`ivanja, Beograd, Srbija

Natriuretski peptidi mogu imati klini~ki zna~aj zapra}enje terapijskih efekata kod pacijenata sa sr~anominsuficijencijom. Cilj ove randomizovane studije je bioda se ispita efikasnost razli~itih terapijskih protokola zasr~anu insuficijenciju kod pacijenata sa sr~anom insufi-cijencijom na osnovu nivoa BNP-a u plazmi pre i posleterapije. [ezdeset dva pacijenta sa sr~anom insuficijen-cijom i povi{enim nivoom BNP-a, dobi 55,82±9,09god, sa II-III NYHA funkcionalnom klasom, ejekcionomfrakcijom <45%, primali su 12 nedelja tradicionalnufarmakolo{ku terapiju za sr~anu insuficijenciju, i to:ACE inhibitori i ß-blokatori (1. grupa); ACE inhibitori iangiotenzin II receptor blokatori (ARBs) (2. grupa); ß-blokatori i ARBs (3. grupa); ACE inhibitori, ß-blokatori iARBs (4. grupa). Ispitivali smo nivo BNP-a u plazmi,hemodinamske parametre (pritisak u plu}nim kapilari-ma (PCWP), cardiac index (CI), EF i du`inu trajanjaoptere}enja. Nivo BNP-a u plazmi se zna~ajno snizio u4. grupi pacijenata, koji su bili na ACE inhibitorima, ß-blokatorima i ARBs, u odnosu na ostale grupe. Tako|eje prime}en zna~ajan pozitivan uticaj na hemodinam-ske parametre i trajanje optere}enja u toj grupi pacije-nata, u pore|enju sa ostalim ispitanicima (rezultati su utabeli).

lesterol (LDL-c), very low-density lipoprotein choles-terol (VLDL-c), index of atherosclerosis IA, establishedrisk factors (RF) and body mass index (BMI). The valueof TCH was significantly higher in the at-risk group ofpatients, while LDL-chol. TG, IA and RF were found tobe significantly higher in students from families at-risk(p 0.01), HDL-chol. was significantly lower (p<0.01) instudents from the at-risk group. The results also sho-wed that a statistically significant difference was found(p<0.5) between BMI and TCH, LDL-c. and TG, how-ever no statistically significant difference wasn foundbetween BMI and HDL-c (p>0.05). These data sug-gest that screening of the lipid status is necessary instudent populations from families at-risk. Primary pre-vention is very important and can be achieved throughlifestyle changes, the promotion of a healthy way of life,as well as modifications of risk factors with the aim ofpreventing atherosclerosis, coronary heart disease, andin some students through a therapeutic intervention inestablished clinical cases.

THE EFFICACY OF DIFFERENT THERAPYPROTOCOLS FOR HEART FAILURE INPATIENTS WITH HEART FAILURE AND

INCREASED NATRIURETIC PEPTIDE LEVEL

R. Kova~evi}, M. Miri}

Cardiovascular Institute »Dedinje«, Biochemistry Laboratory,

Cardiovascular Reseach Center, Belgrade, Serbia

Natriuretic peptide BNP might be clinically usefulfor monitoring treatment effects in patients with heartfailure (HF). In order to investigate the pharmacologi-cal effects of different therapy protocols for patientswith HF based on the BNP level before and after ther-apy, we performed an open randomized comparativetrial. Sixty two HF patients with increased natriureticpeptide level, aged 55.82±9.09, II-III NYHA function-al classes, ejection fraction (EF) <45%, received a 12-week treatment with either traditional pharmacothe-rapy for HF with ACE inhibitors and ß-blockers (1stgroup), or ACE inhibitors and angiotensin II receptorblockers (ARBs) (2nd group), or ß-blockers and (ARBs)(3th group), and ACE inhibitors, ß-blockers and ARBs(4th group). We evaluated the BNP plasma level, he-modynamic state (pulmonary capillary wedge pressure(PCWP), cardiac index (CI), EF and exercise capacity.The BNP plasma level decreased significantly in the 4th

group of patients who received ACE inhibitors, ß-blo-ckers and ARBs, in comparison to other groups. A be-neficial influence on hemodynamic and exercise capa-city was significantly pronounced in this group, compa-red to the other therapy regimes (the results are in thetable).

Mo`e se zaklju~iti da terapijski protokol koji uklju-~uje ACE inhibitore, ß-blokatore i ARBs kod pacijenatasa sr~anom insuficijencijom i povi{enim BNP-om zna-~ajno pobolj{ava kvalitet `ivota, funkciju leve komore,hemodinamske parametre i du`inu trajanja optere}e-nja. Sve te promene su udru`ene sa sni`enjem nivoaBNP-a u plazmi.

In conclusion, the therapeutic protocol: ACE inhi-bitors, ß-blockers and ARBs in HF patients with increa-sed natriuretic peptide level significantly improves thequality of life, left ventricular function, hemodynamicparameters and exercise capacity. All these changeswere accompanied with a decreasing of the BNP plas-ma level.

JMB 2008; 27 (2) 193

BNP, % PCWP, % CI, % EF, % trajanjeopt.

1. grupa –13,5* –9,4* +6,1* +8,8** +11,2**

2. grupa –14,9* –6,3* +4,1* +7,8** +7,2**

3. grupa –15,8* –13,0** +5,3* +7,0* +6,5*

4. grupa –39,8** –20,0** +11,1** +14,6** +14,8**

*–p<0,01; **–p<0,001 u pore|enju sa vrednostima preterapije

BNP, % PCWP, % CI, % EF, % Exercisecap, %

Group 1 –13.5* –9.4* +6.1* +8.8** +11.2**

Group 2 –14.9* –6.3* +4.1* +7.8* +7.2**

Group 3 –15.8* –13.0* +5.3* +7.0* +6.5*

Group 4 –39.8** –20.0** +11.1** +14.6** +14.8**

*–p<0.01; ** –p<0.001 compared with baseline

Promene nivoa BNP-a, hemodinamskih parametara i trajanjaoptere}enja

Changes in BNP, hemodynamic parameters and exercisecapacity

4th EFCC Symposium for Balkan Region

The Impact of the Pre-analytical Phaseon the Quality of the Laboratory Results

POGRE[NA IDENTIFIKACIJA I OSTALE PREANALITI^KE GRE[KE

PA Bonini1,2,3, F. Ceriotti3, G. Mirandola2, C. Signori4

1Katedra za klini~ku biohemiju i klini~ku molekularnu biologiju, Medicinski fakultet,

Univerzitet »Vita Salute San Raffaele«, Milano, Italija2CeSREM (Centro Studi San Raffaele Rischi

Errori in Medicina), Milano, Italija3LABORAF, Centar za laboratorijsku medicinu, Nau~ni institut »San Raffaele«, Milano, Italija

4Menad`ment klini~kog rizika, Klini~ki institut »Humanitas«, Milano, Italija

Najve}i broj laboratorijskih gre{aka doga|a se upreanaliti~koj fazi uglavnom iz razloga koji se ti~u obra-zovanja i organizacije. Bi}e predstavljeno iskustvo na{eustanove, kao i rezultati zajedni~kih napora italijanskihlaboratorija da se otkriju i umanje gre{ke ili rizik od gre-{aka u laboratorijskoj medicini.

POBOLJ[ANJE PREANALITI^KOG PROCESA:FOKUSIRANJE NA KVALITET UZORKA

S. Green

Department of Medical and Scientific Affairs andClinical Operations, BD Diagnostics, Preanalytical

Systems, Franklin Lakes, New Jersey, USA

Trendovi u klini~koj laboratorijskoj praksi posta-vljaju vi{e zahteva za kvalitet uzoraka pacijenata. Una-pre|enje analiti~kih karakteristika (npr. pove}anje auto-matizacije, smanjenje zapremine uzorka, pove}anjeosetljivosti odre|ivanja), kao i efikasnosti i smanjenjetro{kova (npr. prohodnost, vreme obrta testova) je una-predilo medicinsku praksu. Me|utim, ove promene suuzrokovale pove}anje incidencije preanaliti~kih gre{aka{to zahteva potrebu za uzoraka ve}eg kvaliteta. Slede}i~lanak govori o ovim gre{kama kao dodatak za po-bolj{anje kvaliteta preanaliti~ke faze.

MISIDENTIFICATION AND OTHER PREANALYTICAL ERRORS

PA Bonini1,2,3, F. Ceriotti3, G. Mirandola2, C. Signori4

1Chair of Clinical Biochemistry and Clinical Molecular Biology, School of Medicine,

Università Vita Salute San Raffaele, Milano, Italy2CeSREM (Centro Studi San Raffele Rischi

Errori in Medicina), Milano, Italy3LABORAF, Department of Laboratory Medicine

Istituto Scientifico San Raffaele, Milano, Italy4Clinical Risk Management, Istituto Clinico

Humanitas, Milano, Italy

The largest number of Laboratory Errors occurin the pre-analytical phase and are mainly due toeducational and organizational reasons. The experi-ence of our Institution, as well as the results of anItalian interlaboratory effort to detect and reduceerrors/risk of errors in Laboratory Medicine will beillustrated.

IMPROVING THE PREANALYTICAL PROCESS: THE FOCUS ON SPECIMEN QUALITY

S. Green

Department of Medical and Scientific Affairs andClinical Operations, BD Diagnostics, Preanalytical

Systems, Franklin Lakes, New Jersey, USA

Trends in clinical laboratory practice place moredemands on the quality of patient specimens. Advancesin analytical performance (i.e., increased automation,reduced sample volume, increased assay sensitivity), aswell as efficiency and cost reduction gains (i.e., through-put, test turnaround time) have improved medical prac-tice. These changes, however, have caused an increa-sed incidence of preanalytical errors, dictating the needfor higher-quality specimens. The following paperaddresses these errors in addition to methodologies forimproving the quality of the preanalytical phase.

JMB 2008; 27 (2) 197

UDK 577.1 : 61 ISSN 1452-8258

JMB 27: 197–204, 2008 Plenarne sekcije

Plenary sessions

GOVERNANCE OF PREANALYTICAL VARIABILITY. ERRORS DETECTION.

G. Lippi

Clinical Chemistry Section, University of Verona, Italy

In the age of evidence-based medicine, results oflaboratory testing are integral to the clinical decisionmaking, to assist diagnosis, guide or monitor therapyand predict health outcomes. Owing to the increasingdemand placed on laboratory diagnostics and thewedging pressure from cost containment policies, theprimary goal is to achieve a high degree of efficiencywith as little as possible influence on the quality.Although there is widespread perception that mosterrors in medicine occur from mishandled therapies,both medical and surgical, they can also develop with-in the laboratory diagnostics, especially in the mostmanually-intensive preanalytical steps. Since preanaly-tical variability exerts a strong influence on laboratoryorganization, healthcare expenditures and patientsoutcome, governance of this crucial phase of the totaltesting process by reduction of uncertainty offer thegreatest potential for improving total quality andenhance stakeholders satisfaction. Most preanalyticalerrors result from system flaws and insufficient auditwith operators involved in specimen collection/han-dling responsibilities. Therefore, standardization andmonitoring of most, if not all, preanalytical variableswould be associated with the best organizational andclinical revenues. The most reliable strategy shouldhence be tailored to both predict the onset of acci-dental events (incidents) and decrease the vulnerabili-ty of preanalytical steps. The mainstay and necessarypreliminary step in governance of preanalytical vari-ability is errors identification. The magnitude of labo-ratory errors requires, however, the introduction ofcomprehensive reporting systems, encompassing mis-takes falling within the whole diagnostic process, oncea list of performance indicators has been defined on alocal basis. The most suitable approach is to develop asystem including also a variety of representative prean-alytic performance measures, based on criteria forspecimen acceptability. The implementation of a sys-tematic error tracking system in the daily practicewould enable considerations on specific problems andresponsibilities, grant meaningful information on thelocal preanalytic processes more susceptible to errors,thus providing the ideal basis for eliminating bottle-necks and flaws, and redesigning the structure of thetotal testing process.

UPRAVLJANJE PREANALITI^KOM VARIJABILNO[]U I OTKRIVANJE GRE[AKA

\. Lipi

Odeljenje klini~ke hemije, Univerzitet u Veroni, Italija

U vremenu medicine zasnovane na dokazima,rezultati laboratorijskog testiranja su integralan deo kli-ni~kog odlu~ivanja, i poma`u u postavljanju dijagnoze,vo|enju ili pra}enju terapije i predvi|anju klini~kog isho-da. S obzirom na sve {iru upotrebu laboratorijske dijag-nostike i sve ve}i pritisak da se smanje tro{kovi, osnovnicilj je postizanje visokog stepena efikasnosti uz {to manjiuticaj na kvalitet. Iako postoji uvre`eno mi{ljenje da ve-}ina gre{aka u medicini poti~e iz pogre{nih terapija,medicinskih i hirur{kih, one mogu nastati i u laboratori-jskoj dijagnostici, naro~ito u preanaliti~koj fazi kojapodrazumeva najvi{e manuelnog rada. Kako preanali-ti~ka varijabilnost ima veliki uticaj na organizaciju labo-ratorije, tro{kove u zdravstvu i kona~ni ishod pacijenata,upravljanje tom klju~nom fazom ukupnog postupka tes-tiranja putem smanjenja neizvesnosti pru`a najvi{e mo-gu}nosti za pobolj{anje ukupnog kvaliteta i pove}anjezadovoljstva deoni~ara. Najve}i broj preanaliti~kihgre{aka rezultat su manjkavosti sistema i nedovoljno ~e-stih inspekcija u radu operatera ~ija je odgovornostprikupljanje/rukovanje uzorcima. Stoga bi se standard-izacija i pra}enje najve}eg broja, ako ne i svih, preanali-ti~kih varijabli mogli dovesti u vezu sa najboljim organi-zacionim i klini~kim prihodima. Najpouzdanija strategijabi dakle trebalo da bude pravljena tako da predvidi nas-tanak slu~ajnih doga|aja (incidenata) i smanji osetljivostpreanaliti~kih koraka. Glavna stavka i neophodan pre-liminarni korak u upravljanju preanaliti~kom varijabil-no{}u je identifikacija gre{aka. Veliki broj laboratorijskihgre{aka, me|utim, zahteva uvo|enje sveobuhvatnih sis-tema izve{tavanja, koji obuhvataju gre{ke u ukupnomdijagnosti~kom postupku, po{to se prethodno defini{elista pokazatelja performansi na lokalnoj osnovi. Najboljipristup jeste razvijanje sistema koji bi uklju~io i izvestanbroj reprezentativnih mera preanaliti~ke performanse,na osnovu kriterijuma za prihvatljivost uzoraka. Imple-mentacija sistema za sistematsko pra}enje gre{aka usvakodnevnoj praksi omogu}i}e pristupanje specifi~nimproblemima i odgovornostima, pru`iti va`ne informacijeo lokalnim preanaliti~kim postupcima koji su najpod-lo`niji gre{kama, i na taj na~in dati idealnu osnovu za eli-minisanje uskih grla i nedostataka, i preoblikovanje stru-kture ukupnog postupka testiranja.

198

BEZBEDNOSNI STANDARDI U UZIMANJU KRVI

K. Matiuci

Medicinska uprava, Univerzitetska bolnica u Veroni,Verona, Italija

Laboratorijska dijagnostika je zahvaljuju}i zna~aj-nom doprinosu koji daje u postavljanju dijagnoze, pra-}enju i terapeutskom monitoringu pacijenata presudnaza proces klini~kog odlu~ivanja. Praksa flebotomije segeneralno odnosi na uzimanje venske krvi. Iako jeuvo|enje pravih igala za jednokratnu upotrebu i vaku-umskih sistema cevi omogu}ilo uzimanje uzorakaodgovaraju}eg kvaliteta, uz dodatne op{te prednostikako sa stanovi{ta bezbednosti tako i u prakti~nojupotrebi, nekoliko razli~itih aspekata uti~e na celoku-pan postupak u vezi sa uzimanjem uzoraka krvi. Glavnirazlog tako velike prevalence problema je {to je trenut-no te{ko pratiti ve}inu preanaliti~kih varijabli, uklju-~uju}i flebotomiju, koje nisu uvek pod direktnom kon-trolom ili nadzorom osoblja u laboratoriji. Praksa fle-botomije se razlikuje {irom sveta; i evropskim oblastimanema~kog govornog podru~ja za sada je samo lekari-ma dozvoljeno da uzimaju uzorke venske i arterijskekrvi i njih stoga obu~avaju i nadgledaju starije kolegena odeljenjima. U Britaniji se flebolozi obu~avaju kaotehni~ari i delimi~no su pod nadzorom profesionalnihlaboranata. Zahvaljuju}i trendu konsolidacije labora-torijskog testiranja, koje }e neizbe`no obuhvatitiplanove za nabavku u vezi sa prikupljanjem i trans-portom uzoraka, o~ekuje se da }e u budu}nosti poja-~ani nadzor decentralizovanih postupaka u flebotomijidobiti jo{ ve}i zna~aj. Flebotomija je i dalje jedna odnaj~e{}e zapostavljanih procedura u zdravstvu, ali jeneophodna za dobijanje uzoraka krvi u dijagnostici.Pitanje bezbednosti u prikupljanju uzoraka krvi ~esto jepotencirano tokom prethodnih decenija. Me|utim,bezbednost pacijenta bila je na prvom mestu u vizijizdravstva u ~ijem se centru nalazi pacijent, dok su uslo-vi neophodni za pru`anje, odr`avanje i unapre|ivanjebezbednosti osoblja u zdravstvu odgovornog za uzi-manje krvi bili zanemareni. Flebolozi su integralan deozdravstvenog sistema, a njihova aktivnost podrazumevaozbiljne zdravstvene rizike, uglavnom usled slu~ajnihpovreda operatera ubodom igle i uobi~ajenih ozledananesenih pacijentu (ozlede nerva ili tetive), krvarenje,vrtoglavica/sinkopa, limfedem), kao posledica nepravil-no izvr{enog se~enja vene. Stoga bi gotovo svi zdrav-stveni sistemi trebalo ozbiljno da se pozabave pitanjempovreda pacijenta i operatera prilikom uzimanja krvi.Pristup bezbednosti u uzimanju krvi neizbe`no je vi{e-stran, i obuhvata {irenje znanja i preporuka, obuku iizdavanje sertifikata flebolozima. Bez obzira na lokalneuslove, radnici u zdravstvu odgovorni za specifi~nezadatke, poput flebotomije, moraju biti propisno obra-zovani i motivisani za obavljanje tih zadataka sa {to jemanje mogu}e gre{aka i povreda. Glavni ciljevi obukebi trebalo da im pribli`e aspekte koji su naj~e{}e uklju-~eni u zasecanje vene, uklju~uju}i preno{enje znanja o

STANDARDS OF SAFETY IN BLOOD COLLECTION

C. Mattiuzzi

Medical Direction, University Hospital of Verona,Verona, Italy

Laboratory diagnostics is pivotal to the clinicaldecision making, since it substantially contributes todiagnosis, follow-up and therapeutic monitoring ofpatients. The practice of phlebotomy generally refersto the collection of venous blood. Although the intro-duction of disposable straight needles and evacuatedtube systems has allowed collection of specimens ofsuitable quality, with additional general advantagesfrom both a safety and a practical point of view, theoverall procedures linked to blood sample collection,and the phlebotomy success rate itself, are as yetinfluenced by several aspects. The main reason forsuch a high prevalence of problems is that it is cur-rently difficult to monitor most of the pre-analyticvariables, including phlebotomy, which are not bealways under direct control or supervision of the labo-ratory staff. The phlebotomy activity is rather hetero-geneous worldwide; in the German speaking area ofEurope only physicians are presently allowed to drawvenous and arterial blood and are therefore trainedand supervised by the elder colleagues in the ward.In Britain, phlebotomists are educated like techni-cians and are partially supervised by laboratory pro-fessionals. Owing to the expanding trend towardsconsolidation of laboratory testing, which willinevitably entail outsourcing plans for specimen col-lection and transportation, improved vigilance ofdecentralized phlebotomy procedures is expected togain further relevance in the future. Phlebotomy isstill one of the most neglected procedure in health-care, but it is essential to achieve blood specimensfor diagnostics. Much emphasis has been placed onsafety issue in blood collection over the past decades.However, patient safety has been foremost in the pa-tient-centred vision of healthcare and the necessaryrequirements for ensuring, maintaining or enhancingthe safety of healthcare personnel with blood collec-tion responsibilities have been overlooked. Phleboto-mists are integral parts of the healthcare system, andtheir activity still involves serious health risks, mainlyrepresented by accidental needlestick injuries to theoperator and casual lesions inflicted to the patient(nerve or tendon injury, haemorrhage, vertigo/syn-cope, lymphedema) as a result of improperly per-formed venipuncture. Therefore, injuries to bothpatient and operator while collecting blood should bea matter of concern for most health systems. Theapproach to safety in blood collection is necessarymultifaceted, which includes recommendations andknowledge dissemination, training, and certificationof phlebotomists. Regardless of local requirements,healthcare workers responsible for specific tasks,such as phlebotomy, must be properly educated and

JMB 2008; 27 (2) 199

motivated to perform those tasks with as few errorsand injuries as possible. The main objectives of train-ing should encompass those aspects most frequentlyencountered while performing venipunctures, includ-ing notions of anatomy and physiology (location ofveins, nerves, tendons and arteries), characteristics ofdevices (needles, syringes, tourniquets, disinfectants,evacuated tube systems), phlebotomy techniques,employability skills and emergency procedures.Moreover, basic notions of psychology, mostly aimedat providing relational, reassuring skills directedeither to adults or to children should be given as partof the training. Before entering the work force, phle-botomists must achieve a minimum clinical curricu-lum, based on repeated practical experience, whichshould allow them to accomplish the entire phle-botomy process at the best. The diagnostic industryis crucial to this issue, in that it should provide health-care operators with safety devices with the lowestpossible chance of injuries and accidents. Directsupervision of phlebotomists by the laboratory staffwould also be advisable, in that the laboratory pro-fessionals can provide phlebotomists with a compre-hensive view of the complexity of the total testingprocess. Compliance with Best Practice (»BP«) goals,as included in Phlebotomy Personnel Standards andOSHA, JCAHO and CDC recommendations, shouldfinally be regarded as essential means to improve thequality within the whole blood collection procedures.In Germany, for example, a training program avail-able on CD-Rom and called DIAPRO was started.The program contains texts and examples to trainfuture medical professionals in preventing errors dur-ing phlebotomy. Abating the number of unnecessarytests and limiting the number of steps in which speci-mens are delivered to laboratories, tests are per-formed, and results provided to the requesting physi-cian might be additional but not alternative objectives,along with other pre-analytic issues not immediatelyassociated with the phlebotomist’s activity. Finally, thebroad implementation of phlebotomists’ certificationworldwide would enable a high degree of profession-alism in this crucial part of the total testing process.

PREANALYTICAL PHASE IN HAEMATOLOGY

G. Banfi1,2, L. Germagnoli1,3

1IRCCS Galeazzi, Milan, Italy2Dipartimento Tecnologie per la Salute,

School of Medicine, University of Milan, Milan, Italy3Laboraf, Milan, Italy

The preanalytical phase is particularly importantin haematology, where counts of particles and cells areperformed in whole anticoagulated blood. The correctuse and concentration of anticoagulant is mandatoryto avoid spurious results, which can influence clinical

anatomiji i fiziologiji (lokacija vena, nerava, tetiva i arte-rija), osobinama alatki (igle, {pricevi, podvezi, dezinfek-ciona sredstva, vakuumski sistemi cevi), tehnika fle-botomije, ve{tina njihovog primenjivanja i hitnim pos-tupcima. [tavi{e, zahteva se i osnovno poznavanje psi-hologije, u cilju uspostavljanja poverljivijeg odnosa saodraslima i decom. Pre zapo{ljavanja, flebolozi bi mo-rali ste}i minimum klini~kog iskustva, na osnovu pon-avljanog prakti~nog iskustva, koje bi trebalo da imomogu}i uspe{no sprovo|enje postupka flebotomije.Dijagnosti~ka industrija u tome ima najva`niju ulogu, ato je da zaposlenima u zdravstvu pru`i bezbedna sred-stva za rad sa najmanjim mogu}im rizikom od povredaili nezgoda. Tako|e se preporu~uje da profesionalnilaboranti nadziru flebologe, po{to profesionalni labo-ranti mogu flebolozima da pru`e sveobuhvatan uvid uslo`enost ukupnog postupka testiranja. Na kraju, cil-jeve za postizanje najbolje prakse (»BP«), obuhva}eneStandardima za medicinsko osoblje u flebotomiji i pra-vilnicima OSHA, JCAHO i CDC, treba smatrati osnov-nim sredstvima za pobolj{anje kvaliteta u celokupnompostupku uzimanja krvi. U Nema~koj je, na primer,za`iveo program obuke po imenu »DIAPRO« koji jedostupan na CD-romu. Program sadr`i tekstovi i pri-mere za obuku budu}ih medicinskih stru~njaka u spre-~avanju gre{aka tokom flebotomije. Smanjivanje brojanepotrebnih testova i ograni~avanje broja koraka, kojipodrazumevaju isporuku uzoraka laboratorijama, obav-ljanje testova i preno{enje rezultata lekaru koji ih jetra`io, mo`da su dodatni ali ne i protivre~ni ciljevi, uzostala preanaliti~ka pitanja ne nu`no povezana sa ra-dom flebologa. Kona~no, implementacija sertifikacijeflebologa {irom sveta podstakla bi visok stepen profe-sionalizma u tom klju~nom delu ukupnog postupkatestiranja.

PREANALITI^KA FAZA U HEMATOLOGIJI

G. Banfi1,2, L. Germagnoli1,3

1IRCCS Galeazzi, Milano, Italija2Odeljenje za tehnologiju u zdravstvu, Medicinski

fakultet, Univerzitet u Milanu, Milano, Italija3Laboraf, Milano, Italija

Preanaliti~ka faza je naro~ito va`na u hematolo-giji, gde se merenja ~estica i }elija vr{e u celoj antiko-agulisanoj krvi. Ispravna upotreba kao i koncentracijaantikoagulansa je obavezna kako bi se izbegli nejasnirezultati, koji mogu uticati na klini~ko odlu~ivanje.

200

EDTA je preporu~ljivi antikoagulans, mada ima izvesnaograni~enja, posebno u vezi sa o~uvanjem stabilnosti ioblika plo~ica. Stabilnost hematolo{kih parametara jevisoka, sa izuzetkom leukocita i retikulocita. Me|utim,stabilnost (i instrumentalnu preciznost) treba ocenjivatizajedno sa biolo{kom varijabilno{}u i indeksom indi-vidualnosti razli~itih hematolo{kih parametara. Nahematolo{ke testove tako|e uti~u visoke koncentracijelipida i kilomikrona. Postupak me{anja epruveta posleuzimanja krvi a pre analize tako|e je klju~an za dobi-janje ta~nih i validnih podataka. Postoje primeri inter-ferencije na automatizovanim hematolo{kim analiza-torima koji se koriste za dijagnostikovanje i pra}enjepatolo{kih stanja. Krioglobulini i paraziti eritrocita mo-gu izazvati nejasne rezultate WBC, RBC i PLT, ali pon-avljanje takvih interferencija mo`e se iskoristiti zaalarmiranje lekara i mo`e ukazati na prisustvo pato-lo{kih proteina ili parazita u krvi. Moderni hematolo{kianalizatori donose nove parametre, direktno defini-sane ili izra~unate na osnovu tradicionalnih mera, aliklini~ki uticaj tih novih parametara ~esto zavisi od pre-analiti~kih varijabli.

KVALITET DIJAGNOSTI^KIH UZORAKA, PREPORUKE I NASTAVNA SREDSTVA

VG. Guder

Minhen, Nema~ka

Analiti~ki standardi opisani su utvr|enim kriteriju-mima kontrole kvaliteta, me|utim, za prenaliti~ku fazutakvi standardi ne postoje. S druge strane, mnogi pre-analiti~ki aspekti obuhva}eni su poslednjom NormomEN ISO 15189 za rukovo|enje kvalitetom u medicin-skim laboratorijama. Preporuke za kvalitet dijagnosti-~kih uzoraka (1,2) uklju~uju definiciju optimalne koli-~ine uzorka, upotrebu antikoagulanasa i stabilizatora,kao i kriterijume stabilnosti tokom transporta i ~uvanja.Osim toga, preanaliti~ko rukovanje hemoliziranim,lipemi~nim i ikteri~nim uzorcima se ponovo procenju-ju. Tehni~ke preporuke u pogledu uzorkovanja, trans-porta i identifikacije su izdate od strane NCCLS, SZO iIFCC. Nedavno su ti standardi objavljeni kao nastavnasredstva na internetu (3). Ta verzija dopu{ta da se une-su pitanja u vezi sa 350 analita, na koja }e se odgov-oriti preko podataka koji se odnose na izbor antikoagu-lansa, stabilnost i potrebnu koli~inu uzorka. Pored toga,nema~ki program obuke koji je nedavno objavljen naCD-romu sadr`i podatke o preanaliti~kim varijablamakao i preporuke za sve procedure uzorkovanja (4). Tiprogrami su testirani u pet nacionalnih radionica kaomaterijal za obuku i mogu isto tako slu`iti za akredita-cione procedure medicinskih laboratorija, po{to je pre-analiti~ka faza identifikovana kao glavni izvor gre{aka ulaboratorijama {irom sveta.

decision. EDTA is the anticoagulant of choice, but ithas some limits, especially for preserving stability andshape of platelets. Stability of haematological parame-ters is high, with the exception of leukocytes and retic-ulocytes. However, stability (and instrumental preci-sion) should be evaluated together with biological vari-ability and individuality index of various haematologicalparameters. Hematological tests are also influencedand interfered by high amounts of lipids and chylomi-crons. The mixing procedure of the tubes after blooddrawing and before analysis is also crucial for obtain-ing correct and valid data. There are some examplesof interferences on automated haematological analy-sers which are used for diagnosing and screeningpathological conditions. Cryoglobulins and erythro-cytes parasites can induce spurious results of WBC,RBC and PLT, but the repeatability of these interfer-ences could be used for alerting the pathologist andcould reveal the presence of pathological proteins orblood parasites. New parameters have been proposedby modern haematological analysers, directly definedor calculated from traditional measures, but the clini-cal impact of these new parameters is often dependenton preanalytical variables.

QUALITY OF DIAGNOSTIC SAMPLES,RECOMMENDATIONS AND

EDUCATIONAL TOOLS.

WG. Guder

Munich, Germany

Whereas analytical standards are described byestablished quality control criteria, no such standardsexist regarding the preanalytical phase. On the otherhand, many preanalytical aspects are part of the recentNorm EN ISO 15189 on quality management in the me-dical laboratory. Recommendations on the quality of di-agnostic samples (1,2) include the definition of optimalsample size, the use of anticoagulants and stabilizers, aswell as stability criteria regarding transport and storage.Moreover, the preanalytical handling of haemolytic, lipe-mic and icteric samples has been reevaluated. Technicalrecommendations regarding sampling and transportidentification have been published by NCCLS, WHO andIFCC. Recently, these standards have been edited aseducational tools in the internet (3). This version allowsto enter questions regarding 350 analytes, which will beanswered by data regarding the choice of anticoagulant,stability and sample volume needed. In addition, aGerman teaching program on CD-Rom has recentlybeen published containing knowledge on the preanalyti-cal variables as well as recommendations for all samplingprocedures (4). These programs were tested in five na-tional workshops as teaching material and can likewiseserve for accreditation procedures of medical laborato-ries, after the preanalytical phase was identified as themajor source of errors in laboratories worldwide.

JMB 2008; 27 (2) 201

1. The Quality of Diagnostic Samples. Empfehlungender Arbeitsgruppe Präanalytik of the German Socie-ties for Clinical Chemistry and Laboratory Medicine.J Lab Med 2002; 26: 267–83.

2. The Quality of Diagnostic Samples. CD-Rom Chro-nolab, Zug, Switzerland 2003, included in GuderWG, Narayanan S, Wisser H, Zawta B. Samples:From the Patient to the Laboratory 3rd rev Ed,Weinheim, Wiley-VCH, 2003.

3. Internet version including expert forum, 2003:www.//diagnosticsample.com

4. Guder WG, Hagemann P, Wisser H, Zawta B. FokusPatientenprobe. Kompendium Präanalytik. CD-Rom BD Germany, Austria, Switzerland 2007

SAMPLE STABILITY

G. Luca Salvagno

Clinical Chemistry Section, University of Verona, Italy

Over the past years, clinical laboratories havebeen at the vortex of the maelstrom affecting medi-cine. Various strategies to contain and reduce overallcosts of laboratory services are being implemented orhave been advocated. These include centralization,consolidation and integration of services, reengineer-ing the laboratory and increasing the level of auto-mation, optimizing test usage, decentralizing testingto the point-of-care. The growing trend to closuresand mergers of hospital have led to consolidation oflaboratory services for logistic and economic reasons.Moreover, many hospitals, clinics and physician prac-tices find it economic to send out specimens for test-ing to commercial laboratories, because performinglow volume or esoteric testing is not economicallyviable. In areas of the country in which healthcarenetworks with aggressive outreach programs arestrong, laboratories in smaller hospitals are beingclosed, and the tests sent to a central laboratory. Mo-reover, due to the tremendous growth of decentral-ized phlebotomy services worldwide, blood samplesmay arrive in the central laboratory from varying dis-tances, under variable storage and transportationconditions. As a result, central laboratories servinghealthcare networks are experiencing an increase inworkload and facing new problems related to samplestability. The lack of standardized procedures forsample collection, account for most of the errorsencountered within the total testing process. Theseinclude inappropriate procedures for collection andhandling of the specimens, such as the use ofimproper collection tools, prolonged stasis duringvenipuncture, and time before centrifugation or ana-lysis and finally unsuitable storage. The problems inpreparation and transporting of samples from a pe-ripheral facility to a centralised laboratory cannot be

1. The quality of diagnostic samples. Empfehlungender Arbeitsgruppe Praanalytik of the German Socie-ties for Clinical Chemistry and Laboratory Medicine.J Lab Med 2002; 26: 267–83.

2. The quality of diagnostic samples. CD-Rom Chro-nolab, Zug, Switzerland 2003, included in GuderWG, Narayanan S, Wisser H, Zawta B. Samples:From the patient to the laboratory 3rd rev Ed, Wein-heim, Wiley-VCH, 2003.

3. Internet version included expert forum, 2003:www.//diagnosticsample.com

4. Guder WG, Hagemann P, Wisser H, Zawta B. FokusPatientenprobe. Kompendium Praanalytik. CD-RomBD Germany, Austria, Swizerland 2007.

STABILNOST UZORKA

\. Salvanjo

Odeljenje klini~ke hemije, Univerzitet u Veroni, Italija

Klini~ke laboratorije su se poslednjih godinana{le usred vrtloga promena koje su zadesile medi-cinu. Predla`u se i implementiraju razli~ite strategije ~ijije cilj kontrolisanje i smanjivanje ukupnih tro{kova lab-oratorijskih usluga. One uklju~uju centralizaciju, kon-solidaciju i integraciju usluga, rein`enjering laboratori-je i vi{i nivo automatizacije, optimalnu upotrebu testo-va, decentralizaciju postupka testiranja u smeru point-of-care testiranja. Rastu}i trend spajanja bolnica doveoje do konsolidacije laboratorijskih usluga iz logisti~kih iekonomskih razloga. [tavi{e, mnoge bolnice, klinike iprivatne prakse iz ekonomskih razloga praktikuju slan-je uzoraka na testiranje komercijalnim laboratorijama,po{to metode low-volume ili ezoteri~ni testovi nisuekonomski isplativi. U delovima zemlje gde su mre`ezdravstvenih ustanova jake sa agresivnim programimadelovanja zatvaraju se laboratorije u manjim bolnica-ma a testovi se {alju u centralnu laboratoriju. Zahvalju-ju}i izvanrednom porastu broja decentralizovanihslu`bi flebotomije {irom sveta, uzorci krvi ~ak ponekadsti`u u centralnu laboratoriju sa razli~itih udaljenosti,pod razli~itim uslovima skladi{tenja i transporta. Kaorezultat, centralne laboratorije koje opslu`uju mre`ezdravstvenih ustanova iskusile su porast radnog opte-re}enja i suo~ene su sa novim problemima u vezi sastabilno{}u uzorka. Nepostojanje standardizovanihpostupaka za uzimanje uzorka razlog je najve}eg brojagre{aka koje se doga|aju u celokupnom postupkutestiranja. One obuhvataju neodgovaraju}e postupkeza uzimanje i rukovanje uzorcima, kao {to je upotrebapogre{nih alatki, prolongiran zastoj u proticanju krvitokom se~enja vene, vreme pre centrifugiranja ili ana-lize i kona~no neodgovaraju}i na~in skladi{tenja. Oso-blje u laboratoriji ne mo`e u potpunosti da kontroli{eprobleme u pripremi i transportu uzoraka iz periferneustanove do centralizovane laboratorije. Neke indikaci-

202

je date su od strane Nacionalnog komiteta za stan-darde u klini~koj laboratoriji (NCCLS)/Instituta za stan-darde u klini~koj laboratoriji (CLSI) u pravilniku o uzi-manju uzoraka krvi za laboratorijsko testiranje, ali onene govore jasno da li uzorke treba skladi{titi kao celukrvi ili kao plazmu, a koliko smo mi upu}eni samo semali broj istra`ivanja bavio pitanjem stabilnosti uzorkau pogledu temperature i vremena pre centrifugiranjaza koagulaciju ili klini~ko-hemijsku analizu. Op{ti prob-lem s kojim se suo~avaju klini~ke laboratorije jesteintegritet necentrifugiranih uzoraka za hemijske anal-ize. Kako je produ`eni kontakt plazme ili seruma sa}elijama ~est razlog nejasnih rezultata testova, idealnobi bilo odvojiti plazmu i serum {to je pre mogu}e kakobi se spre~io metabolizam }elijskih konstituenata kao iaktivno i pasivno kretanje analita izme|u plazme ili se-ruma i }elijskih pregrada. Me|utim, testovi zgru{a-vanja su naro~ito podlo`ni lo{oj standardizaciji neko-liko postupaka u okviru celokupnog postupka testiran-ja i na njih verovatno vi{e uti~u uslovi skladi{tenja precentrifugiranja. U nau~noj literaturi ima izve{taja oizvesnom broju potencijalnih zamki u vezi sa uslovimaskladi{tenja i centrifugiranjem (temperatura i vreme)uzoraka cele krvi. Stabilnost uzorka je posebno kriti-~na, naro~ito kada uzorci krvi ne mogu biti analiziraniodmah ili kada je potrebno ponovno testiranje. Uprav-ljanje ovom preanaliti~kom varijablom (tj. stabilno{}uuzorka) smanjilo bi laboratorijske tro{kove i podstaklopoverenje izme|u lekara i osoblja u laboratoriji.

RAZVIJANJE LEAN FILOZOFIJE U KLINI^KIM LABORATORIJAMA

A. K. Stankovi}

BD Diagnostics – Preanalytical Systems, FranklinLakes, New Jersey, USA

Ovo je pregled principa Lean i Six Sigma kao sred-stava protoka operacija i pobolj{anja produktivnosti.Proizvo|a~ki sektori su iskoristili ove koncepte sa mnogouspeha. Primarni cilj lean inicijative je da se dajukvalitetni proizvodi i usluge odmah i uvek. Da bi se ovopostiglo, sve aktivnosti koje ne doprinose vrednosti (npr.tro{ak) treba da se elimini{u ili, ako to nije izvodljivo, dase smanje. Zahtevi dana{njeg sistema zdravstvene za-{tite opravdavaju integraciju sistema upravljanja kvalite-tom kao {to su oni za prihvatanje pove}anja radnogoptere}enja, deficit osoblja, i zahtev za brz obrt rezulta-ta uzorka. Ovaj ~lanak diskutuje Lean i Six Sigma strate-gije kao i njihove aplikacije za klini~ke laboratorije,specifi~no upotrebljavaju}i sredstva kao {to su 5S, Kai-zen Blitz, planiranje procesa i planiranje protoka vredno-

fully controlled by the laboratory staff. Some indica-tions were provided by the National Committee forClinical Laboratory Standards (NCCLS) / Clinical La-boratory Standard Institute (CLSI) guidelines on thecollection of blood specimens for laboratory testing,but they do not clearly stipulate whether samplesshould be kept as whole blood or plasma and to thebest of our knowledge, only few investigations havefocused on the sample stability in regard to the tem-perature and the time before centrifugation for coag-ulation or clinical chemistry testing. A general prob-lem facing clinical laboratories is the integrity ofuncentrifuged specimens for chemical analyses.Because prolonged contact of plasma or serum withcells is a common cause of spurious test results, plas-ma and serum should ideally be separated from cellsas quickly as possible to prevent ongoing metabolismof cellular constituents as well as active and passivemovement of analytes between the plasma or serumand cellular compartments. However, clotting assaysare particularly susceptible to poor standardization ofseveral process of the total testing process and theyare probably more influenced by the conditions ofstorage before centrifugation. Several pitfalls relatedto the storage condition and centrifugation (tempera-ture and time) of whole blood samples have beenpreviously reported in scientific literature (1–2). Sam-ple stability may be particularly critical, especiallywhen blood specimens cannot be analyzed immedi-ately or when retesting is required (3–4). The gover-nance of this preanalytical variable (i.e. sample sta-bility), would reduce laboratory costs and enhancethe physician-laboratory confidence.

DEVELOPING A LEAN CONSCIOUSNESS FOR THE CLINICAL LABORATORY

A. K. Stankovi}

BD Diagnostics – Preanalytical Systems, FranklinLakes, New Jersey, USA

This is an overview of the principles of Lean andSix Sigma as a means of streamlining operations andimproving productivity. Manufacturing sectors haveemployed these concepts with much success. The pri-mary goal of a lean initiative is to deliver quality prod-ucts and services the first time and every time. Toaccomplish this, all activities that do not add value (i.e.,waste) must be eliminated or, if not feasible, reduced.The demands of today's healthcare environment war-rant the integration of quality management systemssuch as these to meet increased workloads, staff short-ages, and the demand for rapid turnaround for speci-men results. This paper discusses Lean and Six Sigmastrategies as well as their application for the clinicallaboratory, specifically utilizing tools such as 5S, the

JMB 2008; 27 (2) 203

Kaizen Blitz, process mapping, and value stream map-ping. By implementing these tools to maximize processflow, eliminate waste, and recognize the variations thatcan hinder the delivery of high-quality services, health-care professionals can reach their efficiency goals,reduce costs, and provide customer satisfaction.

sti. Implementacija ovih sredstava maksimalno pove-}ava protok proces, elimini{e tro{kove, i prepoznaje va-rijacije koje mogu spre~iti pru`anje visoko-kvalitetnihusluga, zdravstvenim radnicima omogu}ava dostizanjeciljeva efikasnosti, smanjuje cenu, i obezbe|uje zado-voljstvo korisnika.

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Sekcija 4

BIOHEMIJSKIMARKERI

OBOLJENJA

Session 4

BIOCHEMICALMARKERS OF THE DISEASES

THE CLINICAL IMPACT OF WHO STANDARDIZATION OF PSA ASSAYS

J.-S. Blanchet, T. Brinkmann

Beckman Coulter Eurocenter, Nyon, Switzerland

The determination of serum level of the prosta-te specific antigen (PSA) is widely used for detectionand management of prostate cancer. Analytical vari-ability between the various PSA assays on the markethas been reported. This discrepancy in the PSAresults was shown to be related to non-equimolardetection of total PSA (tPSA) but also to a lack ofassay standardization and could have serious clinicalrepercussions on the diagnostic performance of PSAtesting. The recalibration of equimolar assays tocommon reference preparations (tPSA WHO96/670 and fPSA 96/668) was thought to promotestandardization of PSA assays and limit the clinicalimplication of assay variability. Comparison studieshave demonstrated that PSA assay calibration to theWHO standard certainly improves the harmonisationof PSA testing, but differences between assaysremain. Recent evaluations of the clinical impact ofanalytical variations induced by a calibration to theWHO standard showed that 15% to 30% of prostatecancer could be missed if the historical tPSA cut-offwas used. In order to avoid unacceptable erosion ofthe clinical diagnostic performance of PSA determi-nation for the detection of prostate cancer with WHOcalibrated assays, it is critical to define new specificclinical decision points.

SERUM CYSTATIN C CONCENTRATION IN TRANSPLANTED PATIENTS

TREATED WITH GLUCOCORTICOID IMMUNOSUPPRESSION

T. Gruev1, K. Chakalarovski2, N. Ivanovski2, A. Grueva2, S. Dimitrov3

1Institute for Clinical Biochemistry,University Clinical Centre, Skopje, Macedonia

2Department of Nephrology, University Clinical Centre, Skopje, Macedonia

3Health Center, Skopje, Macedonia

Cystatin C has been described as meeting manyof the characteristics of an ideal GFR marker and has

JMB 2008; 27 (2) 207

KLINI^KI UTICAJ WHO STANDARDIZACIJE NA PSA TESTOVE

J.-S. Blanchet, T. Brinkmann

Beckman Coulter Eurocenter, Nyon, Switzerland

Odre|ivanje nivoa antigena specifi~nog za pro-statu (PSA) u serumu ima {iroku primenu u otkrivanjui pra}enju toka bolesti kod raka prostate. Pokazalo seda izme|u razli~itih testova za odre|ivanje PSA koji po-stoje na tr`i{tu postoji analiti~ka varijabilnost. Utvr|enoje da je neslaganje u rezultatima PSA u vezi sa neekvi-molarnim odre|ivanjem ukupnog PSA (tPSA), ali i sanedostatkom standardizacije testova za odre|ivanje, imo`e imati ozbiljne klini~ke posledice na dijagnosti~kekarakteristike testiranja PSA. Veruje se da rekalibracijaekvimolarnih testova do uobi~ajenih referentnih stan-darda (tPSA SZO 96/670) i tPSA 96/688) poma`estandardizaciju testova za odre|ivanje PSA i ograni~avaklini~ke implikacije varijabilnosti testova. Uporednestudije pokazale su da kalibracija testova za PSA premastandardima SZO zna~ajno unapre|uje harmonizacijuanaliziranja PSA, ali razlike izme|u testova i dalje pos-toje. Nedavna ispitivanja klini~kog uticaja analiti~kihvarijacija zbog kalibracije prema standardu SZO poka-zala su da bi se moglo prevideti 15 do 30% slu~ajevaraka prostate ako bi se koristile zastarele »cut-off« vred-nosti. Kako bi se izbegla neprihvatljiva erozija klini~kedijagnosti~ke performase odre|ivanja PSA u cilju otkri-vanja raka prostate pomo}u testova kalibrisanih premaSZO, neophodno je odrediti nove specifi~ne ta~ke zaklini~ko odlu~ivanje.

KONCENTRACIJA CISTATINA C U SERUMU PACIJENATA POSLE TRANSPLANTACIJE I

TRETMANA IMUNOSUPRESIJE GLUKOKOR-TIKOIDIMA

T. Gruev1, K. Chakalarovski2, N. Ivanovski2, A. Grueva2, S. Dimitrov3

1Institut za klini~ku biohemiju, Univerzitetski klini~ki centar, Skoplje, Makedonija

2Odeljenje nefrologije, Univerzitetski klini~ki centar, Skoplje, Makedonija

3Dom zdravlja, Skoplje, Makedonija

Cistatin C opisan je kao marker koji posedujemnoge osobine idealnog markera za stopu glomeru-

UDK 577.1 : 61 ISSN 1452-8258

JMB 27: 207–210, 2008 Plenarne sekcije

Plenary sessions

been reported to be at least as accurate as the com-monly used serum creatinine to detect impaired renalfunction in various patient groups, including renaltransplant patients. The present study aimed to eluci-date the influence of glucocorticoid immunosuppres-sion on cystatin C concentration in the serum of renaltransplant patients. To evaluate the influence of immu-nosuppressive regimens, especially glucocorticoids, onserum cystatin C, 38 clinically stable patients onimmunosuppression therapy with low-dose glucocorti-coids were matched with 30 clinically stable patientsreceiving cyclosporin A alone and 18 clinically stablepatients receiving cyclosporin A together with azathio-prine. Clinical stability was defined as the absence ofacute rejection, febrile infection, and cyclosporin A tox-icity, as well as stability of creatinine clearance as esti-mated by the formula of Cockroft and Gault. The threegroups were matched for estimated creatinine clear-ance (CrCl) and had comparable gender, age, andtime since transplantation. To reduce the influence ofthe known biologic variation of cystatin C, all patientshad six measurements during subsequent visits thatdemonstrated a stable clinical condition. Means fromcystatin C reciprocals, as well as from CrCl estimatesand CysCGFR were calculated and used for dataanalysis. Furthermore, 10 patients receiving a shortcourse of high-dose methylprednisolone (500 mgintravenously per day for 3 days) for deteriorating renalfunction were analyzed to observe a possible dose-dependent effect of glucocorticoid administration. Thegroup receiving short-course, high-dose methylpred-nisolone had results from four time points available: a)before methylprednisolone commencement (median,15 days); b) the day methylprednisolone was started(before medication); c) after 3 days of methylpredni-solone therapy; and d) on a follow-up 9-10 days afterlast dose. Serum cystatin C was measured by a parti-cle-enhanced turbidimetric immunoassay (PETIA;Dako) on a Cobas Mira (Roche). Serum creatinine wasmeasured with a modified kinetic Jaffe method.Creatinine clearance was estimated by the formula ofCockroft and Gault, and the cystatin C GFR was meas-ured by the formula of Grubb (CysCGFR=84.69.CysC–1.680). Patients receiving long-term, low-dose glucocorticoid therapy showed higher cystatin Cconcentrations than controls (2.25; 1.9–2.9, P<0.05).High-dose methylprednisolone given intravenously ledto significant differences in cystatin C values at differ-ent time points (before administration, after threedoses, and 8 days after discontinuation; P<0.001).After three daily doses of 500 mg, cystatin C concen-trations increased from 2.13 mg/L (1.72–2.80) to2.69 mg/L (2.34–3.5; P<0.05). Eight days after dis-continuation, cystatin C concentrations significantlydecreased to 1.96 mg/L (1.63–2.4; P<0.05). Atthese time points, neither the CrCl estimate (54 ± 13mL · min–1 · 1.73 m–2, 51 ± 15 mL · min–1 · 1.73 m-2, and 56 ± 14 mL · min–1 · 1.73 m–2; P = 0.05). Theserum creatinine concentrations (165 mmol/L,158mmol/L, and 162 mmol/L, P=0.19) underwent signif-

larne filtracije (GFR) i pokazalo se da je u najmanjuruku precizan koliko i kreatinin u serumu koji se obi~nokoristi za otkrivanje oslabljene bubre`ne funkcije u razli-~itim grupama pacijenata, uklju~uju}i pacijente posletransplantacije bubrega. Cilj studije je da rasvetli uticajimunosupresije glukokortikoidima na koncentraciju cis-tatina C u serumu pacijenata posle transplantacije bu-brega. Kako bi se ispitao uticaj imunosupresivnih re`i-ma, naro~ito glukokortikoida, na cistatin C u serumu,oformljene su grupe od 38 klini~ki stabilnih pacijenatana imunosupresivnoj terapiji niskim dozama glukokor-tikoida, 30 klini~ki stabilnih pacijenata koji primajusamo ciklosporin A i 18 klini~ki stabilnih pacijenata kojiprimaju ciklosporin A zajedno sa azatioprinom. Klini~kastabilnost definisana je kao odsustvo akutnog odbijan-ja, febrilne infekcije i toksi~nosti ciklosporina, kao i sta-bilnost klirensa kreatinina izra~unatog pomo}u formuleKokrofta i Golta. ̂ lanovi tri grupe u kojima je odre|ivanklirens kreatinina (CrCl) birani su na osnovu pola, god-ina i vremena koje je pro{lo od transplantacije. Da bi sesmanjio uticaj poznatih biolo{kih varijacija cistatina C,kod svih pacijenata obavljeno je {est merenja tokomuzastopnih poseta koja su pokazala stabilno klini~kostanje. Srednje vrednosti cistatina C, kao i procenjenogCrCl i CysCGFR izra~unate su i kori{}ene u analizipodataka. Analiziranjem deset pacijenata koji primajukratkoro~nu terapiju visokim dozama metilprednizolona(500 mg dnevno intravenski tokom tri dana) za sman-jenu bubre`nu funkciju uo~en je potencijalni dozno-zavisni efekat administracije glukokortikoida. U grupikoja prima kratkoro~nu terapiju visokim dozama metil-prednizolona rezultati su odre|ivani na osnovu ~etiri do-stupne vremenske ta~ke: a) pre po~etka terapije metil-prednizolonom (medijana, 15 dana); b) na dan po~et-ka terapije metilprednizolonom (pre davanja leka); c)posle tri dana terapije metilprednizolonom; i d) na kon-troli 9–10 dana posle poslednje doze. Cistatin C u seru-mu odre|ivan je metodom particle-enhanced turbidi-metric immunoassay (»PETIA«, »Dako«) na analizatoru»Cobas Mira« (»Roche«). Kreatinin u serumu izmerenje modifikovanom kineti~kom Jaffeovom metodom.Klirens kreatinina izra~unat je pomo}u formule Ko-krofta i Golta, a cistatin C GFR pomo}u Grubbove for-mule (CysCGFR=84,69,CysC–1.680). Kod pacijenatakoji primaju dugoro~nu terapiju niskim dozama glu-kokortikoida otkrivene su vi{e koncentracije cistatina Cnego u kontrolnoj grupi (2,25; 1,9–2,9, P<0,05).Visoke doze metilprednizolona date intravenski dovelesu do zna~ajnih razlika u vrednostima cistatina C urazli~itim vremenskim ta~kama (pre administracije, po-sle tri doze, i posle 8 dana od prestanka terapije;P<0,001). Posle tri dnevne doze od 500 mg, koncen-tracije cistatina C sko~ile su sa 2,13 mg/L (1,72–2,80)na 2,69 mg/L (2,34–3,5; P<0,05). Osam dana posleprestanka terapije, koncentracije cistatina C su se zna-~ajno snizile na 1,96 mg/L (1,63–2,4; P<0,05). U timvremenskim ta~kama, ni procenjena vrednost ClCr(54±13 mL·min–1 · 1,73 m–2, 51±15 mL · min–1 ·1,73 m–2, i 56±14 mL · min–1 · 1,73 m–2; P=0,05),niti koncentracije kreatinina u serumu (165 mmol/L,

208

158 mmol/L, i 162 mmol/L, P=0,19) nisu se zna~ajnopromenile. Studija je pokazala da pacijenti posle trans-plantacije bubrega na terapiji glukokortikoidima imajuvi{i cistatin C nego preostale dve grupe na imunosu-presivnoj terapiji bez glukokortikoida. Kako su pacijentikoji primaju 500 mg metilprednizolona imali zna~ajnovi{e vrednosti cistatina C nego pacijenti koji primaju 10mg prednizona, mo`e se govoriti o dozno-zavisnom uti-caju datih doza glukokortikoida. Dakle, glukokortikoididovode to sistematski pogre{no procenjenih ni`ih vred-nosti stope glomerularne filtracije kod pacijenata posletransplantacije bubrega. Primena glukokortikoida kododraslih pacijenata posle transplantacije bubrega pove-zana je na dozno-zavisan na~in sa povi{enim cistatinomC, {to dovodi do sistematski pogre{nog utvr|ivanja ni`estope glomerularne filtracije. To ne isklju~uje mogu}-nost kori{}enja cistatina C u otkrivanju oslabljene bu-bre`ne funkcije kod pacijenata posle transplantacijebubrega na glukokortikoidima, po{to su ova i drugestudije pokazale da je cistatin C zna~ajno precizniji uotkrivanju oslabljene bubre`ne funkcije kod te grupepacijenata. [tavi{e, na{i podaci ukazuju na potrebnu zaspecifi~nim referentnim intervalima kod pacijenata naterapiji glukokortikoidima. U rutinskoj klini~koj praksi,kao i u budu}im klini~kim studijama, va`no je uzeti uobzir glukokortikoide prilikom tuma~enja koncentracijacistatina C u serumu pacijenata posle transplantacijebubrega a potencijalno i u ostalim grupama pacijenata.

ULOGA GLUTATION S-TRANSFERAZA U TUMORIMA URINARNOG TRAKTA

T. Simi}1, A. Savi}-Radojevi}1, M. Plje{a-Ercegovac1,M. Mati}1, T. Sa{i}1, D. Dragi~evi}2, J. Mimi}-Oka1

1Institut of biohemiju, Medicinski fakultet,Univerzitet u Beogradu, Beograd, Srbija

2Institut of nefrologiju i urologiju, Klini~ki centar Srbije, Beograd, Srbija

Jedan od etiolo{kih faktora za nastanak karcino-ma bubrega svetlih }elija i karcinoma prelaznog epitelamokra}ne be{ike je izlo`enost potencijalnim kancero-genima. Karcinom bubrega svetlih }elija pripada tu-morima sa visokim stepenom rezistencije na hemioter-apiju, a karcinom prelaznog epitela mokra}ne be{ikegrupi rekurentnih i multifokalnih tumora. Citosolneglutation S-transferaze (GST) su superfamilija enzimakoji u zdravim }elijama imaju za{titnu ulogu od poten-cijalnih kancerogena, katali{u}i reakcije konjugacijeelektrofilnih jedinjenja sa glutationom. Neki izoenzimiGST poseduju i hidroperoksidaznu aktivnost. Najboljeokarakterisane klase GST su Alfa (GSTA), Mi (GSTM),Pi (GSTP) i Teta (GSTT), a svaka od ovih klasa sadr`ivi{e razli~itih izoenzima. Uo~eno je nekoliko alelskihvarijacija unutar razli~itih klasa, usled kojih GSTM1nulti i GSTT1 nulti genotip imaju za posledicu odsus-

icant changes. This study demonstrates that renaltransplant patients receiving glucocorticoid medicationhave higher cystatin C than two comparable groupswith glucocorticoid-free immunosuppression. Becausepatients receiving 500 mg of methylprednisolone hadsignificantly higher cystatin C values than patients re-ceiving 10 mg of prednisone, a dose-dependent influ-ence of the administered glucocorticoid dose is sug-gested. Thus, glucocorticoid medication leads to sys-tematic underestimation of GFR in renal transplantpatients. Glucocorticoid medication in adult renal tran-splant patients is associated in a dose-dependent man-ner with increased cystatin C, leading to systematicunderestimation of GFR. This does not preclude theuse of cystatin C in detecting impaired renal functionin renal transplant patients with glucocorticoids, beca-use this study and others showed cystatin C to be sig-nificantly more accurate in detecting impaired renalfunction in this patient group. Moreover, our data illus-trate the need for specific reference intervals in patientson glucocorticoid therapy. In clinical routine settings, aswell as in future clinical studies, it is important to takeglucocorticoid medication into account when interpret-ing serum cystatin C concentrations in renal transplantpatients and presumably in other patient groups.

THE ROLE OF GLUTATHIONE S-TRANSFERASES IN URINARY

TRACT TUMORS

T. Simi}1, A. Savi}-Radojevi}1, M. Plje{a-Ercegovac1,M. Mati}1, T. Sa{i}1, D. Dragi~evi}2, J. Mimi}-Oka1

1Institute of Biochemistry, Faculty of Medicine,University of Belgrade, Belgrade, Serbia

2Institute of Nephrology and Urology, Clinical Centre of Serbia, Belgrade, Serbia

Exposure to potential carcinogens is among etio-logical factors for renal cell carcinoma (RCC) and tran-sitional cell carcinoma (TCC) of urinary bladder. RCCis very resistant, while TCC exhibits high recurrence ra-te and multifocality. Cytosolic glutathione S-transferas-es (GST) are a superfamily of enzymes which protectnormal cells by catalyzing conjugation reactions bet-ween electrophylic compounds, including carcino-gens, and glutathione. Some GST enzymes posses hy-droperoxidase activity. The most well characterizedclasses have been named Alpha (GSTA), Mu (GSTM),Pi (GSTP) and Theta (GSTT) and each of these class-es contain several different isoenzymes. Several typesof allelic variation have been identified within classes,among which GSTM1-null and GSTT1-null conferimpaired catalytic activity. Individuals with the GSTM1null genotype carry a substantially higher risk for blad-

JMB 2008; 27 (2) 209

der carcinogenesis. The effects of glutathione S-trans-ferase T1 polymorphism on the increased susceptibili-ty to RCC and TCC of urinary bladder depend on thepresence of specific chemical exposures to com-pounds metabolized via GSTT1-1 pathway. In the pro-cess of kidney cancerisation expression of GST alphaisoenzymes tends to decrease, consequently favoringprooxidant environment necessary for growth of RCC.GST pi enzyme activities are generally retained in RCCand might contribute to their chemotherapy resistanceof RCC. In the malignant phenotype of TCC of urinarybladder up regulation of various GST classes occurs.Up regulation of GSTT1-1 and GSTP1-1 might haveimportant consequences on the tumor growth, by pro-viding reduced environment and inhibition of apopto-tic pathways.

tvo kataliti~ke aktivnosti. Kod osoba koje su homozig-oti za GSTM1 nulti alel uo~en je zna~ajno ve}i rizik zanastanak karcinoma mokra}ne be{ike. Uticaj polimor-fizma GSTT1 na pove}anu osetljivost za nastanak kar-cinoma bubrega svetlih }elija i karcinoma prelaznogepitela mokra}ne be{ike u velikoj meri zavisi odizlo`enosti odre|enim hemijskim agensima koji semetaboli{u pomo}u GSTT1-1. U procesu nastankatumora bubrega dolazi do smanjenja ekspresije izoen-zima, pripadnika GST klase alfa, sa posledi~nompromenom redoks statusa, {to pogoduje razvoju karci-noma bubrega svetlih }elija. Enzimska aktivnost GSTpi je neizmenjena u karcinoma bubrega svetlih }elija imo`e doprinositi hemorezistenciji ovih tumora. U nas-tanku malignog fenotipa karcinoma prelaznog epitelamokra}ne be{ike zna~ajnu ulogu ima pove}anaekspresija razli~itih klasa GST. Pove}ana ekspresijaGSTT1-1 i GSTP1-1 mo`e zna~ajno uticati na rasttumora, pomeranjem ravnote`e izme|u pro- i antioksi-danasa ka redukovanom stanju, {to pogoduje inhibici-ji apoptotskih puteva.

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Sekcija 5

BIOHEMIJSKIMARKERI

OBOLJENJA

Session 5

BIOCHEMICALMARKERS OF THE DISEASES

FUNKCIONALNA HRANA-ULOGA U UNAPRE\ENJU ZDRAVLJA

I. Mileti}, S. [obaji}, B. \or|evi}

Institut za bromatologiju, Farmaceutski fakultet, Beograd, Srbija

Funkcionalna hrana je hrana koja ima povoljanuticaj na ljudsko zdravlje mimo uobi~ajenih nutri-tivnih funkcija. Biolo{ki aktivna jedinjenja su nosiocipovoljnog dejstva funkcionalne hrane. Brojni nau~nidokazi govore u prilog tome da je ishrana bogatapojedinim namirnicama (kao {to su to na primer vo}ei povr}e) direktno u vezi sa smanjenim rizikom odhroni~nih, nezaraznih bolesti, tako da se na tim saz-nanjima razvio koncept funkcionalne hrane. Otkrivajuse funkcionalne osobine tradicionalnih namirnica, alise dizajniraju i nove funkcionalne namirnice. Uobi~a-jene izjave koje prate tu vrstu namirnica mogu sesvrstati u dve kategorije: (1) izjave o odnosu struk-ture i funkcije (engl. Structure and function claims)moraju da budu istinite i da ne dovode u zabludupotro{a~a. Te izjave ne moraju da budu odobrene odstrane FDA. (2) Zdravstvene izjave (engl. Healthclaims ili disease-specific claims) moraju da buduautorizovane od strane FDA i da poseduju zna~ajnunau~nu potvrdu (Hillovi kriterijumi). Neophodno jerangiranje dokaza razli~itih tipova studija kojepodr`avaju zdravstvenu izjavu. Veliki broj biolo{kiaktivnih jedinjenja su nestabilna tokom tretmana i~uvanja. Ona podle`u mnogobrojnim hemijskimreakcijama, kao {to su to oksidacija, hidroliza, ter-mi~ka degradacija i Maillardova reakcija, {to rezultirasmanjenjem bioiskoristljivosti. Povoljan efekat biolo-{ki aktivnih jedinjenja direktno zavisi od primenjenogtretmana.

THE FUNCTIONAL FOODS – THE ROLE IN HEALTH IMPROVEMENT

I. Mileti}, S. [obaji}, B. \or|evi}

Institute of Bromatology, Faculty of Pharmacy, Belgrade, Serbia

Functional foods are foods that may provide ahealth benefit beyond basic nutrition. Numerous sci-entifically proven pieces of evidence in many epi-demiological studies indicate that nutrition abundantin certain foods (e.g. fruits and vegetables) is directlycorrelated with a decreased risk of degenerative dis-eases. Biologically active compounds in functionalfoods may impart health benefits or desirable physio-logical effects. Functional attributes of many tradi-tional foods are being discovered, while new foodproducts are being developed with beneficial compo-nents. These results are closely related to nutrition'spotentials in preventing chronic diseases. Based onthese facts the concept of functional foods has beendeveloped. Rigorous scientific investigation has toconfirm the positive physiological effects of thesecompounds upon health. Labeling claims that areused on functional foods are of two types: (1) struc-ture and function claims, which describe effects onnormal functioning of the body, but not claims thatthe food can treat, diagnose, prevent, or cure a dis-ease (claims such as »promotes regularity«, »helpsmaintain cardiovascular health«, and »supports theimmune system« fit into this category); and (2) dis-ease-risk reduction claims, which imply a relationshipbetween dietary components and a disease or healthcondition. Structure and function claims do notrequire preapproval by the FDA, and they requiremuch less stringent scientific consensus than disease-risk reduction claims. Many biologically active com-pounds are unstable during treatments and storage.They undergo many common chemical reactionssuch as oxidation, hydrolysis, thermal degradationand Maillard reaction, contributing to the lowering ofbioavailability. Anyhow, beneficial effect of bioactivecompounds depends directly on the applied treat-ment in the production of foods.

JMB 2008; 27 (2) 213

UDK 577.1 : 61 ISSN 1452-8258

JMB 27: 213–218, 2008 Plenarne sekcije

Plenary sessions

THE SECRET LIFE OF FAT: WHAT FAT CELLSDOING FOR REGULATION OF METABOLISM

A. Tzontcheva

Medical University of Sofia, Chair of Clinical Laboratoryand Clinical Immunology, Sofia, Bulgaria

Adipose tissue has long been regarded as anorgan the sole purpose of which was to store excessenergy as triglycerides, and release energy as freefatty acids, which itself is an essential self-defensesystem for survival during starvation. This point ofview has now changed, fat tissue has emerged as anendocrine and secretory organ affecting more thanone metabolic pathway. Its major endocrine functionis secreting several hormones, notably leptin andadiponectin. Also adipose tissue releases adipokinesinvolved in inflammation and hemostasis: growth fac-tors (TNFa, transforming growth factor-beta, nervegrowth factor, VEGF), cytokines (IL-1b, IL-6, IL-10),chemokines (IL-8), acute-phase proteins (haptoglo-bin, serum amyloid A) and prothrombotic factor(plasminogen activator inhibitor-1). This review aimsto present some of the recent topics of selectedadipokine research that may be of particular impor-tance.

CLINICAL IMPACT OF CYTOCHROME P4503A4 INHIBITION

B. Miljkovi}, S. Vezmar

Institute of Pharmacokinetics, Faculty of Pharmacy,University of Belgrade, Serbia

Cytochrome P450 3A4 (CYP3A4) is an enzymeinvolved in the metabolism of the majority of drugs.Induction or inhibition of the enzyme may induce re-duction or elevation of drug plasma concentrations. Ifplasma levels are significantly altered treatment effica-cy may be jeopardized or the patient may be at risk ofincreased toxicity. Clinically significant drug interac-tions which have the inhibition of CYP3A4 as theunderlying mechanism are likely to occur in drugs withnarrow therapeutic index or low bioavailability. Severalsevere adverse effects due to CYP3A4 inhibition havebeen reported. Among the most important are ventric-ular arrhythmias (torsades de pointes), rhabdomyoly-sis, excessive sedation, ataxia, ergotism and fatalhypotension. Drugs may also develop side effects afterCYP3A4 inhibition which normally do not occur suchas acute hepatic insufficiency after administration ofbudesonide. Also lack of treatment efficacy such asresistance to clopidogrel has been attributed toCYP3A4 inhibition. Nevertheless, drug interactionsmay also have a positive outcome, such as decrease ofnecessary dose of the high cost treatment with cy-

TAJNI @IVOT MASTI: [TA MASNE ]ELIJE^INE ZA REGULISANJE METABOLIZMA

A. Con~eva

Odeljenje klini~ke laboratorije i klini~ke imunologije,Medicinski univerzitet u Sofiji, Sofija, Bugarska

Adipozno tkivo se dugo smatralo organom ~ija jejedina svrha skladi{tenje vi{ka energije u obliku trigliceri-da i otpu{tanje energije u obliku slobodnih masnih kiseli-na, {to samo po sebi predstavlja neophodni sistem sa-moodbrane u cilju pre`ivljavanja tokom gladovanja.Takav stav je danas izmenjen, a masno tkivo se pokaza-lo kao endokrini i sekretorni organ koji uti~e na nekolikometaboli~kih puteva. Njegova glavna endokrina funkci-ja je lu~enje izvesnog broja hormona, pre svega leptinai adiponektina. Adipozno tkivo tako|e otpu{ta adipokineuklju~ene u inflamaciju i hemostazu: faktore rasta(TNFa, transformi{u}i faktor rasta-beta, nervni faktorrasta, VEGF), citokine (IL-1b, IL-6, IL-10), hemokine (IL-8), proteine akutne faze (haptoglobin, serumski amiloidA) i protrombotski faktor (plazminogen aktivatorainhibitor-1). Cilj ove studije je da predstavi odabranepredmete skora{njih istra`ivanja adipokina koji bi mogliimati poseban zna~aj.

KLINI^KI ZNA^AJ INHIBICIJECITOHROMA P450 3A4

B. Miljkovi}, S. Vezmar

Institut za farmakokinetiku, Farmaceutski fakultet,Univerzitet u Beogradu, Srbija

Citohrom P450 3A4 (CYP3A4) je enzim uklju-~en u metabolizam velikog broja lekova. Indukcija iliinhibicija enzima mogu uzrokovati sni`enje ili pove}a-nje koncentracija lekova u plazmi. Ukoliko su plazmakoncentracije lekova zna~ajno izmenjene mo`e bitiugro`ena efikasnost terapije ili pacijent mo`e imatipove}ani rizik od toksi~nosti. Klini~ki zna~ajne interak-cije lekova koje u osnovi imaju inhibiciju CYP3A4 naj-verovatnije }e se pojaviti kod lekova sa uskim terapi-jskim indeksom ili niskim stpenom bioraspolo`ivosti.Ozbiljna ne`eljena dejstva uzrokovana inhibicijomCYP3A4 su poznata. Me|u najzna~ajnijima se nalazeventrikularne aritmije (torsades de pointes), rabdomi-joliza, poja~ana sedacija, ataksija, ergotizam i fatalnahipotenzija. Zabele`ena su tako|e ne`eljena dejstvanakon inhibicije CYP3A4 koja se u odsustvu inhibitorane javljaju poput akutne hepati~ne insuficijencije kodprimene budesonida. Izostanak terapijske efikasnostido koje dolazi pri rezistenciji na klopidogrel se tako|epripisuje inhibiciji CYP3A4. Nasuprot tome, interakcijelekova mogu imati i pozitivan ishod poput sni`enjaneophodne doze skupe terapije ciklosporinom nakon

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inhibicije CYP3A4. Farmaceuti moraju imati aktivnuulogu u prevenciji ili potenciranju interakcija lekova, aza to moraju uzeti u obzir osobine lekova, karakteristikepacijenta i vreme primene lekova.

MALE NEKODIRAJU]E RNK – NOVINE U DIJAGNOSTICI I TERAPIJI

G. Koci}

Medicinski fakultet, Ni{, Srbija

Nekodiraju}e RNK predstavljaju specifi~nu kla-su RNK koje ne slu`e u procesu translacije proteina.One u~estvuju u nizu razli~itih regulatornih procesavezanih za proces transkripcije i posttranskripcioninivo. Pomenuta familija RNK sadr`i razli~ite tipove,ali su gotovo sve oko 20–30 nukleotida duga~ke,nastale iz prekursora ve}e du`ine. Neke od njih, kao{to je siRNK (mala interferentna RNK) formiraju senakon razgradnje dvostrukospiralnih RNK (dsRNK) iudru`ene su sa virusnom infekcijom. Druge familije,poznate kao mikro RNK (miRNK), kodiraju se odstrane specifi~nih gena. Glavna funkcija je inhibicijatranslacije, razgradnja mRNK ili indukcija razgradnjemRNK, poznata kao uti{avanje mRNK. Prepoznava-nje target mRNK mogu}e je po principu komple-mentarnosti baza. Stepen komplementarnosti odre-|uje uspe{nost uti{avanja. Kori{}enje iRNK u anti-virusnom odgovoru predstavlja novi terapijski izazov.Dok neke siRNK imaju zadatak da suprimiraju razvojoboljenja (virus), druge su uklju~ene u razvoj huma-nih kancera. Stoga je njihova identifikacija va`na sadijagnosti~kog i terapijskog aspekta kod razli~itih ti-pova karcinoma i razli~itih stadijuma oboljenja. Nekitipovi miRNK su udru`eni sa razvojem degenerativnihi metaboli~kih oboljenja. S obzirom na to da pravilanrazvoj nervnog sistema, insulinska sekrecija, kao iva`ni metaboli~ki procesi zahtevaju preciznu kontroluna nivou ekspresije gena, kontrola stabilnosti itranslacije mRNK putem malih nekodiraju}ih RNKpredstavlja dominantnu regulaciju razli~itih metabo-li~kih, endokrinih i neurolo{kih procesa.

closporine after CYP3A4 inhibition. Pharmacistsshould have an active role in preventing or potentiat-ing drug interactions and should therefore take intoconsideration drug properties, patient characteristicsand time of drug administration.

SMALL NON-CODING RNAs – DIAGNOSTIC AND THERAPEUTIC NEWS

G. Koci}

Medical Faculty, Ni{, Serbia

Non-coding RNAs represent a specific type ofRNAs that are non translated into proteins. They par-ticipate in a number of different regulatory processesconcerning the transcriptional and posttranscription-al level. This family contains different types, eachmainly 20-30 nucleotides long, excised from longerprecursors. Some of them, such as siRNAs (smallinterfering RNAs) are formed during longer double-stranded dsRNA cleavage associated with the viralinfection. Another family, named micro RNAs(miRNA) are encoded by specific genes. The mainfunction is to repress or degrade mRNA translation orto induce mRNA degradation, know as RNA silen-cing. They recognize target mRNA according to thesequence-specific base pairing. The degree of com-plementarity directs the outcome of the reaction.The use of RNAi as a defence against different virus-es was documented as a new therapeutic tool. SomesiRNAs have a potential of silencing disease (virus)specific RNAs, while miRNAs are involved in develop-ment of human cancers. Their profiling can be usedas diagnostic and prognostic tool in different cancertypes and different cancer stages. Some of themiRNA-associated genes are implicated in the deve-lopment of degenerative and metabolic diseases.Since the proper development of the nervous system,insulin secretion and important metabolic processesrequires precise control of gene expression, controlof the translation and stability of many mRNAs by thesmall non-coding RNAs is emerging as an importantregulator of various metabolic, endocrine and neuro-logical processes.

JMB 2008; 27 (2) 215

THE SIGNIFICANCE OF URINARY MARKERS IN THE

EVALUATION OF DIABETIC NEPHROPATHY

T. Cvetkovi}1,2, P. Vlahovi}2, V. \or|evi}1,3, L. Zvezdanovi}3, D. Pavlovi}1,

G. Koci}1, D. Sokolovi}1

1Institute of Biochemistry, Medical Faculty, Ni{, Serbia2Clinic of Nephrology, Clinical Centre, Ni{, Serbia

3Centre for Medical Biochemistry, Clinical Centre, Ni{, Serbia

Oxidative stress is considered to be a unifying linkbetween diabetes mellitus (DM) and its complications,including nephropathy (DN). The aim of this study wasto determine the parameters of oxidative injury of lipidsand proteins as well as the activity of ectoenzymes inthe urine of DN patients. The study included 40 indi-viduals: 10 patients with type 2 diabetes mellitus andmicroalbuminuria (DMT2-MIA), 10 type 2 diabeticpatients with macroalbuminuria (DMT2-MAA), 10 pati-ents with type 1 diabetes and microalbuminuria(DMT1-MIA) and 10 age- and sex-matched healthysubjects (control). In the urine we determined TBAreactive substances (TBARS), reactive carbonyl groups(RCG), and the activity of ectoenzymes N-acetyl-b-d-glucosaminidase (NAG), plasma cell differentiationantigen (PC-1), aminopeptidase N (APN) and dipep-tidyl peptidase IV (DPP IV). A higher concentration ofTBARS in the urine was found in DMT2-MIA andDMT1-MIA, compared to the control group (p<0.001and P<0.05). The urine concentration of RCD showssimilar results with a significant elevation in the groupswith DMT2-MAA and DMT1-MIA, compared to theDMT2-MIA (p<0.001) and control group (p<0.001).Activities of NAG, APN and DPPIV were significantlyhigher in the urine of DMT2-MAA, compared to thecontrol (p<0.01). The activity of PC-1 was slightlyincreased in that group, but not significantly. In conclu-sion, the level of oxidative stress markers and activitiesof brush border ectoenzymes in the urine may be a use-ful non-invasive and easily repeatable test in DN.

ZNA^AJ ODRE\IVANJA URINARNIH MARKERA U PRA]ENJU

DIJABETESNE NEFROPATIJE

T. Cvetkovi}1,2, P. Vlahovi}2, V. \or|evi}1,3, L. Zvezdanovi}3, D. Pavlovi}1,

G. Koci}1, D. Sokolovi}1

1Institut za biohemiju, Medicinski fakultet, Ni{, Srbija2Klinika za nefrologiju, Klini~ki centar, Ni{, Srbija

3Centar za medicinsku biohemiju,Klini~ki centar, Ni{, Srbija

Oksidativni stres mo`e se smatrati jedinstvenimfaktorom koji povezuje dijabetes melitus (DM) i nje-gove komplikacije, uklju~uju}i nefropatiju (DN). Ciljovog istra`ivanja bio je da se u urinu pacijenata sa DNodrede parametri oksidativnog o{te}enja lipida i pro-teina kao i aktivnost ektoenzima. Istra`ivanjem je obuh-va}eno 40 pacijenata: 10 pacijenata sa dijabetes meli-tusom tipa 2 i mikroalbuminurijom (DMT2-MIA), 10dijabeti~nih pacijenata tipa 2 sa makroalbuminurijom(DMT2-MAA), 10 pacijenata sa dijabetom tipa 1 i mi-kroalbuminurijom (DMT1-MIA) i 10 zdravih osoba(kontrola). U urinu je odre|ivana koncentracija TBAreaktivnih supstanci (TBARS), reaktivne karbonilnegrupe (RCG) i aktivnost ektoenzima N-acetil-b-d-gluko-zaminidaze (NAG), plazma }elijski diferenciraju}i anti-gen (PC-1), aminopeptidaza N (APN) i dipeptidil pep-tidaza IV (DPP IV). Ve}a koncentracija TBARS u urinuna|ena je u DMT2-MIA i DMT1-MIA grupi, u odnosuna kontrolu (p<0,001 i p<0,05). Koncentracija RCDu urinu pokazuje sli~ne vrednosti sa statisti~kizna~ajnim pove}anjem u DMT2-MAA i DMT1-MIAgrupi u odnosu na DMT2-MIA (p<0,001) i kontrolnugrupu (p<0,001). Aktivnost NAG, APN i DPPIVzna~ajno je ve}a u urinu pacijenata sa DMT2-MAA uodnosu na kontrolu (p<0,01). Aktivnost PC-1 pokazu-je lagani, ali ne i signifikantan porast u toj grupi pacije-nata. Zaklju~ujemo da pra}enje markera oksidativnogstresa i aktivnost brush border ektoenzima u urinumogu biti korisni, neinvazivni i lako primenljivi testovi upra}enju DN.

216

DIJAGNOSTI^KI ZNA^AJ ANTI-CIKLI^NIHCITRULINIRANIH PEPTIDNIH ANTITELA,AKTIVNOSTI ADENOZIN DEAMINAZE I

DRUGIH POTENCIJALNIH BIOMARKERA U OTKRIVANJU I PRA]ENJU REUMATOIDNOG ARTRITISA

J. Mehanovi}-Nikoli}1, J. Lalo{-Milju{2, M. Staj~i}-Nalesnik1, Lj. Laki}2, @. Bobi}3,

J. Bogdani}4, N. Vaji}4

1Medicinski fakultet, Banja Luka2Klini~ki centar, Banja Luka3Dom zdravlja Banja Luka

4Dom zdravlja Banja Vru}ica, Tesli},Bosna and Hercegovina

Cilj rada je bio utvr|ivanje prisustva anti-cikli-~nih citruliniranih peptidnih antitela (anti-CCP2) kodpacijenata sa i bez reumatoidnog artritisa (RA) i po-re|enje tih auto-antitela s drugim markerima kao {tosu reumatoidni faktor (RF) i C-reaktivni protein(CRP). Odre|ivana je kataliti~ka aktivnost adenozindeaminaze (ADA) u serumima pacijenata sa reuma-toidnim artritisom, bez terapije i sa terapijom meto-treksatom. Cilj rada bio je i utvr|ivanje mogu}nostiuvo|enja navedenih biohemijskih parametara udijagnostikovanje i pra}enje reumatoidnog artritisa una{u regiju. Anti-CCP2 su analizirana ELISA testom,kataliti~ka aktivnost ADA odre|ivana je spektrofo-tometrijskom metodom uz adenozin kao supstrat. RFje odre|ivan Latex testom. CRP je meren imunotur-bidimetrijskom metodom. Odre|ivanje anti-CCP2 jekorisno u dijagnostikovanju RA zbog visoke osetljivo-sti i specifi~nosti, me|utim, kombinovano odre|iva-nje anti-CCP2 sa RF je mnogo korisnije. Kataliti~kaaktivnost ADA u serumu mo`e da se koristi kao mark-er upalnog procesa u RA. Odre|ivanje CRP u serumupacijenata sa RA nema dijagnosti~ki zna~aj.

OKSIDATIVNI STRES – KLINI^KO-DIJAGNOSTI^KI ZNA^AJ

M. \uki}1, M. Ninkovi}2, M. Jovanovi}2

1Farmaceutski fakuletet Univarziteta u Beogradu2Institut za medicinska istra`ivanja,

Vojnomedicinska akademija, Beograd

Pove}ano stvaranje slobodnih radikala (SR) i/ilinedovljna antioksidativna za{tita stimuli{e }elijski odgo-vor na nastali oksidativni stres (OS). Produ`eni oksida-tivni insult ili u intezivnoj formi ostvaren prevazilazi }eli-jski odbrambreni kapacitet, dolazi do o{te}enja makro-molekula, gibi se }elijska funkicja, o{te}uju se mem-brane {to dovodi do smrti }elije. Oksidativni stres kaopatofiziolo{ki mehanizam je potvr|en u brojnim patolo-

THE DIAGNOSTIC VALUE OF ANTI-CYCLICCITRULLINATED PEPTIDE ANTIBODIES,ADENOSINE DEAMINASE ACTIVITY ANDOTHER POTENTIAL BIOMARKERS FOR

PREDICTING AND MONITORING RHEUMATOID ARTHRITIS

J. Mehanovi}-Nikoli}1, J. Lalo{-Milju{2, M. Staj~i}-Nalesnik1, Lj. Laki}2, @. Bobi}3,

J. Bogdani}4, N. Vaji}4

1Medical Faculty, Banja Luka2Clinical Center, Banja Luka3Health Centre Banja Luka

4Health Center Banja Vru}ica, Tesli},Bosnia and Herzegovina

The aim of this study was to investigate the pres-ence of anti-cyclic citrullinated peptide antibodies (anti-CCP2) in RA and non-RA patients and to evaluate thecombination of these auto-antibodies with some othermarkers such as RF and CRP. We examined the enzy-matic activity of ADA in RA patients without therapyand RA patients treated with methotrexate. Therefore,the aim of this study was also to assess the possibility ofintroducing these biochemical parameters in the diag-nosis and monitoring of the RA patients in this region.Serum antibodies directed to cyclic citrullinated pep-tide were analyzed using an anti-CCP2 antibody ELISA.Serum ADA activity was measured by a spectropho-tometer using adenosine as substrate (Giusti method).Rheumatoid factor (IgG-RF) was analyzed using a latexagglutination test. Serum CRP was measured by animmunoturbidimetric assay. The measurement of anti-CCP2 by itself, is useful for the diagnosis of RA for itshigh sensitivity and specificity, however, a combineduse of anti-CCP2 with RF is much more useful. SerumADA activity can be used as a biochemical marker ofinflammation in RA. Measuring CRP in the RA patientshas no diagnostic value.

OXIDATIVE STRESS – CLINICAL-DIAGNOSTIC SIGNIFICANCE

M. \uki}1, M. Ninkovi}2, M. Jovanovi}2

1Phaculty of Pharmacy at University of Belgrade2Institute for medical research Military Medical

Academy, Belgrade

Increased free radical (FR) production and/orinsufficient antioxidative defense stimulates cellular oxi-dant stress responses. Sustained oxidative insult or suf-ficient intensity overcome cell defenses, damage tomacromolecules can accumulate, leading to loss of cellfunction, membrane damage, and ultimately to celldeath. Oxidative stress (OS) as a pathophysiologicalmechanism is confirmed in numerous pathologies, poi-

JMB 2008; 27 (2) 217

sonings, ageing and body conditions such as enhancedphysical exercise, exposure to environmental pollutionand xenobiotics, smoking, etc. Principal reactive spe-cies, reactive oxygen species (ROS) and reactive nitro-gen species (RNS), endogenously or exogenously pro-duced, can readily attack all classes of macromolecules(proteins, DNA, unsaturated fatty acid). Perturbatedoxidative-reductive milieu, proceeded through lipid pe-roxidation development, changed activities of antioxi-dative enzymes (AOE) and depletion of non enzymaticendogenous antioxidants can be recognized in pre-symptomatic phase of many diseases, and thus mightbe marker of changed metabolic and functional disor-der. Therefore, in daily clinical- diagnostic practice, itwould be useful to routinely analyze OS parameters inorder to prevent and/or advance adequate diseasetreatment.

gijama, trovanjima, starenju i stanjima organizma kao{to su pove}ana fizi~ka aktivnost, izlo`enost zaga|enju~ovekove okoline, ksenobioticima, pu{enje itd. Najdo-minantnije reaktivne vrste u organizmu, reaktivne ki-seoni~ne vrste (RKV) i reaktivne azotove vrste (RNS),endogeno ili egzogeno stvorene, mogu lako da napad-nu sve klase biomolekula (proteni, DNK, nezasi}enemasne kiseline). Naru{en oksido-reduktivni milje, krozpove}anje lipidne peroksidacije, promenu aktivnostidirektnih ili indirektnih antioksidativnih enzima, kao ismanjenog sadr`aja neenzimskih antioksidanasa mo`ebiti prepoznat u presimptomatskoj fazi brojnih bolesti, iu tom smislu mo`e biti pokazatelj izmenjenih metabo-li~kih i funkcionalnih zbivanja. U svakodnevnoj klini~ko-dijagnosti~koj praksi analize parametara OS u biolo-{kom materijalu bi trebalo da imaju svoje mesto, radiprevencije bolesti i unapre|ivanja terapije bolesti.

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