Student’s Scientific Researches in the Field of Medical Biochemistry

and Pharmaceutical Science

Nau~no istra`iva~ki radovi studenata medicinsko biohemijskih

i farmaceutskih nauka

Predavanja / LecturesPosteri / Posters

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ASSOCIATION BETWEEN THE RS143383 MUTATION IN

THE GENE GDF5 AND THE DEVELOPMENT OF OSTEOPOROSIS

Marko @arak

Faculty of Pharmacy-Biochemistry in Zagreb andFaculty of Pharmacy in Ljubljana

Osteoporosis is a progressive, systemic meta-bolic disorder characterized by decreased bone mass,disturbed microarchitecture of the bones and in -creased bone brittleness, the consequence of whichis a risk of fractures. The disease results in frequentfractures of certain bones, most often compressivespinal fractures, femoral neck and distal radius frac-tures. Research has shown that osteoporosis is acomplex disease that is, along with specific etiologicfactors, also due to mutations in certain genes. TheGDF5 gene codes for a protein of the same name. Sofar, GDF5 mutations have been linked to the follow-ing diseases: acromesomelic dysplasia (Hunter-Thompson type), brachydactyly type C and chon-rodysplasia (Grebe type). The association with thesediseases suggests that the gene product of GDF5 isinvolved in skeletal development. GDF5 belongs tothe family of bone morphogenetic proteins involvedin the formation of cartilage and bone, and its defi-ciency may lead to the shortening of leg bones aswell as formation of bones with reduced density andincreased brittleness. The mutation rs143383 is asingle nucleotide polymorphism (SNP) in which cyto-sine (C) is replaced by thymine (T) within the 5'-UTRof GDF5. This polymorphism was first mentioned in2007 in relation to hip osteoarthritis in a Japanesepopulation. Later investigations showed a true asso-ciation between rs143383 and the development ofhip, knee and spinal osteoarthritis in Caucasian andAsian populations. Studies completed thus far haverevealed a close association between osteoarthritisand certain genetic mutations. One of the bestknown is the SNP rs143383 mutation in the GDF5gene. Since osteoarthritis and osteoporosis are bonediseases with similar clinical pictures, molecular andpathophysiological bases, it seems logical to assumethat the same specific genetic mutations are behindboth diseases. Therefore, the following hypotheseshave been proposed also as the aims of this paper: toprove the association of SNP rs143383 mutation in

POVEZANOST RS143383 MUTACIJE

GDF5 GENA S RAZVOJEM OSTEOPOROZE

Marko @arak

Farmaceutsko-biokemijski fakultet u Zagrebu iFarmaceutski fakultet u Ljubljani

Osteoporoza je progresivna, sistemska meta bo -li~ka bolest koju karakterizira smanjena masa ko{ta -nog tkiva, poreme}ena mikroarhitektura ko{tanestrukture te pove}anje lomljivosti kosti, {to za poslje -dicu ima rizik od nastajanja fruktura. Posljedica ovebolesti su ~esti lomovi pojedinih kostiju, naj~e{}ekom presivni prijelomi kralje{nice, prijelom vratabedrene kosti i distalne podlaktice. Dosada{nja istra -`ivanja pokazala su da je osteoporoza kompleksnabolest koja je uz odre|ene etiolo{ke ~imbenikeposljedica i mutacije pojedinih gena. GDF5 gen kodi-ra za protein istog imena. Dosada su mutacije GDF5gena povezane sa slijede}im bolestima: akrome-zomeli~na displazija (Hunter-Thompson tip), bahi-daktilija tipa C i hondrodisplazija (Grebe tip).Povezanost s ovim bolestima upu}uje da je genskiprodukt GDF5 gena uklju~en u razvoj skeleta. GDF5spada u porodicu ko{tanih morfogenetskih proteinakoji su uklju~eni u formaciju hrskavice i kosti, a u nje-govom nedostatku mogu} je razvoj skra}enih kostinogu te tvorba kosti slabije konzistencije i poja~anelomljivosti. Mutacija rs143383 je polimorfizam jednebaze (SNP) u kojem je zamijenjen citozin (C) s timi-nom (T) unutar 5'-UTR GDF5 gena. Ovaj polimor-fizam se prvi put spominje u vezi s osteoartritisomkuka u japanskoj populaciji 2007. godine. Kasnijaistra`ivanja pokazala istinitost povezanosti rs143383mutacije s razvojem osteoartitisa kuka, koljena i kra -lje{aka u populacijama bijelaca i azijata. Dosada{njaistra`ivanja pokazala su usku povezanost osteoartri -tisa s pojedinim genetskim mutacijama. Jedna odnajpoznatijih je SNP rs143383 mutacija GDF5 gena.S obzirom da su osteoartritis i osteoporoza ko{tanebolesti sli~nih klini~kih slika, molekularnih i patofizio-lo{kih osnova, logi~no je pretpostaviti da su odre|enegenetske mutacije jednake u pozadini obiju bolesti.Stoga su postavljena slijede}e hipoteze i ciljevi ovogarada: dokazati povezanost SNP rs143383 mutacijeGDF5 gena s razvojem osteoporoze u slovenskoj po -pulaciji, prona}i povezanost s promjenama BMD-a i

UDK 577.1 : 61 ISSN 1452-8258

J Med Biochem 34: 106–115, 2015 Lectures Predavanja

biokemijskih biljega ko{tane pregradnje te potvrditidosada{nje hipoteze o povezanosti SNP rs143383mutacije s razvojem osteoartritisa. U tu svrhu je mo -le kularnim tehnikama genotipizirano ukupno 900uzoraka od kojih je za 750 unaprijed postavljenadijagnoza osteoporoze, a za 150 osteoartritisa. Po -tencijalnim dokazivanjem povezanosti SNP rs143383mutacije GDF5 gena s razvojem osteoporoze, posta -vili bi se dokazi o novim patofiziolo{kim mehanizmi-ma te bi se olak{alo razumijevanje molekularnihosno va same bolesti. Doprinijelo bi se novim spozna-jama u vidu prevencije i lije~enja osteoporoze te otvo-rili novi aspekti u postavljanju hipoteza i daljnjemistra ̀ ivanju osteoporoze.

Klju~ne re~i: osteoporoza, mutacije GDF5, poli -morfizam

GDF58 with osteoporosis development in a Slavicpopu lation, to establish a correlation with thechanges in BMD and the biochemical markers ofbone turnover, and to confirm the current hypothesisof an association between the SNP rs143383 muta-tion and development of osteoarthritis. To this end,using molecular techniques, we genotyped a total of900 samples, of which in 750 a diagnosis of osteo-porosis and in 150 of osteoarthritis had been previ-ously established. Potential evidence of an associa-tion between the SNP rs143383 mutation in theGDF5 gene and the development of osteoporosiswould provide data about new pathophysiologicalmechanisms and contribute to the understanding ofthe molecular basis of this disorder. Also, it wouldenable new insights valuable for the prevention andtreatment of osteoporosis and open new aspects inproposing hypotheses and in further investigations ofosteoporosis.

Keywords: osteoporosis, GDF5 mutation, poly-morphism

J Med Biochem 2015; 34 (1) 107

ODRE\IVANJE ADIPONEKTINA,REZISTINA I IL-6 KOD PACIJENATASA KORONARNOM ARTERIJSKOM

BOLE[]U

Ana [ikora, Tijana Le~i}Mentor: prof. dr Vesna Spasojevi}

Kalimanovska, Sopi} Miron, Joksi} Jelena

Katedra za medicinsku biohemiju, Farmaceutski fakultet Univerziteta u Beogradu

Ateroskleroza je hroni~na inflamatorna bolestkoja se karakteri{e akumulacijom lipida i inflama-tornih }elija du` zidova arterija, i smatra se glavnimuzrokom koronarne arterijske bolesti (KAB). Rezistinje adipokin koji se primarno sekretuje iz monocita imakrofaga i poseduje zna~ajna proinflamatorna svo-jstva. Proinflamatorni medijatori, kao {to je IL-6 u invitro uslovima dovode do pove}anog lu~enja rezistinai vice versa. Adiponektin je adipokin koji se primarnolu~i iz adipocita, pobolj{ava osetljivost na insulin,poseduje zna~ajna antiaterogena i antiinflamatornasvojstva. Odre|ivanje nivoa rezistina, adiponektina iIL-6 kod pacijenata sa KAB-om u cilju rasvetljavanjanjihove uloge u razvoju bolesti. Adiponektin, rezistin iIL-6 su odre|ivani u plazmi 78 pacijenata sa KAB (41`ena i 37 mu{karaca), starosti od 32 do 82 godine,ELISA metodom. Pacijenti su na osnovu nalazakoronarografije razvrstani prema prisustvu (jednosu-dovna, dvosudovna, trosudovna KAB) i odustvu st e - noze. Dobijene vrednosti analizirane su neparame -tarskim Mann-Whitney testom i prikazane kao

ASSESSMENT OF ADIPONECTIN,RESISTIN AND IL-6 IN

PATIENTS WITH CORONARY ARTHERY DISEASE

Ana [ikora, Tijana Le~i}Menthor: prof. dr Vesna Spasojevi}

Kalimanovska, Sopi} Miron, Joksi} Jelena

Institute of Medical Biochemistry, Faculty of Pharmacy, University of Belgrade

Atherosclerosis is chronic inflammatory diseasecharacterized by the accumulation of lipids andinflammatory cells along the inner walls of arteries. Itis considered to be one of major causes of coronaryartery disease (CAD). Resistin, adipokine primarilysecreted by monocytes and macrophages, shows sig-nificant proinflammatory properties. Proinflam -matory mediators, such as IL-6 lead to increasedresistin secretion in vitro and vice versa. Adiponectin,adipocyte-derived protein, improves insulin sensitivityand shows important antiaterogenic and antiinflam-matory activity. Measurement of resistin, adiponectinand IL-6 levels in patients with CAD in order to assesstheir involvement in disease development. Resistin,adiponectin and IL-6 concentrations were measuredin plasma of 78 patients with CAD (41 female and37 male), aged from 32 to 82 year, using ELISAmethod. Patients have been classified by presence(single, double or triple-vessel stenosis) or absence ofcoronary stenosis, based on coronarography results.Results were analyzed using nonparametric Mann-

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Whitney test and shown as median values withinterquartile range. Resistin levels in patients withoutstenosis [11.8(9.4–17.9) ng/mL] were significantlylower compared with patients with stenosis [15.3(12.3–21.1) ng/mL], p=0,018, and compared withthose with triple-vessel stenosis [16.4(15.7–23.6)ng/mL], p=0,011. Adiponectin levels were signifi-cantly lower in patients with triple-vessel stenosis[11.2(8.9–13.6) ng/mL] compared with patientswithout stenosis [15.9(10.9–23.3) ng/mL], p=0.042.Patients on statin therapy also had significantly loweradiponectin levels [12.2(8.8–20.4) ng/mL], compa -red with those without therapy [16.1(11.1–24.9)ng/mL] p=0,025. Difference in IL-6 levels was notsignificant between patients with [1,21(0.79–7.25)pg/mL] and without stenosis [1.94(0.95–5.31)pg/mL]. Increased concentrations of proinflammato-ry resistin and decreased concentrations of vasculo-protective adiponectin indicate their potential role inatherosclerosis development. Decreased adiponectinlevels in patients on statin therapy suggest negativepleiotropic effect of these drugs. In this study directcorrelation between IL-6 and degree of the diseasehas not been proven.

Key words: resistin, adiponectin, IL-6

medijane sa interkvartilnim rasponima. Nivoi rezistinakod pacijenata bez stenoze [11,8(9,4–17,9) ng/mL]su bili zna~ajno ni`i u odnosu na sve pacijente sastenozom [15,3(12,3–21,1) ng/mL], p=0,018 i uodnosu na pacijente sa trosudovnom KAB [16,4(15,7–23,6) ng/mL], p=0,011. Koncentracija adi -po nektina je bila zna~ajno ni`a kod pacijenta sa tro-sudovnom stenozom [11.2(8.9–13.6) ng/mL] uodnosu na pacijente bez stenoze [15,9(10,9–23,3)ng/mL], p=0,042. Nivoi adiponektina su bili zna ~aj -no ni`i kod pacijenata na terapiji statinima [12,2(8,8–20,4) ng/mL], p=0,025 u odnosu na pacijentekoji nisu uzimali terapiju [16,1(11,1–24,9) ng/mL].Nivoi IL-6 se nisu zna~ajno razlikovali izme|u pacije-nata sa [1,21(0,79–7,25) pg/mL] i bez stenoze[1,94(0,95–5,31) pg/mL]. Pove}ane koncentracijeproinflamatornog rezistina i sni`ene koncentracijevaskuloprotektivnog adiponektina kod pacijenta saprisutnom stenozom ukazuju na njihovu mogu}u ulo -gu u razvoju ateroskleroze. Sni`ene vrednosti adipo -nektina kod pacijenata na terapiji statinima ukazujuna negativni plejotropni efekat ovih lekova. U ovomistra ̀ ivanju nije dokazana direktna povezanost IL-6 istepena razvoja bolesti.

Klju~ne re~i: rezistin, adiponektin, IL-6, KAB

POZIVANJE MEMORIJE U MORISOVOM VODENOM

LAVIRINTU U MODELU LIPOPOLISAHARIDOM-IZAZVANOGNEURORAZVOJNOG O[TE]ENJA

KOD PACOVA

Aleksandra Vidojevi}, Nikola Go{nji},Aleksandra @ivkovi}

Mentor: asist. dipl. farm. Bojan Batini}, dipl.farm. Branka Divovi}, prof. dr Miroslav Savi}

Katedra za farmakologiju, Katedra za fiziologiju,

Farmaceutski fakultet Univerziteta u Beogradu

Prenatalno izlaganje mladunaca pacova lipo -polisaharidu (LPS) i posledi~na inflamacija dovode doneurorazvojnih o{te}enja koja su prepoznata kaomogu}i animalni model {izofrenije. U ovom radusproveden je segment validacije LPS modela kroz testpozivanja memorije u Morisovom vodenom lavirintu(MVL). Mladunci pacova soja Wistar bili su prenatal-no izlo`eni LPS-u ili fiziolo{kom rastvoru (FIZ) u 15. i16. danu embriogeneze. Nakon ro|enja mu`jaci (M)su odvojeni od `enki (F) i formirano je osam grupa(n=11–12) u zavisnosti od pola, intrauterinog tret-mana i uzrasta u kome je bilo predvi|eno da, u okviruMVL eksperimenta, u|u u test pozivanja memorije i

MEMORY RETRIEVAL IN MORRIS WATER MAZE IN

THE RAT MODEL OF LIPOPOLYSACCHARIDE-INDUCED

NEURODEVELOPMENTAL IMPAIRMENT

Aleksandra Vidojevi}, Nikola Go{nji},Aleksandra @ivkovi}

Mentor: asist. dipl. farm. Bojan Batini}, dipl.farm. Branka Divovi}, prof. dr Miroslav Savi}

Department of Pharmacology, Department of Physilogy,

Faculty of Pharmacy University of Belgrade

Prenatal exposure to lipopolysaccharide (LPS)and consequential inflammation lead to neurodevel-opmental impairment in offspring rats that has beenrecognized as a putative animal schizophreniamodel. This study was conducted as a segment ofLPS model validation using memory retrieval test(probe trial) in Morris water maze (MWM). OffspringWistar rats were exposed to LPS or saline (SAL) atprenatal days 15 and 16. After birth, males (M) andfemales (F) were separated, and eight groups(n=11–12) were formed with respect to the sex,intrauterine treatment and the planned age for mem-ory retrieval testing, at postnatal days 40 and 60,

J Med Biochem 2015; 34 (1) 109

to 40. odnosno 60. dana postnatalno (P40, P60).Tokom pet dana eksperimenta `ivotinje su u~ile da uMVL-u prona|u platformu ispod povr{ine vode. Testpozivanja memorije je izvo|en {estog dana, bez plat-forme i analizirani su parametri deklarativne memo -rije: efikasnost putanje i latencija do ulaska u ciljnuzonu, kao i procenat tigmotaksi~nog kretanja kaoparametar proceduralne memorije. Kod P40 `ivoti njanije bilo zna~ajnih razlika u pra}enim parametrima.Studentov T- test je pokazao da su pacovi u grupiM/LPS/P60 imali statisti~ki zna~ajno manju efikas-nost putanje do ulaska u ciljnu zonu (r=0,015) kao ive}u latenciju do ulaska u ciljnu zonu (r=0,002) uodnosu na grupu M/FIZ/P60, dok u datim para-metrima me|u grupama F/LPS/P60 i F/FIZ/P60 nijebilo statisti~ke razlike. Trend pove}anja procenta vre-mena provedenog u tigmotaksi~noj zoni pokazan jepri pore|enju F/LPS/P60 sa F/FIZ/P60 (p=0,068),ali ne i pri pore|enju M/LPS/P60 sa M/FIZ/P60.Dobijeni rezultati ukazuju na to da se neurorazvojnao{te}enja u LPS modelu mogu manifestovati kogni-tivnim deficitima u periodu kasne adolescencije(P60). Mo`e se postaviti hipoteza da su mladuncipacova mu{kog roda podlo`niji LPS-indukovaniminflamatornim procesima u centralnom nervnom sis-temu, {to je moglo da rezultuje zapa`enim deficitommemorije.

Klju~ne re~i: lipopolisaharid, {izofrenija, neuro-razvojni model, MVL, pacov

respectively (P40, P60). During five days of Morriswater maze acquisition, animals have been learningto find a platform, hidden beneath water surface.The probe trial without platform was performed onthe sixth day. The analysis used included the declar-ative memory parameters: path efficiency to the tar-get zone and latency to the target zone, as well aspercentage of thigmotaxis locomotion, as a proce-dural memory parameter. P40 animals did not differin any of analyzed parameters. The Student’s t testshowed that M/LPS/P60 animals had significantlydecreased path efficiency to the target zone(p=0.015), and prolonged latency to the target zone(p=0.002), compared to M/SAL/P60 group, whilefor these parameters F/LPS/P60 and F/SAL/P60groups showed no statistical difference. A trend ofincrease in the percentage of time spent in thigmo-taxis area was shown for F/LPS/P60 in comparisonto F/SAL/P60 group (p=0.068), but not when com-paring male groups. The results demonstrate thatneurodevelopmental impairment in the LPS modelcan be manifested through cognitive deficits duringthe late adolescence (P60). We hypothesize that ratmale pups are more susceptible to the LPS-inducedinflammatory processes in central nervous system,which may have resulted in the observed memorydeficit.

Keywords: lipopolysaccharide, schizophrenia,neurodevelopmental model, MWM, rat

GENOTIPIZACIJA POLIMORFIZAMAU GENU OCT1 I NJIHOV ODNOS SA

TRETMANOM IMATINIBOM KODOBOLELIH OD HRONIČNEMIJELOIDNE LEUKEMIJE

Judita Avbelj, Irena Prodan, Barbara Ostanek

Farmaceutski fakultet, Univerzitet u Ljubljani

Imatinib je inhibitor tirozinske kinaze koji sekoristi za le~enje hroni~ne mijeloidne leukemije(HML) (1). Nedavno objavljeni podaci u literaturipokazuju da efikasnost le~enja imatinibom mo`dazavisi od aktivnosti prenosioca leka, transporteraorganskih katjona 1 (OCT1) (2). Cilj na{e studije bioje da se istra`i povezanost efikasnosti le~enja imati -nibom sa pet odabranih genskih polimorfizama inivoom genske ekspresije OCT1 gena SLC22A1 kodobolelih od HML. Na{a teza bila je da odabranipolimorfizmi u genu za OCT1 uti~u na aktivnostprenosioca imatiniba {to ima za posledicu slabijiodgovor na le~enje kod bolesnika sa varijantnim ale-lima u pore|enju sa subjektima bez mutacija. Tako|esmo `eleli da utvrdimo da li su za postizanje zna -

GENOTYPING POLYMORPHISMS IN OCT1 GENE

AND THEIR RELATION TO TREATMENT WITH

IMATINIB IN CML PATIENTS

Judita Avbelj, Irena Prodan, Barbara Ostanek

Faculty of Pharmacy, University of Ljubljana

Imatinib is a tirosine kinase inhibitor used fortreatment of chronic myeloid leukemia (CML) (1).The resently published literature data shows thattreatment efficasy of imatinib could depend on activ-ity of drug transporter, organic cation transporter 1(OCT1) (2). The aim of our study was to researchthe association of imatinib treatment efficacy withfive selected gene polymorphisms and the level ofgene expression of OCT1 gene SLC22A1 in CMLpatients. We postulated that the selected polymor-phisms in the gene for OCT1 affect the imatinibtransporter activity and result in poorer response totreatment in patients with variant alleles compared towild type individuals. We also wanted to determine, ifhigher doses of imatinib are needed to achieve the

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~ajnog molekularnog odgovora kod pacijenata samutacijama u genu OCT1 potrebne ve}e doze ima-tiniba. Genotipizacija je izvr{ena za 5 polimorfizamau genu OCT1: rs 12208357, rs72552763,rs34130495, rs628031 i rs683369 kori{}enjemtehnologije TaqMan. Aktivnosti prenosioca imatinibau granulocitima i mononuklearnim }elijama perifer nekrvi nisu se razlikovale izme|u genotipova ni u jed-nom slu~aju testiranih polimorfizama. Pored toga, naosnovu genotipova OCT1 za testirane polimorfizme,kod bolesnika nisu otkrivene zna~ajne razlike u po sti -zanju odnosno nedovoljnom postizanju mole kular -nog odgovora tokom 18 meseci. Me|utim, bolesnici-ma sa dva varijanta alela za polimorfizam rs683369bile su potrebne zna~ajno ve}e doze imatiniba da bise postigao zna~ajan molekularni od govor u pore -|enju s heterozigotima i subjektima bez mutacija.Prema na{im rezultatima, razlog za to nije uticaj naaktivnost prenosioca imatiniba. Mo`emo zaklju~iti danismo prona{li nikakvu vezu izme|u polimorfizma ugenu OCT1 i aktivnosti prenosioca imatiniba ili posti-zanja zna~ajnog molekularnog od go vora. U budu}imstudijama trebalo bi istra`iti mo gu}u povezanostizme|u rs683369 i doze imatiniba.

Klju~ne re~i: genotipizacija polimorfizma, ima-tinib, hroni~na mijeloidna leukemija

major molecular response in patients with mutationsin the OCT1 gene. Genotyping was performed for 5polymorphisms in the OCT1 gene rs12208357,rs72552763, rs34130495, rs628031 and rs683369using TaqMan genotyping technology. Activity ofimatinib transporter in granulocytes and peripheralblood mononuclear cells did not differ betweengenotypes for any of the tested polymorphisms. Inaddition, no significant differences in achievement /underachievement of molecular response within 18months were discovered between patients accordingto OCT1 genotypes for the tested polymorphisms.However, patients with two variant allels forrs683369 polymorhpism required significantly high-er doses of imatinib for reaching major molecularresponse compared to heterozygous and wild-typeindividuals. According to our results, this is not due toinfluence on imatinib transporter activity. In conclu-sion we did not find any association of polymorphismin the OCT1 gene with imatinib transporter activity orachievement of major molecular response. The pos-sible association of rs683369 with imatinib doseshould be investigated in future studies.

Keywords: genotyping, polymorphisms, Ima ti -nib, chronic myeloid leukemia

U^ESTALOST TRADICIONALNIH I NOVIH KARDIOVASKULARNIH FAKTORA RIZIKA U RAZLI^ITIMPATOFIZIOLO[KIM STANJIMA

POVEZANIM SA ATEROSKLEROZOM

Sonja Lazi}, Branko Pavlovi} Mentori: prof. dr Zorana Jeli}-Ivanovi},

doc. dr Aleksandra Zeljkovi}

Katedra za medicinsku biohemiju, Farmaceutski fakultet Univerziteta u Beogradu

Ishemijske bolesti mozga i srca predstavljajuvode}i uzrok smrtnosti u svetu. U njihovoj osnovi le`irazvoj ateroskleroze, pa je primarna prevencija orijen-tisana na identifikaciju faktora zna~ajnih za rano pre-poznavanje i procenu rizika za razvoj bolesti. Poredtradicionalnih faktora rizika (starost, pol, gojaznost,pu{enje, hipertenzija, hiperlipidemija), uvedeni su inovi parametri kao {to su veli~ina i raspodela subfrak-cije lipoproteina niske gustine (engl. Low densitylipoprotein, LDL) i lipoproteina visoke gustine (engl.High density lipoprotein, HDL). Ispitati u~estalosttradicionalnih i novih lipidnih i nelipidnih parametaraza razvoj ateroskleroze kod pacijenata sa ishemijskimbolestima srca i mozga i proceniti njihovu dijagnos-ti~ku ta~nost. U istra`ivanju je u~estvovalo 168 paci-

PREVALENCE OF TRADITIONAL AND NOVEL CARDIOVASCULAR

RISK FACTORS IN ATHEROSCLEROSIS

– RELATED DISEASES

Sonja Lazi}, Branko Pavlovi} Mentori: prof. dr Zorana Jeli}-Ivanovi},

doc. dr Aleksandra Zeljkovi}

Department of Medical Biochemistry, Faculty of Pharmacy, University of Belgrade

Ischemic heart and brain diseases are the maincauses of mortality worldwide. Since development ofatherosclerosis is a common basis for these condi-tions, primary prevention is oriented towards identifi-cation of risk factors which are useful in disease pre-diction and early recognition. Beside traditional riskfactors (age, gender, obesity, smoking, hypertension,hyperlipidemia), new parameters, such are size anddistribution of low-density lipoprotein (LDL) andhigh-density lipoproteins (HDL) subfractions, areinvestigated. To determine the prevalence of tradi-tional and new lipid and non-lipid cardiovascular riskfactors in patients with ischemic heart and brain dis-eases and to estimate their clinical accuracy. Thestudy involved 168 patients with coronary artery dis-

J Med Biochem 2015; 34 (1) 111

jenata sa angiografski dokazanom koronarnombole{ }u, 167 pacijenata sa akutnim ishemijskimmo` da nim udarom i 174 zdrava ispitanika. Svimispitanicima odre|ene su antropometrijske ka rakte -ristike, lipidni profil i raspodela lipoproteinskih sub-frakcija metodom elektroforeze na gradijentu poli-akrilamida. Dijagnosti~ka ta~nost je ispitivanametodom »Receiver Operating Characteristic (ROC)«krive. Pacijenti sa kardiovaskularnim bolestima (KVB)imaju zna~ajno vi{e vrednosti indeksa telesne mase ikoncentracije triglicerida (engl. TG), a statisti~ki zna -~ajno manje koncentracije HDL-holesterola, dijame -tar LDL i dijametar HDL ~estica u odnosu na kontrol-nu grupu. Pacijenti sa cerebrovaskularnim bolestima(CVB) imaju zna~ajno vi{e koncentracije TG i kon-centracije LDL-holesterola, a manje koncentracijeHDL-holesterola, dijametar LDL ~estica, te ve}i udeomalih HDL i LDL subfrakcija od zdravih ispitanika.U~estalost tradicionalnih faktora rizika je i kod KVB ikod CVB pacijenata ve}a u odnosu na zdrave ispi-tanike. ROC analizom smo utvrdili da se dodatkomdijametra LDL i HDL ~estica dijagnosti~ka ta~nosttradicionalnih faktora rizika zna~ajno pobolj{ala (po -vr {ina ispod ROC krive=0,900, p<0,05). U~estalosttradicionalnih lipidnih i nelipidnih faktora rizika kodKVB i kod CVB pacijenata je ve}a u odnosu na zdraveispitanike. Odre|ivanje LDL i HDL subfrakcija dopri-nosi dijagnosti~koj ta~nosti.

Klju~ne re~i: LDL ~estice, HDL ~estice, kardio-vaskularne bolesti, cerebrovaskularne bolesti

ease (CAD), 167 patients with acute ischemic stroke(AIS) and 174 healthy participants. Lipoprotein sub-fractions were assessed by polyacrylamide gradientgel electrophoresis. Clinical accuracy was tested withReceiver Operating Characteristic (ROC) curve. CADpatients have significantly higher values of body massindex and concentrations of triglyceride (TG), but sig-nificantly lower concentrations of HDL-cholesterol,and diameters of LDL particle and HDL particlescompared to the controls. AIS patients have signifi-cantly higher concentrations of TG and LDL-choles-terol, but lower HDL-cholesterol and LDL particle sizeand a larger proportion of small HDL and LDL sub-fractions than healthy subjects. Prevalence of tradi-tional risk factors is higher in CAD and AIS patientsthan in healthy subjects. ROC analysis showed thatby adding the diameter of LDL and HDL particlesclinical accuracy of traditional risk factors is signifi-cantly improved (area under the ROC curve =0.900, p <0.05). Prevalence of traditional lipid andnon-lipid risk factors is higher in patients withischemic heart and brain diseases than in healthysubjects. The determination of LDL and HDL sub-fractions improves clinical accuracy.

Keywords: LDL particles, HDL particles, cardio-vascular disease, cerebrovascular disease.

POVEZANOST SALIVARNIHBIOKEMIJSKIH BILJEGA STRESA

S PSIHOLO[KIM POKAZATELJIMA ISTUPNJEM TJELESNE AKTIVNOSTI

U STUDENTSKOJ POPULACIJI

Renata Lipovec, Marija Radi~ekMentor: doc. dr. sc. Nada Vrki}

Farmaceutsko-biokemijski fakultet, Domagojeva 2;KBC Sestre Milostrdnice, Vinogradska cesta 29;

10 000 Zagreb, Hrvatska

Slina kao uzorak je pogodna zbog jednos-tavnosti uzorkovanja, a u njoj se analiti nalaze u slo-bodnom tj. aktivnom obliku. Stres je stanje u kojemje poreme}ena psihofiziolo{ka ravnote`a organizma.Aktivira osovinu hipotalamus–hipofiza-kora nad-bubre`ne `lijezde pa je salivarni kortizol (sCORT)indikator slobodnog kortizola, dobro korelira saserumskom vrijedno{}u i odra`ava slobodnu frakcijukoja je biolo{ki aktivna. Istovremeno, stresna stanjastimuliraju simpati~ki neuronski sustav {to uzrokujepromjenu sekrecije salivarne a-amilaze (sAA) iz sali-

CORRELATION BETWEEN SALIVARYBIOCHEMICAL MARKERS OF

STRESS AND PSYCHOLOGICAL INDICATORS OF PHYSICAL ACTIVITYLEVELS IN STUDENT POPULATION

Renata Lipovec, Marija Radi~ekMentor: doc. dr. sc. Nada Vrki}

Faculty of Pharmacy and Biochemistry, University of Zagreb, KBC Sestre Milostrdnice,

Zagreb, Croatia

Saliva as a sample is suitable for easy samplingand analyts are found in free, active form. Stress is acondition in which is disturbed inner (psychophysio-logical) balance of the organism. Activates the hypo-thalamic – pituitary – adrenal cortex and salivary cor-tisol (sCORT) is an indicator of free cortisol, correlateswell with serum free value and the reflected fraction isbiologically active. At the same time, stressful situa-tion stimulate the sympathetic neural system whichcauses a change in the secretion of salivary alphaamylase from salivary gland (sAA). The hypothesis of

112

varnih `lijezda. Hipoteza ovog istra`ivanja bila je dasu studenti sa svakodnevnom intenzivnom fizi~komaktivno{}u izlo`eni kroni~nom stresu u odnosu nadruge studente koji imaju minimalne fizi~ke aktiv -nosti. Svi ispitanici najprije su ispunili psiholo{ki test,a potom su pristupiti davanju uzorka silne za odre -|ivanje koncentracija sCORT i sAA. Utvr|ena jeumjerena negativna povezanost izme|u koncentraci-ja sCORT i sAA kod svih studenata neovisno o fizi~kojaktivnosti, ali nije dokazana statisti~ka zna~ajnost.Najbolja povezanost koncentracije sCORT i sAA inaj~vr{}a statisti~ka zna~ajnost pokazala se u skupinistudenata s umjerenom fizi~kom aktivnosti. Aktivnijei pozitivnije suo~avanje sa stresnim stanjima zapa -`eno je kod studenata kineziologije na temelju psiho-lo{kog testiranja. Usporedbom biokemijskih i psiho-lo{kih pokazatelja u odnosu na stres zaklju~ujemo dasu dvije skupine studenata izjedna~ene unato~ bitnorazli~itim fizi~kim aktivnostima.

Klju~ne rije~i: slina, stres, kortizol, amilaza, psi-holo{ko testiranje

this study was that students with intensive daily physi-cal activity are exposed to chronic stress compared toother students who have a minimum of physical activ-ity. All participants first filled out a psychological test,and then gave saliva samples to determine the con-centration of sCORT and sAA. There was a weak tomoderate negative correlation between the concen-tration of salivary cortisol and sAA in all students, butnot statistically significant. The best correlationbetween cortisol and sAA and the strongest statisticalsignificance showed in a group of students who havemoderate physical activity. There was no statisticallysignificant correlation between biochemical indicatorsof stress and results obtained using psychological test-ing. Actively and positively coping with stressful situa-tions is observed in physical education students on thebasis of psychological testing, coping with stressful sit-uations. On a comparison of biochemical and physio-logical parameters in relation to stress, we concludethat the two groups of students are equal despite thevery different physical activities.

Keywords: saliva, stress, amilase, cortisol, psy-chological testing

BIOLO[KI U^INCI METIL-TRANSFERAZE RMTC

U BAKTERIJSKIM STANICAMAESCHERICHIA COLI DH5a

Andrea ^eriMentor: doc. dr. sc. Gordana Maravi}

Vlahovi~ek

Sveu~ili{te u Zagrebu, Farmaceutsko – biokemijski fakultet, Zavod za biokemiju i molekularnu biologiju, A. Kova~i}a 1,

10 000 Zagreb

Pojava otpornosti na antibiotike jedan je odnajve}ih problema u lije~enju bolesti uzrokovanihbakterijama zbog smanjenja broja djelotvornih lijeko-va. Jedan od mehanizama otpornosti je modifikacijaciljnog mjesta djelovanja lijeka. Enzimi metil-transfer-aze metiliraju 16S rRNA ribosoma te onemogu}ujuvezanje i djelovanje aminoglikozidnih antibiotika. Tajmehanizam uobi~ajen je za sojeve bakterija koji suprirodni proizvo|a~i antibiotika, ali se ubrzano {iri ikod patogenih bakterijskih sojeva otpornih na antibi-otike. Cilj ovog rada bio je ispitati biolo{ki u~inakmetil-transferaze RmtC iz porodice Arm u stanicamaE. coli DH5a. U tu svrhu provedeni su testovi kojimasu ispitane karakteristike rasta stanica s enzimom uodnosu na stanice bez enzima usporedbom generaci-jskih vremena i provedbom testa konkurencije rastastanica. b-galaktozidaznim testom ispitan je utjecaj

BIOLOGICAL EFFECTS OF METHYLTRANSFERASE

RMTC IN ESCHERICHIA COLI DH5a

Andrea ^eriMentor: doc. dr. sc. Gordana Maravi}

Vlahovi~ek

University of Zagreb, Faculty of Pharmacy andBiochemistry, Department of Biochemistry and

Molecular Biology, A. Kova~i}a 1, 10 000 Zagreb

Appearance of antimicrobial resistance is one ofthe greatest issues in treatment of diseases caused bybacteria due to reduction in number of effectiveantibiotics. One mechanism of resistance is modifica-tion of target sites of action of the drug. Enzymesmethyltransferases perform methylation of 16S rRNAin ribosomes and thereby inhibit binding and actionof aminoglycoside antibiotics. This mechanism iscommon for bacterial strains that are natural produc-ers of antibiotics, but it is spreading rapidly in patho-genic strains resistant to antibiotics. The aim of thiswork was to examine biological effects of methyl-transferase RmtC from Arm family in E. coli DH5a.To examine characteristic of growth of E. coli cellsexpressing RmtC in relation to cells without enzyme,determination and comparison of generation timeand growth competition assay were done. The effect

J Med Biochem 2015; 34 (1) 113

enzima na to~nost dekodiranja mRNA tijekom sin-teze proteina. Rezultati su pokazali da stanice E. coliDH5a koje eksprimiraju metil-transferazu RmtC rastusporije od stanica bez enzima. U uvjetima u kojima suse morale natjecati za hranjive tvari i `ivotni prostor,stanice s enzimom su pokazale da mogu konkuriratistanicama bez enzima nakon po~etnog perioda pri-lagodbe. Prisutnost metil-transferaze RmtC uzrokujeu~estalije pogre{ke prilikom dekodiranja na A- i P-mjestu ribosoma tijekom translacije. Prou~avanjemehanizama otpornosti od velike je va`nosti u borbiprotiv {irenja otpornosti na aminoglikozidne antibi-otike i pronalasku novih na~ina lije~enja bolestiuzrokovanim otpornim sojevima bakterija. Dobivenirezultati mogu poslu`iti kao podloga za daljnjaistra`ivanja, iako predstavljaju tek malen korak naputu za ostvarenje ovoga cilja.

Klju~ne rije~i: aminoglikozidni antibiotici,otpornost, metil transferaza RmtC

of enzyme to accuracy of decoding mRNA duringproces of protein translation was examined using b-galactosidase assay. Results showed that bacteria E.coli DH5a expressing methyltransferase RmtC growslower than cells without the enzyme. In conditionswhen they had to compete for nutrients and livingspace, cells with enzyme showed that they can com-pete with cells without enzyme but after initial adjust-ment period. The presence of methyltransferaseRmtC causes more frequent errors in decoding on P-and A-site during translation. The results of this workcan serve as a basis for further research, which couldhelp to find new opportunities in combating antibiot-ic resistance based on ribosomal RNA methyltrans-ferases.

Keywords: aminoglycoside antibiotics, resist-ance, methyltransferase RmtC

BIOHEMIJSKI MARKERIPOREME]AJA OKSIDATIVNO-

STRESNOG STATUSA KOD DECEKOJA @IVE U PETROHEMIJSKOJ

INDUSTRIJSKOJ SREDINI

Biljana Miljkovi}, Simona Tatovi}Mentor: Prof. dr Vesna Spasojevi}-Kalimanovska

Katedra za medicinsku biohemijuUniverzitet u Beogradu, Farmaceutski fakultet

Ko-mentor: Asist. dr Ana Nini} Katedra za medicinsku biohemiju

Univerzitet u Beogradu, Farmaceutski fakultetKo-mentor: Dipl. farm. med. biohem. Milica

Miljkovi}Katedra za medicinsku biohemiju

Univerzitet u Beogradu, Farmaceutski fakultet

Negativan efekat zaga|enog vazduha `ivotnesredine na krvne sudove smatra se va`nim uzro~ -nikom nastanka kardiovaskularnih bolesti. U posebnoosetljive grupe spadaju gerijatrijska i pedijatrijskapopulacija. Studija je sprovedena sa ciljem da bi seispitalo da li postoji uticaj zaga|enog vazduha `ivotnesredine na markere oksidativnog stresa (OS) i antiok-sidativne za{tite (AOZ) kao faktore rizika za razvojkardiovaskularnih oboljenja. Markeri OS statusa,lipid nog statusa, koncentracija glukoze i broj leukoci-ta su odre|ivani kod 40 zdrave, {kolske dece. Jednugrupu je ~inilo 20 dece (10 devoj~ica i 10 de~aka)uzrasta od 12 do 13 godina iz Pan~eva (najve}eindustrijske zone u Srbiji). Drugu grupu je ~inilo pre-ostalih 20 dece oba pola podjednako zastupljenih,uzrasta od 13 do 15 godina, iz Kova~ice (varo{ice

BIOCHEMICAL MARKERS OF OXIDATIVE STRESS STATUS

IN CHILDREN LIVING IN PETROCHEMICAL INDUSTRIAL AREA

Biljana Miljkovi}, Simona Tatovi}Mentor: Prof. dr Vesna Spasojevi}-Kalimanovska

Department of medical biochemistryUniversity of Belgrade, Faculty of Pharmacy

Co-mentor: Ass. dr Ana Nini}Department of medical biochemistry

University of Belgrade, Faculty of PharmacyCo-mentor: BSc. Milica Miljkovi}

Department of medical biochemistryUniversity of Belgrade, Faculty of Pharmacy

Negative effect of environmental air pollutionon blood vessels is considered to be an importantcause of cardiovascular diseases. Geriatric and pedi-atric populations are particularly vulnerable groups.This study was conducted in order to determine pos-sible influence of environmental air pollution onoxidative stress (OS) and antioxidative defense (AOD)status markers as risk factors for cardiovascular dis-eases development. OS and lipid status markers, glu-cose concentration and leukocytes count were meas-ured in the group of 40 healthy schoolchildren. Onegroup of 20 children (10 girls and 10 boys), whowere 12 to 13 years old, were from Pan~evo (thebiggest industrial area in Serbia). The second groupconsisted of the rest of 20 children both genderequally presented who were 13 to 15 years old from

114

udaljenoj 30 kilometara severno od Pan~eva).Odabrane grupe dece su bile homogene, tj. nisu pos-tojale statisti~ki zna~ajne razlike u ispitivanim marker-ima izme|u de~aka i devoj~ica u okviru svake grupe.Deca iz Kova~ice su imala zna~ajno ve}i indeks te -lesne mase (ITM) i bila su starija od dece iz Pan~eva.Statisti~ki zna~ajna razlika nije postojala u vrednosti-ma biohemijskih i lipidnih markera izme|u grupa.Pove}an nivo OS prikazan kroz pove}ane koncen-tracije tiobarbituratna kiselina-reaguju}ih supstanci(TBKRS) (p<0,001) utvr|en je kod dece koja su`ivela u industrijskoj u odnosu na decu koja su `ivelau ruralnoj sredini. Tako|e, smanjen nivo AOZ kaoposledica smanjene aktivnosti superoksid-dismutaze(SOD) (p<0,01) je pokazan kod dece koja su `ivelau industrijskoj sredini. Razlike u koncentraciji TBKRSi aktivnostima SOD su zadr`ale statisti~ku zna~ajnosti nakon analize kovarijanse za ITM i uzrast. Pove}annivo OS pokazan je kod dece koja su odrastala uindustrijskoj sredini. Dugotrajno izlaganje pove}animkoncentracijama zaga|iva~a u vazduhu predstavljapotencijalan faktor rizika po zdravlje dece.

Klju~ne re~i: oksidativni stres, antioksidativnaza{tita, SOD, TBKRS, zaga|enje vazduha

Kova~ica (village located 30 km north of Pan~evo).Groups of children were homogeneous; no signifi-cant differences were determined in tested markersbetween boys and girl within each group. Childrenfrom Kova~ica had statistically significant higher bodymass index (BMI) and they were older then childrenfrom Pan~evo. No statistically significant differenceswere found in biochemical and lipid status markersbetween groups. High level of OS demonstrated assignificantly higher concentration of tiobarbituricacid-reacting substances (TBARS) (p<0.001) wasfound in children from industrial environment com-paring to children from rural environment. Low levelof AOD demonstrated by lower superoxide dismutase(SOD) activity (p<0.01) was observed in group ofchildren from industrial area. Differences in TBARSlevel and SOD activity remained statistically signifi-cant after analysis of covariance for BMI and age.Increased level of OS was found in children who livedin an industrial area. Long-term exposition toincreased air pollutants concentrations presents apotential risk factor for children’s health.

Keywords: oxidative stress, antioxidativedefense, TBARS, SOD, air pollution

ZNA^AJ ODRE\IVANJA ADIPONEKTINA I REZISTINA KODPACIJENATA NA HEMODIJALIZI

Branislava [upljeglav, Jelena Bojovi}Mentori: prof. dr. Vesna Spasojevi}-

Kalimanovska, ass. Miron Sopi}, ass. Jelena Joksi}

Katedra za medicinsku biohemiju, Farmaceutskifakultet Univerziteta u Beogradu

Hroni~na bubre`na insuficijencija (HBI) je sin-drom koji nastaje kao posledica progresivnog,ireverzibilnog smanjenja glomerularne filtracije dokona~nog stadijuma uremije. Bolesnici sa HBI imajuznatno ve}i rizik od kardiovaskularne smrti nego {toje to slu~aj sa op{tom populacijom. Adiponektin jemultifunkcionalni protein adipocita sa antiatero -genom i antiinflamatornom aktivno{}u. Rezistin, kodljudi prona|en u }elijama bele loze, uklju~en je upatolo{ke procese poput inflamacije, endotelne dis-funkcije i angiogeneze. Odre|ivanje adiponektina irezistina u plazmi kod pacijenata na hemodijalizi (DP)i zdravih ispitanika u cilju razmatranja povezanostiovih parametara sa HBI. Adiponektin i rezistin suodre|ivani u uzorcima plazme kod ukupno 67 ispi-tanika od kojih su 33 DP (14 `ena, 19 mu{karaca) i34 zdrava ispitanika (18 `ena, 16 mu{karaca), sta -rosti 27–76 godina, ELISA metodom. Dobijene vred-

THE SIGNIFICANCE OFADIPONECTIN AND RESISTIN IN

PATIENTS ON HEMODIALYSIS

Branislava [upljeglav, Jelena Bojovi} Mentor: prof. dr Vesna Spasojevi}-Kalimanovska, ass. Miron Sopi},

ass. Jelena Joksi},

Institute for Medical Biochemistry, Faculty of Pharmacy, University of Belgrade

Chronic kidney disease (CKD) is a syndromethat occurs as a result of progressive, irreversibledecrease in the glomerular filtration rate until thefinal stage, uremia. The patients with CKD have asignificantly higher risk of cardiovascular death thangeneral population. Adiponectin is a multifunctionaladipocyte protein with antiatherogenic and anti-inflammatory properties. Resistin, in humans foundin white blood cells, is involved in pathologicalprocesses such as inflammation, endothelial dysfunc-tion and angiogenesis. Determination of adiponectinand resistin concentrations in patients on dialysis(PD) and healthy subjects in order to access the rela-tionship of these parameters with CKD. Adiponectinand resistin were measured in plasma of 67 patients,33 PD (14 women, 19 men) and 34 healthy subjects(18 women, 16 men), aged 27–76 years, usingELISA method. The obtained data were analyzed by

J Med Biochem 2015; 34 (1) 115

nosti su analizirane studentovim t-testom i prikazanekao srednje vrednosti sa standardnim devijacijama.Nivoi rezistina i adiponektina su bili zna~ajno vi{i kodDP (60,1±12,6 ng/mL, 16,0±8,4 ng/mL, respek-tivno) u odnosu na zdrave ispitanike (17,4±5,8ng/mL, p<0,001; 12,0±7,4 ng/mL, p=0,041). DPsu podeljeni u dve grupe prema vrednostima HDLholesterola (HDL-h) (grani~na vrednost: CHDL-h=0,91 mmol/L). Pacijenti sa ve}im vrednostimaHDL-h su imali zna~ajno vi{i nivo adiponektina uodnosu na DP sa ni`im vrednostima HDL-h(p=0,028). Kod DP pokazana je korelacija adiponek-tina sa HDL-h (r=0,618, p<0,001), trigliceridima(r=–0,457, p=0,007) i serumskim kreatininom(r=–0,368, p=0,035), a vi{estrukom regresionomanalizom pokazano je da je HDL-h jedini nezavisniprediktor adiponektina (b=0,618, adjR2=0,361,p<0,001). Razlike u nivoima rezistina i adiponektinau odnosu na zdrave ispitanike ukazuju na zna~aj kojiovi adipokini mogu imati u razvoju HBI. Pozitivnakorelacija adiponektina sa HDL-h i ve}e vrednostiadiponektina kod pacijenata sa ve}im HDL-h ukazu-ju na njegov potencijalno za{titni efekat kod pacijena-ta na hemodijalizi.

Klju~ne re~i: adiponektin, rezistin, hemodijaliza

Student’s t-test and presented as means with stan-dard deviations. Adiponectin and resistin levels weresignificantly higher in PD (60.1±12.6 ng/mL,16.0±8.4 ng/mL, respectively) compared withhealthy subjects (17.4±5.8 ng/mL, p<0.001;12.0±7.4 ng/mL, p=0.041). PD were divided intotwo groups according to the values of HDL-choles-terol (HDL-c) (cut-off value: CHDL-C=0.91mmol/L). Patients with higher HDL-c had a signifi-cantly higher level of adiponectin compared with DPwith lower HDL-c (p=0.028). In PD groupadiponectin correlated with HDL-c (r=0.618,p<0.001), triglycerides (r=–0.457,p=0.007) andserum creatinine (r=–0.368,p=0.035), and multipleregression analysis showed that HDL-c is the onlyindependent predictor of adiponectin (b=0.618,adjR2=0.361, p<0.001). Different levels of resistinand adiponectin found in PD compared to healthysubjects suggest the importance of these adipokinesin development of CKD. The positive correlation ofadiponectin with HDL-c, and higher values ofadiponectin in patients with higher HDL-c indicatespotential protective effect of adiponectin in patientson hemodialysis.

Keywords: adiponectin, resistin, hemodialysis

Poster Sessions

118

UDK 577.1 : 61 ISSN 1452-8258

J Med Biochem 34: 118–128, 2015 Poster sessionsPosterske sekcije

SP001 UPOTREBA QSAR STUDIJA U

DIZAJNIRANJU NOVIH DVOSTRUKIHINHIBITORA PI3K/MTOR KINAZE

Jelena Olui}, Katarina Nikoli}

Katedra za farmaceutsku hemiju, Farmaceutski fakultet Univerzitet u Beogradu

PI3K/mTOR kinaze spadaju u porodicu PIKKkinaza (kinaze povezane sa fosfatidilinositol-3-kina-zom). Signalni put PI3K/mTOR kinaze igra veomava`nu ulogu u }elijskom rastu, pre`ivljavanju i regu-laciji proliferacije. Uo~eno je da pokazuju znatnuhomologiju u strukturi svojih aktivnih mesta i izvesnobi mogle postati potencijalne mete za nove antineo-plastike. Kako bi se popravila in vitro biolo{ka aktiv -nost zajedno sa fizi~ko-hemijskim karakteristikama iodredile farmakokineti~ke karakteristike strukturaizabranih iz uzorka od 120 molekula, na osnovu po -zitivnih interakcija na enizimima PI3K/mTOR. Cilj jerazviti mo}nije 3D-strukture farmakofora za dvo stru -ke inhibitore PI3K/mTOR kinaze pomo}u kompjuter-ski pomognutog dizajna lekova. Prethodno formirani3D-QSAR modeli koriste se za predvi|anje aktivnostinovih, optimizovanih struktura, MarvinSketch, Chem-Office paket i Gaussian 98W bili su programi zacrtanje i optimizovanje molekula a za evaluaciju far-makokineti~kih parametara izabranog jedinjenja po -stojao je ADME/tox. Birana su i modifikovana jedi -njenja sa odgovaraju}im pIC50 vrednostima za PI3Ki mTOR kako bi se intenzivirale pozitivne interakcije.Pravilnim menjanjem hemijskih struktura u okvirusupstituenata, uz po{tovanje strategija u dizajnu leko-va, formiran je skup molekula sa sli~nim ili ~akpovi{enim aktivnostima. Poja~avanje aktivnosti bilo jeprioritet za mTOR kinaze; dok su aktivnosti za PI3Kkinaze pode{avane da budu stalne ili povi{ene.Upotreba QSAR studija u dizajniranju novih dvostruk-ih inhibitora PI3K/mTOR kinaze po kazale se koris-nom za analizu karakteristika i poprav ljanje biolo{kihaktivnosti jedinjenja. Ovo se mo`e ~initi kao dobarosnov za odabir potencijalnih kandidata za sintezu injihovu analizu u budu}im istra`iva njima.

Klju~ne re~i: 3D-QSAR, PI3K/mTOR kinaze,dvostruki inhibitori, dizajn lekova

SP001THE USAGE OF QSAR STUDIES IN

THE DESIGN OF NEW DUALPI3K/MTOR KINASE INHIBITORS

Jelena Olui}, Katarina Nikoli}

Department of Pharmaceutical chemistry, Faculty of Pharmacy University of Belgrade

PI3K/mTOR kinases belong to the PIKK (phos-phatidylinositol-3-kinase-related kinase) family ofkinases. PI3K/mTOR kinase signaling pathway playsa very important role in cell growth, survival and pro-liferation regulation. It has been noticed that theycontain a considerable homology in the structure oftheir active sites and there is a certain possibility forthem to become potential target places for new anti-neoplastics. To enhance in vitro biological activityaltogether with physicochemical characteristics andto determine the pharmacokinetic characteristics ofstructures chosen from a sample of 120 molecules,based on positive interactions on PI3K/mTORenzymes. The goal is to develop more potent 3D-structures of pharm cophores for dual PI3K/mTORkinase inhibitors with the help of computer-aideddrug design. Previously formed 3D-QSAR modelshave been used to predict the activities of new, opti-mized structures (Pentacle), MarvinSketch, Chem -Office package, and Gaussian 98W were tools fordrawing and optimizing the molecules, and for theevaluation of the chosen compound’s pharmacoki-netic parameters there was ADME/tox. Compoundswith adequate pIC50 values for PI3K and mTORwere chosen and modified in order to intensify thepositive interactions. With the correct alteration ofchemical structures within substituents, while res -pecting the strategies of drug design, a set of mo -lecules with corresponding or even higher activitieshas been formed. Enhancing the activities for mTORkinases was a priority; while activities for PI3K kina -ses were opt to be constant or higher. The usage ofQSAR studies in the design of new, dual PI3K/mTORkinase inhibitors was proven to be useful for theanalysis of characteristics and the improvement ofbiological activities of compounds. This may seem tobe a good basis for the choice of possible candidatesfor synthesis and their analysis in further research.

Keywords: 3D-QSAR, PI3K/mTOR kinases,dual inhibitors, drug design

J Med Biochem 2015; 34 (1) 119

SP002 TROVANJE ALKOHOLNIM PI]IMA

SA POVE]ANIM SADRŽAJEMMETANOLA

Marija Vidosavljevi}, Jovana Pelivanovi}Mentor: prof. dr Vesna Matovi}, asist. dipl.

farm. Aleksandra Buha

Katedra za toksikologiju »Akademik DaniloSoldatovi}«, Univerzitet u Beogradu-Farmaceutski

fakultet

Pove}ane koli~ine metanola u alkoholnimpi}ima mogu biti rezultat neadekvatne proizvodnjeprirodnih alhoholnih pi}a ili falsifikovanja, i pri tomemogu izazvati naj~e{}e masovna trovanja samogu}im te{kim efektima kao {to su o{te}enje vida iacidoza. Prema podacima SZO od 1998. do 2012.godine je zabele`eno ukupno 59 slu~ajeva masovnihtrovanja metanolom, a u Srbiji je 1998. godinezabele`eno ~ak 43 smrtna ishoda. Ispitivanje sadr`ajametanola i etanola u krvi pacijenata koji su konzumi-rali alkoholna pi}a. Uzorci krvi 14 pacijenata koji suprimljeni na VMA tokom 2013. godine u alkoholi -sanom stanju su analizirani headspace gasnom hro-matografijom sa plameno-jonizuju}im de tektorompod slede}im uslovima: kolona Rtx-BAC1, 30 m ×0,32 mm, debljina filma 1,8 mm, temperatura 40 °C,uz protok gasa 2 mL/min, a kao interni standardkori{}en je 1‰ rastvor 2-propanol. Du`ina trajanjaanalize iznosila je 5 minuta. Analizom uzoraka krviizmerene su koncentracije metanola u rasponu od0,1–0,26‰, dok su se koncentracije etanola kretaleod 0,80–5,18‰ {to ukazuje na alkoholisanost ali ipove}an unos metanola. Prema literaturnim podaci-ma, ve} niske koncentracije metanola u krvi (bez pris-ustva etanola) mogu rezultirati zna~ajnim toksi~nimefekatima, a koncentracije ve}e od 0,10‰ mogu iza-zvati i letalni ishod. Kod dva pacijenta su zabele`eninivoi metanola u krvi ve}i i od 0,20‰ (0,22 i0,26‰). U istim uzorcima krvi je i sadr`aj etanola bionajvi{i i iznosio je 4,52, odnosno 5,18‰. RezultatiNacionalnog Centra za kontrolu trovanja VMA,prikazani u ovom radu, ukazuju da je tokom 2013.godine registrovano vi{e slu~ajeva trovanja alkohol-nim pi}ima sa pove}anim sadr`ajem metanola, iakonije bilo smrtnih ishoda. Navedeni rezultati pokazalisu da alkoholna pi}a mogu sadr`ati pove}ane kon-centracije metanola {to potvr|uje zna~aj odre|ivanjametanola u krvi osoba u alkoholisanom stanju.

Klju~ne re~i: alkoholna pi}a, metanol, trovanje,headspace gasna hromatografija

SP002 POISONING WITH ALCOHOLIC

BEVERAGES CONTAININGINCREASED LEVELS OF METHANOL

Marija Vidosavljevi}, Jovana Pelivanovi}Mentors: prof. Dr Vesna Matovi}, mr pharm.

Aleksandra Buha

Department of Toxicology »Akademik DaniloSoldatovi}«, University of Belgrade-Faculty of

Pharmacy

The increased amount of methanol in alcoholbeverages can be the result of inadequate productionof natural alcoholic drinks or its falsification and cancause massive poisonings with possible severe toxiceffects such as visual impairments and acidosis.According to WHO 59 cases of massive methanolpoisonings were reported from 1998 to 2012, and inSerbia methanol caused death of 43 persons in1998. Analysis of methanol and ethanol contents inblood of patients who consumed alcoholic bever-ages. Blood samples of 14 alcohol-intoxicatedpatients who were admitted to the MMA during2013 were analyzed with headspace gas chromatog-raphy with flame ionization detector under the follow-ing conditions: Rtx-BAC1 column, 30 m × 0.32 mm,film thickness 1.8 mm, temperature 40 °C, gas flow 2mL/min, internal standard 1‰ 2-propanol. Run timewas 5 minutes. The concentrations of methanol inanalyzed blood samples were in the range between0.10 and 0.26‰ while ethanol concentrations were0.80–5.18‰ indicating increased intake of alcoholicbeverages which contained increased levels ofmethanol. According to literature data, rather lowmethanol levels (in absence of ethanol) can causedevelopment of significant toxic effects, and evenlethality with concentrations higher than 0.10‰. Twopatients had methanol concentrations even higherthan 0.20‰ (0.22 and 0.26‰). Furthermore, inthese samples ethanol levels were the highest as well,4.52 and 5.18‰ respectively. The results of theNational Poison Control Centre of MMA presented inthis study indicate that several cases of poisoningwith alcoholic drinks containing increased amount ofmethanol occurred during 2013, although with nodeath outcomes. Obtained results have shown thatalcoholic beverages can contain increased levels ofmethanol justifying the importance of methanolanalysis in the blood of alcohol-intoxicated patients.

Keywords: alcoholic beverages, poisoning,methanol, headspace gas chromatography

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SP003FORMULACIJA I PROCENA

VI[ESTRUKIH V/U/V EMULZIJA PRIMENOM

EKSPERIMENTALNOG DIZAJNA

Sergej JakimenkoMentor: Doc. dr Dragana Vasiljevi}

Katedra za farmaceutsku tehnologiju i kozmetologiju, Farmaceutski fakultet,

Univerzitet u Begradu

Multiple emulzije (v/u/v ili u/v/u) su kompleksnidisperzni sistemi, poznati i kao »emulzije emulzija«.Neke od prednosti vi{estrukih emulzija v/u/v tipa,koje ih ~ine pogodnim za primenu u farmaciji ikozmetologiji, su: relativno velika mogu}nost inkor-poriranja hidrofilnih supstanci, za{tita inkapsuliranihsupstanci od degradacije, mogu}nost inkorporiranjainkompatibilnih supstanci u isti sistem, kao i pro -du`eno osloba|anje aktivnih sastojaka. Me|utim,vi{e struke emulzije su termodinami~ki izuzetnonestabilni sistemi. Cilj ovog rada bila je procena uti-caja razli~itih formulacijskih faktora na reolo{kekarakteristike i fizi~ku stabilnost v/u/v vi{etrukihemul zija, primenom metode 24-1 frakcionog faktori-jalnog dizajna, kao i njihova optimizacija. Primenomdvostepenog postupka izra|eno je osam formulacijavi{estukih emulzija (F1 – F8), koje su se razlikovale ukoncentracijama: primarnog emulgatora PEG-30dipolihidroksistearata (1% i 3%), sekundarnog emul-gatora poloksamera 407 (0,8% i 1,2%), magnezijum-sulfata, heptahidrata (0,1% i 0,5%) i natrijum-hlorida(0,1% i 0,5%). Prisustvo slo`enih kapi je potvr|enoposmatranjem uzoraka opti~kim mikroskopom.Rezul tati, dobijeni 72h posle izrade (vrednosti elek-tri~ne provodljivosti, volumeni odvojenog sloja nakoncentrifugiranja, prividni viskoziteti i histerezne po vr{i -ne) su analizirani primenom softvera Design-Expert7.0. Druga faza eksperimentalnog rada bila jeposve}ena optimizaciji formulacija vi{estrukih v/u/vemulzija. Sve izra|ene vi{estruke emulzije bile suhomogene, bele boje, konzistencije kremova ililosiona. Vrednosti elektri~ne provodljivosti su bile urasponu od 113 mS/cm do 1563 mS/cm, a zapremi-na odvojeno g sloja nakon centrifugiranja bila je 0 do0,5 ml. Rezultati reolo{kih merenja su pokazali da sveispitivane emulzije ispoljavaju nenjutnovsko tiksotrop-no proticanje. Vrednosti maksimalnog prividnogviskoziteta su bile u rasponu od 421 mPas do 45433mPas. Karakterizacija i ispitivanje fizi~ke stabilnostioptimizovanih formulacija (izra|enih u skladu sapred logom softvera), dali su slede}e rezultate: nakontesta centrifugiranja nije uo~ena separacija faza,vrednosti elektri~ne provodljivosti bile su 158 mS/cmi 192 mS/cm, a maksimalnog prividnog viskoziteta19800 mPas odnosno 22300 mPas (za formulacije

SP003FORMULATION AND

EVALUATION OF W/O/W MULTIPLE EMULSIONS USING

EXPERIMENTAL DESIGN

Sergej JakimenkoMentor: Doc. dr Dragana Vasiljevi}

Department of Pharmaceutical Technology and Cosmetology, Faculty of Pharmacy,

University of Belgrade

Multiple emulsions (w/o/w or o/w/o) are com-plex dispersion systems, known also as «emulsions ofemulsions«. Relatively high entrapment capacity forhydrophilic compounds, protection of the encapsu-lated substances towards degradation, the ability tointroduce incompatible substances into the same sys-tem and sustained active substance release are someof the advantages of w/o/w type of these emulsionsystems that make them potentially interesting forapplication in pharmaceutics and cosmetics. The aimof this study was to evaluate significance of the influ-ence of different formulation parameters on the rhe-ological properties and physical stability of the w/o/wmultiple emulsions (using 24-1 fractional factorialdesign) as well as their optimization. Eight samples(F1 – F8) were prepared using two-step procedurevarying the following factors: concentrations of pri-mary emulsifier PEG 30-dipolyhydroxystearate (1%and 3%), secondary emulsifier Poloxamer 407 (0.8%and 1.2%), magnesium sulfate heptahydrate (0.1%and 0.5%) and sodium chloride (0.1% or 0.5%).Multiple emulsions were analyzed under microscopeto confirm the multiple characters. Results obtained72h after the preparation (values of electrical con-ductivity, the volumes of separate layers after cen-trifugation test, apparent viscosities and hysteresisarea) were analyzed using software Design-Expert7.0. In the second phase of experiments, optimiza-tion of the w/o/w multiple emulsions was performed.All prepared multiple emulsions appeared as whiteand homogenous creams or lotions. The conductivi-ty values were in range from 113–1563 mS/cm, andthe volumes of after centrifugation test were 0 to 0.5ml. Results of the rheological measurements haveshown that the investigated emulsions exhibited non-Newtonian thyxotropic behavior. The maximal appar-ent viscosities were from 421–45433 mPas. When itcomes to optimized formulations (which were pre-pared based on software recommendation), therewas no phase separation after centrifugation. Theconductivity values were 158 mS/cm and 192mS/cm; the maximal apparent viscosities were 19800mPas and 22300 mPas (for formulations F9 andF10, respectively). Based on the results obtainedusing the fractional factorial design it was concluded

J Med Biochem 2015; 34 (1) 121

F9 i F10, respektivno). Na osnovu rezultata, dobi-jenih primenom metode frakcionog faktorijalnogdizajna, mo`e se zaklju~iti da najve}i uticaj na reo -lo{ke karakteristike i fizi~ku stabilnost ispitivanihv/u/v vi{estrukih emulzija imaju koncentracije pri -mar nog emulgatora PEG-30 dipolihidroksistearata imagnezijum-sulfata, heptahidrata. Eksperimentalnodobijeni rezultati optimizovanih formulacija V/U/Vemulzija bili su veoma bliski predvi|enim, te primenametoda eksperimentalnog dizajna mo`e biti od veli -kog prakti~nog zna~aja u razvoju novih formulacijafarmaceutskih i kozmeti~kih emulzija.

Klju~ne re~i: v/u/v emulzije, polimerni emulga-tori, eksperimentalni dizajn

that the concentrations of primary emulsifier PEG 30-dipolyhydroxystearate and magnesium sulfate hepta -hydrate had the greatest influence on the rheologicalproperties and physical stability of the inves tigatedw/o/w multiple emulsions. Experi mentally obtainedresults of the optimized formulations were very closeto predicted values.Therefore, application of anexpe rimental design can significantly improve thedevelopment of pharmaceutical and cosmetic emul-sions.

Keywords: w/o/w emulsions, polymeric emulsi-fiers, experimental design

SP004 DEFINISANJE KINETIKE

REAKCIJE DEGRADACIJEMIKOFENOLAT MOFETILA

Jelena Zari}, Olivera @u`aMentori: Doc. dr Biljana Ota{evi},

Dr sc. Ana Proti}

Katedra za analitiku lekova, Farmaceutski fakultet Univerziteta u Beogradu

Mikofenolat mofetil je imunosupresiv koji senaj~e{}e koristi nakon transplantacije organa. Studijeforsirane degradacije izvode se prema smernicamaICH regulative Q1A. Procena uspe{nosti transferaHPLC na U-HPLC metodu mora biti u skladu sa Ph.Eur. 7 i USP 30. Skratiti vreme analize transferomHPLC na U-HPLC metodu, a zatim primeniti U-HPLCmetodu za pra}enje kinetike degradacije mikofenolatmofetila pri kontrolisanim stres uslovima. Kinetikareakcija degradacije mikofenolat mofetila ispitivanaje u baznoj sredini (0,01 mol/L NaOH), kiseloj sredi-ni (1 mol/L HCl), pod uticajem oksidativnog agensa(3 % H2O2) i na povi{enoj temperaturi (70 °C).Hromato grafski uslovi obuhvatali su: Atlantis C18(150 × 4,6 mm, 3 mm) kolonu, talasnu du`inudetekcije od 215 nm, mobilnu fazu (10 mmol/Lamonijum-acetat pH 6 : acetonitril (60 :40, v/v)) saprotokom od 0,7 mL min–1; Prilikom transfera na U-HPLC metodu kori{}ena je Hypersil Gold aq kolona(100 × 2,1 mm, 1,9 mm), protok je smanjen na 0,23mL min–1 i injekciona zapremina je smanjena sa 10na 1,39 mL. Izlaganje mikofenolat mofetila oksida-tivnom stresu dovodi do nastanka ~etiri mogu}adegradaciona proizvoda, zbog ~ega je transfermetode ra|en sa ovim uzorkom. Prilikom transferabilo je va`no da stacionarna faza ostane hemijski ista.Na osnovu svojih karakteristika, predlo`ena U-HPLCmetoda se mo`e smatrati novom metodom i zato

SP004 DEFINITION OF KINETIC

DEGRADATION OF MYCOPHENOLATE MOFETIL

Jelena Zari}, Olivera @u`aMentori: Doc. dr Biljana Ota{evi},

Dr sc. Ana Proti}

Department of Analytics of drugs, Faculty ofPharmacy, University of Belgrade

Mycophenolate mofetil is an immunosuppres-sant most used after organ transplantation. Forceddegradation studies are conducted by quidelines ofICH Q1A. Evaluation of the success of transfer fromHPLC to U-HPLC must be in accordance with the Ph.Eur. 7 and USP 30. Shorten the analysis time usingthe transfer from HPLC to U-HPLC, and then use U-HPLC method to follow the kinetics of degradation ofmycophenolate mofetil in controlled stress condi-tions. Kinetic of reaction of degradation of mycophe-nolate mofetil was tested in base environment (0.01mol/L NaOH), in acid environment (1 mol/L HCl), inthe conditions of oxidative stress (3% H2O2) and atelevated temperature (70 °C). Chromatographic con-ditions included: Atlantis C18 (150 × 4,6 mm, 3mm) column, wavelenght of detection in 215 nm,mobile phase (10 mmol/L mmonium-acetate pH 6 :acetonitrile (60 :40, v/v)) with flow of 0.7 mL min–1;During the transfer to U-HPLC method was usedHypersil Gold aq column (100 x 2.1 mm, 1.9 mm),flow was reduced to 0.23 mL min–1 and the injectionvolume was reduced from 10 to 1.39 mL. Exposureof myco phenolate mofetil to oxidative stress leads tothe creation of 4 degradation products which is whythe transfer of method was made with this sample.During the transfer, it was important that the the sta-tionary phase remains chemically the same. Basedon its characteristics, proposed U-HPLC method can

122

zahteva kompletnu validaciju. U-HPLC metodauspe{no je primenjena za pra}enje kinetikedegradacije mikofenolat mofetila. Transferom HPLCmetode skra}eno je vreme hromatografske analize sa22 na 6 minuta. U-HPLC metoda kori{}ena je zapra}enje kinetike degradacije mikofenolat mofetila.Na osnovu dobijenih rezultata mo`e se zaklju~iti damikofenolat mofetil prati kinetiku nultog reda prioksidativnoj degradaciji, prvog reda pri baznojdegradaciji i reakciju drugog reda pri kiseloj itermi~koj degradaciji.

Klju~ne re~i: mikofenolat mofetil, kinetika reak-cija degradacije, transfer HPLC u U-HPLC.

be regarded as a new method and therefore requiresthe complete validation. U-HPLC method has beensuccessfully applied to monitor the kinetics of degra-dation of mycophenolate mofetil. With the transfer ofHPLC method, was shortened the time of chromato-graphic analysis from 22 to 6 minutes. Based on theobtained results it can be concluded that themycophenolate mofetil follows zero-order reaction inoxidative stress degradation, first-order reaction inbase degradation, and second-order reaction in acidand thermal degradation.

Keywords: mycophenolate mofetil, kinetics ofdegradation reactions, HPLC to U-HPLC methodtransfer

SP005OKSIDATIVNO-STRESNI STATUS LDL LIPOPROTEINSKE FRAKCIJEKOD PACIJENATA SA AKUTNIM

INFARKTOM MIOKARDA

Milica \oki}, Katarina Joki}evi}Mentor: Dr sc. Jelena Kotur-Stevuljevi},

Doc. dr Aleksandra Zeljkovi}

Farmaceutski fakultet Univerziteta u Beogradu, Katedra za Medicinsku biohemiju

Oksidativna modifikacija lipoproteina male gus-tine (LDL) ima zna~ajnu ulogu u patogenezi atero -skleroze koja predstavlja komplikovan vaskularniporeme}aj i vode}i uzrok nastanka infarkta miokarda.Pra}enje efekata oksidativnog stresa na modifikacijuLDL ~estica kod pacijenata sa akutnim infarktommiokarda. U izolovanoj LDL frakciji seruma 16 paci-jenata sa akutnim infarktom miokarda i 10 zdravihosoba odre|ivani su parametari lipidnog statusa,oksidativnog stresa-totalni oksidativni status (TOS) itotalni antioksidativni status (TAS), kao i koncen-tracije malondialdehida (MDA), pre i posle oksidacijevodonik-peroksidom u prisustvu soli bakra. Pokret -ljivost LDL frakcija pre i nakon egzogene oksidacijedetektovana je metodom elektroforeze. Koncen tra -cija malondialdehida pre oksidacije LDL frakcije nijebila zna~ajno razli~ita kod pacijenata u odnosu nakontrolnu grupu, dok je koncentracija TAS ni`a.Nakon in vitro oksidacije LDL frakcije u obe ispitivanegrupe dolazi do zna~ajnog pove}anja koncentracijemalondialdehida, kao i do pada totalnog antioksi-dantnog statusa (p<0,001), pri ~emu je smanjenjekoncentracije TAS-a izra`enije kod zdravih ispitanika.Elektroforetskim merenjem pokretljivosti izolovanihLDL frakcija kod pacijenata sa akutnim infarktommiokarda i zdravih osoba utvr|ena je inicijalna zastu-pljenost oksidovanog LDL-a, kao i pove}anje njego -

SP005 OXIDATIVE STRESS STATUS

OF LDL LIPOPROTEIN FRACTION IN PATIENTS WITH ACUTE MYOCARDIAL INFARCTION

Milica \oki}, Katarina Joki}evi}Mentor: Dr. sc. Jelena Kotur-Stevuljevi},

Doc. Dr. Aleksandra Zeljkovi}

Faculty of Pharmacy, University of Belgrade,Department of Medical Biochemistry

Oxidative modification of low density lipopro-tein (LDL) plays an important role in the pathogene-sis of atherosclerosis, which is a complex vascular dis-order and a leading cause of myocardial infarction.Monitoring the effects of oxidative stress on the mo -dification of LDL particles in patients with acutemyocardial infarction. The LDL fraction was isolatedfrom serum of 16 patients with acute myocardialinfarction, and 10 healthy subjects. Lipid statusparameters, oxidative stress-total oxidative status(TOS) and total antioxidative status (TAS), also mal-ondialdehyde concentration (MDA) were determinedbefore and after hydrogen peroxide oxidation in thepresence of a copper salt. Mobility of LDL fractionsbefore and after oxidation was detected using elec-trophoresis. Malondialdehyde concentration beforeLDL fraction oxidation was not different comparing tocontrol group, while TAS concentration was signifi-cantly lower. After in vitro oxidation of LDL fraction inboth groups there was a significant increase in MDA,and a decline in the total antioxidant status(p<0.001), whereby the decrease in TAS is morepronounced in healthy subjects. he initial presenceof oxidized LDL, and its increase after the in vitro oxi-dation was determined measuring the electrophoret-ic mobility of isolated LDL fraction in patients withacute myocardial infarction and in healthy persons.

J Med Biochem 2015; 34 (1) 123

vog udela nakon in vitro oksidacije. Oksidovane LDL~estice pokazale su ve}u elektroforetsku pokretljivostu odnosu na neoksidovane. Migraciona daljinapove}ala se za 19% kod obolelih, a za pribli`no 40%kod kontrolne grupe. Na osnovu rezultata ovog ispi-tivanja mo`e se zaklju~iti da je kontrolna grupa sa -dr`ala ve}i udeo neizmenjenog LDL-a pa je naknad-na, egzogena oksidacija u ve}em procentu uticala namodifikaciju ovih ~estica. Manji procenat dodatnemodifikacije ovih ~estica kod pacijenata nakon oksi-dacije, ukazuje da su usled bolesti LDL ~estice ve}bile zna~ajno modifikovane endogenom oksidacijom.

Klju~ne re~i: infarkt miokarda, oksidovani LDL

Oxidized LDL particles showed a higher electro -phoretic mobility compared to unoxidized. Migrationdistance was increased by 19 % in the patients, andby about 40 % in the control group. Based on theresults of this study conclusion is that the controlgroup contains a higher amount of unmodified LDLand the subsequent, exogenous oxidation resulted ina higher percentage of particles’ modification. Lowerpercent of additional LDL particles modification inpatients’ group showed that these particles werealready endogenously modified because of the dis-ease process.

Keywords: myocardial infarction, oxidized LDL

SP006ZNANJE STUDENATA

U VEZI SA REGISTROM DAVALACA KOSTNE SR@I

SRBIJE

Jelena Lazarevi}Mentori: Du{anka Krajnovi}, Andrijana Milo{evi} Georgiev

Farmaceutski fakultet, Univerzitet u Beogradu, Srbija

Registar davalaca kostne sr`i (RDKS) sadr`ipodatke o osobama koje `ele da se prijave za poten-cijalne donore kostne sr`i. Trenutno je u RDKS pri-javljeno preko 4000 potencijalnih davalaca. Cilj jeispitati znanje, stavove i pona{anje studentske popu-lacije Farmaceutskog, Stomatolo{kog i Medicinskogfakulteta Univerziteta u Beogradu, Srbija, u vezi sadobrovoljnim davanjem kostne sr`i i postojanjemRDKS, uporediti dobijene rezultate me|u fakultetimazdravstvene struke i istra`iti mogu}e uzroke nedo-voljnog broja potencijalnih davalaca u RDKS. Ura |e -na je prospektivna studija preseka na uzorku studen-ta farmacije, medicine i stomatologije, kori{}e njemnamenski kreiranog upitnika. Anketiranje studenataje ura|eno na Univerzitetu u Beogradu. Rezultati suanalizirani deskriptivnom statistikom koriste}i MSExcel 2013. Anketirano je ukupno 574 studenata:195 (34%) studenata farmacije, 190 (33,1%) stude-nata medicine i 189 (32,9%) studenata stomatologi-je. Od anketiranih studenata, njih 320 (55,7%) znada postoji Svetski registar davalaca kostne sr`i(BMDW), dok za postojanje RDKS zna 141 (24,6%)student. Me|u anketiranim studentima bilo je i 199(34,7%) dobrovoljnih davaoca krvi, od kojih je~etvoro (0,7%) prijavljeno u RDKS, 32 (5,6%) stude-nata poznaje osobu iz okru`enja, upisanu u RDKS.Rezultati ukazuju da je samo 20,0% ispitanikarazmi{ljalo da se prijavi u RDKS, a 64,7% nije uop{terazmi{ljalo ili nije sigurno. Skoro ~etvrtina ispitanika

SP006BIOMEDICINE STUDENTS’

KNOWLEDGE RELATED TO BONE MARROW DONATION

REGISTRY IN SERBIA

Jelena Lazarevi}Mentors: Du{anka Krajnovi}, Andrijana

Milo{evi} Georgiev

University of Belgrade, Faculty of Pharmacy, Serbia

Serbian Bone Marrow Donor Registry (SBMDR)contains information concerning potential bone mar-row donors. Currently there are over 4000 potentialdonors listed in the SBMDR. The aim is to examinethe knowledge, attitudes and behavior of students inthe health professions at the University of Belgrade,Serbia, concerning bone marrow donation and theexistence of SBMDR. Also the aim is to compare theresults between different faculties and to discoverpossible causes of insufficient bone marrow donors inthe SBMDR. A prospective cross sectional study wasconducted among students of pharmacy, medicineand dentistry, using a specifically designed question-naire. Data collection was conducted at theUniversity of Belgrade. Results were analyzed bydescriptive statistics using MS Excel 2013. A total of574 students were asked to fill in the questionnaire:195 (34%) from the Faculty of Pharmacy, 190(33.1%) from the Faculty of Medicine and 189(32.9%) from the Faculty of Dentistry. Among sur-veyed students, 320 (55.7%) of them know thatthere is the Bone Marrow Donors Worldwide(BMDW), while 141 (24.6%) know of the existenceof SBMDR. Amid 34.7% (199 students) of study par-ticipants who are blood donors, only four (0.7%) arealso registered with SBMDR. Only 32 students(5.6%) know someone who is registered withSBMDR. Results point to the fact that only 20% ofinterviewees have considered registering with

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je izjavilo da bi voleli da budu kontaktirani od stranezaposlenih u Registru kako bi se prijavili. Naj~e{}irazlog za neprijaviljivanje u RDKS je strah od bola,kod 30,1% studenata. Veliki broj studenata je upoz-nat sa postojanjem Svetskog registra, a dosta manjesa postojanjem nacionalnog RDKS. Relativno malibroj studenata je razmiljao o prijavljivanju u isti.Po`eljno bi bilo da se rade promocije i edukacije ucilju informisanja i pro{irenja svesti o postojanju iva`nosti upisivanja u RDKS.

Klju~ne re~i: registar davalaca kostne sr`i, do -nori, studenti

SBMDR, while 64.7% have not at all considered orare uncertain about it. Almost a quarter of the sam-ple report they would like to be contacted in order toapply for registration. The most common reason fornot applying for registration with SBMDR is fear ofpain (30.1% students). A large number of studentsare acquainted with the existence of the BMDW,although far fewer know of the existence of theSBMDR. Relatively few students have consideredjoining the national registry. Promotion and educa-tion with aim to inform and increase awarenessabout the existence and importance of joining theSBMDR are highly desirable.

Keywords: bone marrow donor registry, donors,students

SP007 ISHEMIJOM MODIFIKOVAN

ALBUMIN KOD PACIJENATA SAAKUTNIM INFARKTOM MIOKARDA

Katarina Joki}evi}, Milica \oki}Mentor: Dr sc. Jelena Kotur-Stevuljevi},

dipl. farm.-med. biohem. Milica Miljkovi}

Farmaceutski fakultet Univerziteta u Beogradu, Katedra za Medicinsku biohemiju

Dijagnoza akutnog infarkta miokarda u hitnimslu~ajevima je ~esto te{ka zbog nejasne klini~ke slikei nedostatka brzog i pouzdanog dijagnosti~kog testa.Otud poti~e i potencijalni zna~aj ishemijom modifiko-vanog albumina (IMA) kao osetljivog biomarkeraishemije miokarda koji se pojavljuje pre razvojanekroze }elija sr~anog mi{i}a. Utvrditi zna~aj IMAkao potencijalnog biomarkera u ranom otkrivanjuinfarkta miokarda, kao i njegovu korelaciju sa para-metrima oksidativnog stresa. Kod 109 pacijenata saakutnim bolom u grudima i 60 zdravih ispitanikaodre|ivana je koncentracija IMA, metodom vezivanjakobalta za albumin. Ispitanicima je odre|ena i kon-centracija troponina I (TnI), kao i parametri oksida-tivno – stresnog statusa: totalni antioksidantni status(TAS), totalni oksidantni status (TOS), paraoksonaznaaktivnost (PON1), prooksidativno – antioksidativnibalans (PAB). Rezultati ovog ispitivanja pokazali suzna~ajno vi{e vrednosti koncentracije IMA (p<0,01)kod pacijenata u odnosu na zdrave osobe. Podelompacijenata prema koncentracijama troponina I, speci-fi~nog markera akutnog infarkta miokarda premagrani~nim koncentracijama za klini~ku odluku o pos-tojanju infarkta miokarda utvr|ene su vi{e koncen-tracije IMA kod pacijenata koji su imali klini~kizna~ajno vi{e koncentracije TnI, i ta razlika je bila sta-tisti~ki zna~ajna (p<0,05). Vrednosti PAB su bilezna~ajno vi{e kod obolelih pacijenata u pore|enju sa

SP007 ISCHEMIA – MODIFIED ALBUMIN

IN PATIENTS WITH ACUTE MYOCARDIAL INFARCTION

Author: Katarina Joki}evi}, Milica \oki}Menthor: Dr sc. Jelena Kotur Stevuljevi},

Ass. Milica Miljkovi}

Department of Medical Biochemistry, Faculty of Pharmacy, University of Belgrade

The diagnosis of acute myocardial infarction inemergency patients with acute coronary symptoms isoften difficult due to an unclear clinical presentationand lack of a rapid, reliable diagnostic test. There -fore, ischemia – modified albumin (IMA) might havepotential value as a sensitive biomarker of ischemia,possibly detected before myocardial cell necrosis. Todetermine the significance of IMA as a potential bio-marker in early myocardial infarction diagnosis andits correlation with oxidative stress status parameters.IMA concentration has been determined in 109patients with acute chest pain and 60 controls, bycobalt binding method. Also, troponin I (TnI) con-centration has been measured and oxidative stressparameters: total antioxidant status ( TAS), total oxi-dant status (TOS), paraoxonase (PON1), pro oxi-dant/antioxidant balance (PAB). Results have shownsignificantly higher IMA concentrations in patientscompared to control group (p<0.01). Dividingpatients according to troponin I concentration limitsfor establishing myocardial infarction diagnosis, high-er IMA concentrations have been established inpatients with significantly higher TnI concentrations.Furthermore, that difference has been significant(p<0.05). PAB values have been significantly higherin patients compared to control group (p<0.01), alsoTOS values (p<0.05). PON1 activity has been signif-icantly lower in patients (p<0.01), while TAS values

J Med Biochem 2015; 34 (1) 125

zdravim sredove~nim osobama koje su ~inile kontrol-nu grupu (p< 0,01), kao i vrednosti TOS (p<0,05).Aktivnost PON1 je bila zna~ajno ni`a kod pacijenata(p <0,01), dok se koncentracija TAS nije zna~ajnorazlikovala. Primenom Spearman–ove nepara-metarske korelacije utvr|ena je zna~ajna korelacijaizme|u koncentracija IMA kao markera ishemije ikoncentracije TOS kao markera prooksidativnihprocesa ( =+0,368, p<0,001). Na osnovu rezulta-ta ove studije mo`e se zaklju~iti da IMA nije markerspecifi~niji od klasi~nih biomarkera za akutni infarktmiokarda. Korelacija ovog parametra sa prooksidan-sima u krvi ukazuje na ulogu slobodnih radikala unjegovom nastanku.

Klju~ne re~i: ishemijom modifikovan albumin,infarkt miokarda

have not been significantly different. Spearman’s non–parametric correlation has shown significant corre-lation between IMA, marker of ischemia, and TOS,marker of pro oxidant processes, has been deter-mined (r=+0.368, p<0.001). Based on results ofthe current study it could be concluded that IMA isnot more specific marker than other classic myocar-dial infarction markers. The correlation of this mark-er with blood pro oxidants implies the role of free rad-icals in his producement in blood.

Keywords: ischemia – modified albumin,myocardial infaction

SP008 ZNANJA I STAVOVI STUDENATA

U VEZI SA FARMACEUTSKIMOTPADOM

Miona Mihajlovi}Mentori: doc. dr Du{anka Krajnovi}, stru~ni saradnik Jelena Manojlovi}

Katedra za socijalnu farmaciju i farmaceutsko zakonodavstvo, Univerzitet u Beogradu,

Farmaceutski fakultet

Farmaceutski otpad (FO) ~ine svi lekovi i medi-cinska sredstva zajedno sa primarnom ambala`om,koji su neupotrebljivi zbog isteka roka upotrebe, neis-pravnosti u pogledu njihovog kvaliteta, kontaminaci-je ili rasipanja. Ispitati znanja i stavove studenataBeogradskog univerziteta o pojmu FO, njegovimupravljanjem i ulozi studenata Farmaceutskog fakul-teta (FF) u ovoj oblasti. Deskriptivno istra`ivanje sanamenski kreiranim upitnikom za ovu studiju koji sudobrovoljno popunjavali studenti razli~itih godinaUniverziteta u Beogradu tokom meseca februara2014. godine. Od ukupnog broja ispitanih studenatanjih 198 je popunilo upitnik {to daje stepen odgovo-ra od 99%. Veoma mali procenat studenata, svega2%, zna {ta se podrazumeva pod pojmom FO, dokdaleko ve}i zna koje su boje kontejneri za odlaganjeFO (30,8%), koji je krajnji korak upravljanja konte-jnerima (25,3%). Svi studenti su jedinstvenog stavada je svest gra|ana u vezi sa FO nedovoljno razvijenaa velika ve}ina ispitanika (72,1 %) smatra da bi stu-denti FF trebalo da podsti~u aktivnosti u oblasti FO:edukacijom gra|ana (39,4%) i organizovnim prikupl-janjem FO (4,5%). Saznanja studenata u vezi saupravljanjem FO nisu dovoljno razvijena, te je potreb-no preduzimati edukacije studenata BeogradskogUniverziteta, a studenti FF bi mogli uzeti u~e{}e u

SP008 STUDENTS’ KNOWLEDGE AND ATTITUDES ABOUT

PHARMACEUTICAL WASTE

Miona Mihajlovi} Mentor: Doc. dr Du{anka Krajnovi},

Assist. Jelena Manojlovi}

Department of Social Pharmacy and Pharmaceutical Legislation,

University of Belgrade, Faculty of Pharmacy

Pharmaceutical waste includes all drugs andmedical devices, including primary packaging, unus-able due to their expiration date, failure in the termsof their quality, contamination or dissipation. Enquireinto the knowledge and attitudes of BelgradeUniversity students about the term of pharmaceuticalwaste, managing of it, and Faculty of Pharmacy stu-dents’ participation in this area. Materials and meth-ods: Descriptive research with purposefully madequestionnaire for this study which has been filled outvoluntarily by University of Belgrade students fromdifferent study levels, in February 2014. Out of thetotal number of the examinees 198 have fiiled outthe questionnaire which provides us with theresponse level of 99%. Very low percent of students,only 2%, know what the term of pharmaceuticalwaste implies, whereas a larger number know whatthe colour of containers for pharmaceutical wastedisposal is (30.8%), which is the last step of contain-ers managing (25.3%). All of them are unanimous intheir attitude that citizens’ awareness about pharma-ceutical isn’t developed well enough. Most of theexaminees (72.1 %) think that students of Faculty ofPharmacy should stimulate the activities in this area:with education of citizens (39.4%) and organizedpharmaceutical waste collection (4.5%). Students’

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programu edukacije op{te javnosti, koja premami{ljenju studenata nema dovoljno razvijenu svest oovom pitanju.

Klju~ne re~i: farmaceutski otpad, znanja istavovi, studenti farmacije

knowledge about pharmaceutical waste managingisn’t developed well enough, thus it’s necessary toeducate students of Belgrade Univesity, and studentsof Faculty of Pharmacy should be involved in educa-tion programs of general public.

Keywords: pharmaceutical waste, knowledgeand attitudes, pharmacy students

SP009 VALIDACIJA POPULACIONOG

FARMAKOKINETI^KOG MODELATOPIRAMATA

Majda Martinac, Jovan Savi}Mentor: prof. dr Branislava Miljkovi},

asist. Marija Jovanovi}

Katedra za Farmakokinetiku i Klini~ku farmaciju,Univerzitet u Beogradu, Farmaceutski fakultet

Populaciono modelovanje omogu}ava ispitiva njeuticaja faktora varijabilnosti na farmakokinetiku leka iposledi~no njegovu koncentraciju. Da bi mogao da sekoristi, populacioni farmakokineti~ki model je potreb-no validirati nekim od tehnika interne ili eksterne vali-dacije. Validacija populacionog farmakokineti~kogmodela topiramata radi njegove primene u individu -alizaciji re`ima doziranja. Model je izgra|en na poda -cima dobijenim od 49 odraslih pa cijenata sa epilep -sijom koji su na monoterapiji topiramatom ilikombi novanoj terapiji nekim drugim antiepileptikom.Validacija finalnog modela je vr{ena tehnikamainterne validacije, koje su obuhvatile: gra fi~ku pro-cenu (goodness of fit), numeri~ku (numerical predic-tive check, NPC) i vizuelnu procenu predikcije (visualpredictive check, VPC), kao i bootstrap analizu. Eks -terna validacija je ra|ena na setu poda taka dobijenomod 29 pacijenata. Odnos popu lacionih/individualnihpredvi|enih koncentracija i merenih koncentracijapokazuje visok stepen ko relacije. Tako|e, i grafik za -visnosti kondicionalnih ponderisanih reziduala (condi-tional weighted residuals, CWRES) u zavisnosti od vre-mena nakon primene leka je pokazao da su vrednostive}ine reziduala u rasponu od –2 do +2. Nume -ri~kom i vizuelnom procenom predikcije je pokazanoda se merene vrednosti nalaze uglavnom u okviruintervala pouzdanosti simuliranih podataka. Bootstrapanalizom nije pokazana zna~ajna razlika u srednjimvrednostima, standardnim gre{kama i 95% intervalupouzdanosti izme|u bootstrap replikata i originalnihpodataka. Eksternom validacijom je pokazana dobrakorelacija izme|u predvi|enih i merenih vrednosti.Rezultati potvr|uju da je model stabilan i da se mo`ekoristiti u identifikaciji i kvantifikaciji faktora farma ko -kineti~ke varijabilnosti sa ciljem njegove primene uindividualizaciji re`ima doziranja topiramata.

Klju~ne re~i: validacija, populaciono modelo-vanje, topiramat, NONMEM®

SP009 VALIDATION OF A POPULATION

PHARMACOKINETIC MODEL OF TOPIRAMATE

Majda Martinac, Jovan Savi}Mentor: Prof. dr Branislava Miljkovi},

Ass. Marija Jovanovi}

Department of Pharmacokinetics and ClinicalPharmacy, Faculty of Pharmacy

Population modeling allows investigation ofinfluence of variability factors on drug pharmacoki-netics and consequently on drug level. For the modelto be used, it is needed to perform its validation,using techniques of internal or external validation.Validation of the population pharmacokinetic modelof topiramate for its use in the individualization of thedosage regimen. Data were collected from 49 adultepileptic patients on mono- or co-therapy of TPMand other antiepileptic drug(s). The validation of thefinal model was performed by the techniques ofinternal validation, which included: goodness of fit,numerical predictive check, visual predictive check,and bootstrap method. For external validation wasused dataset from 29 patients. Population/individualpredicted concentrations correlated well with theobserved values. Also, conditional weighted residualsvs. time after dose plot showed that the most of theresiduals were within –2 to +2 range. Numerical andvisual predictive check showed that observed con-centrations were mostly within confidence intervals(CIs) for the simulated data. Bootstrap analysis didn’tshow any significant difference in mean values, stan-dard errors and 95% CIs. External validation showedgood correlation between predicted and observedvalues. The results confirmed that the model is stableand that it can be used in identification and quantifi-cation of pharmacokinetic variability factors, with agoal of individualisation of topiramate dosage regi-men.

Keywords: validation, population modeling, top-iramate, NONMEM®

J Med Biochem 2015; 34 (1) 127

SP010 ODRE\IVANJE AKTIVNOSTI

ENZIMA LECITIN-HOLESTEROLACILTRANSFERAZE (ENGL. LCAT)

I HOLESTEROL-ESTAR TRANS-FERNOG PROTEINA (ENGL. CETP)

KOD GOJAZNE DECE

Marijana Jevti}, Tanja GligorovMentori: doc. dr Aleksandra Zeljkovi},

dipl. farm-med. biohem. Tamara Gojkovi}, dipl. farm-med. biohem. Jelena Joksi}

Katedra za medicinsku biohemiju, Farmaceutski fakultet Univerziteta u Beogradu

Gojaznost predstavlja jedan od osnovnih zdrav -stvenih problema u savremenom svetu, a sve ~e{}ese javlja i u pedijatrijskoj populaciji. Poreme}aji kon-centracije serumskih lipida su u velikoj meri uslovljenipromenjenim aktivnostima enzima lecitin-holesterolaciltransferaze (engl. LCAT) i holesterol-estar trans-fernog proteina (engl. CETP). Ispitati lipidni profil,aktivnost enzima LCAT i CETP kod gojazne dece iadolescenata i uporediti sa kontrolnom grupom nor-malno uhranjene dece. U ispitivanju je u~estvovalo90 gojazne dece i 27 normalno uhranjene dece, kojisu ambulantno le~eni na Univerzitetskoj de~joj kliniciTir{ova. Lipidni profil je odre|en rutinskim meto-dama. Aktivnosti LCAT i CETP su odre|ene meto -dom po Asztalos-u i saradnicima. Gojazna deca imalasu zna~ajno vi{i indeks telesne mase i koncentracijutriglicerida (P<0,001), a ni`u koncentraciju holes-terola sadr`anog u lipoproteinima niske gustine(engl. High-density lipoprotein, HDL) (R<0,001) uodnosu na negojaznu decu. Nije na|ena statisti~kizna~ajna razlika u aktivnostima enzima LCAT i CETPkod gojazne i normalno uhranjene dece. Kod go -jazne dece aktivnost LCAT je bila u statisti~ki zna -~ajnoj korelaciji sa koncentracijom ukupnog holes-terola (P<0,05) i aktivno{}u CETP (R<0,01), dok jeaktivnost CETP bila u zna~ajnoj korelaciji sa glikozili-ranim hemoglobinom (engl. HbA1C) (P<0,05).Aktivnost enzima CETP bila je zna~ajno povi{ena kodgojazne dece sa hipertenzijom u odnosu na nor-motenzivne ispitanike (P<0,05). Aktivnosti LCAT iCETP nisu se zna~ajno razlikovale izme|u gojazne inormalno uhranjene dece, ali su gojazna deca sahipertenzijom imala zna~ajno vi{e aktivnosti CETP.

Klju~ne re~i: gojazna deca, lipidni profil, LCAT,CETP.

SP010 ASSESSMENT OF THE ACTIVITY

OF LECITHIN-CHOLESTEROL ACYLTRANSFERASE AND

CHOLESTEROL ESTER TRANSFER PROTEIN IN OBESE CHILDREN

Marijana Jevti}, Tanja GligorovMentors: doc. dr Aleksandra Zeljkovi}, BSc.

Tamara Gojkovi}, BSc. Jelena Joksi}

Department of Medical Biochemistry, Faculty of Pharmacy, University of Belgrade

Obesity is one of the major health problems inthe modern world and it has been more and morefrequent in pediatric population. Disturbances ofserum lipid concentrations in obese individuals arehighly related to changes of activities of lecithin:cho-lesterol acyltransferase (LCAT) and cholesterol estertransfer protein (CETP). To examine lipid profile,activities of LCAT and CETP in obese children andadolescents and compare the results with a controlgroup of normal weight children. The study included90 obese children and 27 normal weight childrenwho are outpatients treated at the UniversityChildren’s Hospital Tir{ova. Lipid profile is deter-mined by routine methods. LCAT and CETP activitieswere determined by the method introduced byAsztalos and associates. Compared to normal weightchildren, obese children had significantly higherbody-mass index and concentrations of triglycerides(P<0.001), while lower concentration of high-densi-ty lipoprotein (HDL) cholesterol (P<0.001). Wefound no differences in activities of LCAT and CETPbetween obese and normal-weight children. In obesechildren, LCAT activity significantly correlated withconcentrations of total cholesterol (P<0.05) and theactivity of CETP (P<0.01). The activity of CETP andglycosylated hemoglobin (HbA1c) were in statistical-ly significant correlation (P<0.05 ). Activity of CETPwas significantly elevated in obese children withhypertension compared to normotensive subjects(P<0.05). LCAT and CETP activities were not signif-icantly different between obese and normal weightchildren, but obese children with hypertension hadsignificantly higher CETP activity.

Keywords: obese children, lipid profile, LCAT,CETP.

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SP011 ATEROPROTEKTIVNA ULOGA ENZIMA PARAOKSONAZE 1 U HRONI^NOJ BUBRE@NOJ

INSUFICIJENCIJI

Predrag Stojakovi}, Vuka{in \uki}Mentori: doc. dr Aleksandra Stefanovi}, asist.

dipl. farm. med.biohem. Milica Miljkovi}

Katedra za medicinsku biohemiju, Farmaceutski fakultet Univerziteta u Beogradu

Hroni~na bubre`na insuficijencija (HBI) pred-stavlja hroni~no, ireverzibilno gubljenje bubre`nefunkcije koje je blisko povezano sa ubrzanim razvo-jem ateroskleroze. Rezultati prethodnih istra`ivanjanedvosmisleno ukazuju na postojanje veze izme|uoksidativnog stresa i HBI. Parametri oksidativnogstresa i antioksidativne za{tite se sve vi{e prou~avajukao potencijalni novi markeri za procenu toka i isho-da ove bolesti. Cilj ovog istra`ivanja bio je da se ispi-ta ateroprotektivna uloga antioksidativnog enzimaparaoksonaze 1 (PON 1) kod pacijenata sa HBI. Uovoj studiji je u~estvovalo 20 predijaliznih, 20 dija -liznih pacijenata i 20 zdravih ispitanika. Kod svih ispi-tanika u serumu su odre|eni parametri lipidnog pro-fila, totalni oksidativni status (TOS) i status enzimaPON 1. Iz seruma ispitanika izdvojena je subfrakcijaseruma sa holesterolom sa lipoproteinskih ~esticavisoke gustine (HDL-C) u kojima je kvantifikovanakoli~ina paraoksonaze-1 i paraoksonaze-3 imuno-hemijskom metodom (ELISA test). Koncentracijetriglicerida bile su statisti~ki zna~ajno ve}e kod paci-jenata u odnosu na kontrolnu grupu (p<0,05), dokje koncentracija holesterola u ~esticama visoke gus-tine (HDL-C) bila zna~ajno ni`a (p<0,05). TOS jezna~ajno povi{en (p<0,05), dok je aktivnost enzimaPON 1 bila zna~ajno ni`a kod pacijenata. Ispitiva -njem koncentracije PON 1 u HDL subfrakcijamauo~eno je da je koncentracija zna~ajno manja kodpacijenata u odnosu na kontrolnu grupu, dok je kon-centracija PON 3 bila pribli`no ista u obe ispitivanegrupe. Na osnovu rezultata ove studije mo`e sezaklju~iti da se pacijenti sa HBI nalaze u stanjuuznapredovalog oksidativnog stresa, sni`ene koncen-tracije PON 1 i PON 3, kao i aktivnosti enzimaparaoksonaze 1, ukazuju na promene u strukturiHDL-a i rezultuju}u sni`enu antioksidativnu za{titu.

Klju~ne re~i: PON-1, PON-3, HBI, Oksidativnistres

SP011 THE ATHEROPROTECTIVE ROLE OF PARAOXONASE 1 ENZYME IN CHRONIC RENAL FAILURE

Predrag Stojakovi}, Vuka{in \uki}Mentors: Assoc. professor Aleksandra

Stefanovi}, dipl. pharm. med. biochem. Milica Miljkovi}

Department of Medical Biochemistry, Faculty of Pharmacy, University of Belgrade

The chronic renal failure (CRF) is chronic, irre-versible lost of kidney function associated with pro-gressive development of atherosclerosis. Results ofprevious studies have unambiguously showed therelation between oxidative stress and CRF. The oxida-tive stress and antioxidant parameters are increasing-ly being investigated as potential new markers for theassessment of progress and outcome of this disease.The aim of this study was to investigate the athero-protective role of paraoxonase-1 (PON-1), an antiox-idative enzyme in patients with CRF. This studyincluded 20 predialysis patients, 20 patients on dial-ysis and 20 healthy participants. In the serum of allparticipants the lipid profile parameters wereassessed, total oxidative status (TOS) and status ofthe PON-1 enzyme. The concentrations of paraox-onase-1 and paraoxonase-3 enzymes in the isolatedsubfractions of high density lipoprotein cholesterol(HDL) were quantified by immunoassay method(ELISA). The concentrations triglycerides were statis-tically significantly higher in patients compared withcontrol group (p<0.05), while the concentrations ofhigh density lipoprotein cholesterol (HDL-C) weresignificantly lower (p<0.05). TOS was significantlyhigher (p<0.05), while the enzyme activity of PON-1was significantly lower in patients. Results of assessedPON-1 concentration on HDL subfractions showedthat the concentrations were significantly lower inpatients compared with control group, while thePON-3 concentrations were approximately the samein the both groups. Based on results of this study wecan conclude that patients with CRF are in state ofadvanced oxidative stress, with decreased concentra-tion of PON-1 and PON-3, decreased paraoxonase 1activity, which indicates the change in HDL structureand consequent compromised antioxidative protec-tion.

Keywords: PON-1, PON-3, CRF, Oxidativestress