Uvod: Ronjenje s komprimiranim plinovima je često praćeno stvaranjem plinskih mjehurića u krvi i hiperoksijom, što se smatra mogućim
uzrocima endotelne disfunkcije nakon ronjenja. Endotelna ...disfunkcija se povezuje s asimptomatskim kardiovaskularnim i neurološkim
promjenama nakon ronjenja s komprimiranim plinovima.
Cilj ove disertacije je istražiti fiziološke i biokemijske promjene vaskularne i endotelne funkcije, te ispitati ima li znakova narušenog
integriteta SŽS-a nakon ronjenja s komprimiranim plinskim smjesama. Pritom su testirane sljedeće hipoteze: asimptomatski zaroni s
komprimiranim zrakom će uzrokovati promjene biokemijskih parametara endotelne funkcije i oksidacijskog stresa; asimptomatski zaroni će
imati različiti utjecaj na vaskularnu funkciju s obzirom na vrstu korištene plinske smjese za disanje; asimptomatski zaroni će u prisutnosti
arterijalizacije plinskih mjehurića uzrokovati porast koncentracije humoralnih biljega oštećenja endotela i središnjeg živčanog sustava.
Metode i ispitanici: Istraživanje je obuhvatilo tri studije. U prvoj studiji na 15 ispitanika istražene su promjene sljedećih parametara nakon
ronjenja s komprimiranim zrakom: vWf-a, PAI-1-a, hsCRP-a kao pokazatelja endotelne funkcije, te adenozina, TBARS i FRAP kao
pokazatelja oksidacijskog statusa uz istovremeno praćenje stupnja stvaranja mjehurića. U drugoj studiji na 10 ispitanika uspoređivan je
utjecaj ronjenja sa zrakom i ronjenja s nitrox-om na vaskularnu funkciju ispitivanjem promjena pokazatelja arterijske elastičnosti: brzine
pulsnog vala i augmentacijskog indeksa; pokazatelja endotelne funkcije mjerenjem protokom posredovane dilatacije (FMD); te koncentracije
nitrita u plazmi. Usporedba je uključivala i stupanj stvaranja mjehurića nakon zarona. U trećoj studiji na 16 ispitanika istraženo je dovodi li
arterijalizacija mjehurića do promjene koncentracije endotelina-1, S100-B i NSE u krvi odnosno do pojave znakova narušenog integriteta
endotela i središnjeg živčanog sustava.
Rezultati: Ronjenje je uzrokovalo značajno stvaranje plinskih mjehurića u sve tri studije. Arterijalizacije su zabilježene u sve tri studije i bile
su praćene visokim stupnjem stvaranja mjehurića. Značajno veći stupanj stvaranja mjehurića i češće arterijalizacije uslijedili su nakon
ronjenja sa zrakom u odnosu na nitrox. Indikatori arterijske elastičnosti pokazali su oprečne promjene nakon ronjenja s obje plinske smjese.
Brzina pulsnog vala je porasla sugerirajući porast krutosti arterija, dok se augmentacijski indeks smanjio nakon ronjenja. Ronjenje s nitroxom
pokazuje veći učinak na protokom posredovanu dilataciju u odnosu na ronjenje sa zrakom upućujući na veći utjecaj hiperoksije na
endotelnu funkciju od plinskih mjehurića. Ronjenje sa zrakom u prvoj studiji nije polučilo promjenu oksidacijskog statusa izuzev blagog
porasta antioksidacijskog kapaciteta nakon ronjenja. Mjerenje endotelnih parametara nije otkrilo promjene izuzev PAI-1 u prvoj studiji koji
pokazuje pad aktivnosti nakon ronjenja. Promjenu humoralnih biljega oštećenja SŽS-a u vidu porasta nakon ronjenja otkriva samo S-100B,
ali bez dokaza o oštećenju integriteta SŽS-a budući se sličan porast očitovao i nakon kontrolnog zarona bez stvaranja plinskih mjehurića i
arterijalizacije.
Zaključak: Ovim istraživanjem je pokazano da asimptomatski zaroni sa zrakom ne uzrokuju promjene biokemijskih pokazatelja endotelnog
oštećenja unatoč prisutnosti plinskih mjehurića u krvi, te da je potaknuta antioksidacijska obrana mogući mehanizam zaštite od endotelne
disfunkcije. Značajniji negativni učinak nitrox-a na endotel u odnosu na zrak, unatoč manjem stvaranju plinskih mjehurića i njihovoj rjeđoj
arterijalizaciji, upućuje na endotelno oštećenje uzrokovano hiperoksijom i na potrebu za dodatnim istraživanjima mehanizama endotelnih
promjena.
Uvod: Ronjenje s komprimiranim plinovima je često praćeno stvaranjem plinskih mjehurića u krvi i hiperoksijom, što se smatra mogućim
uzrocima endotelne disfunkcije nakon ronjenja. Endotelna disfunkcija se povezuje s asimptomatskim kardiovaskularnim i neurološkim
promjenama nakon ronjenja s komprimiranim plinovima.
Cilj ove disertacije je istražiti fiziološke i biokemijske promjene vaskularne i endotelne funkcije, te ispitati ima li znakova narušenog
integriteta SŽS-a nakon ronjenja s komprimiranim plinskim smjesama. Pritom su testirane sljedeće hipoteze: asimptomatski zaroni s
komprimiranim zrakom će uzrokovati promjene biokemijskih parametara endotelne funkcije i oksidacijskog stresa; asimptomatski zaroni će
imati različiti utjecaj na vaskularnu funkciju s obzirom na vrstu korištene plinske smjese za disanje; asimptomatski zaroni će u prisutnosti
arterijalizacije plinskih mjehurića uzrokovati porast koncentracije humoralnih biljega oštećenja endotela i središnjeg živčanog sustava.
Metode i ispitanici: Istraživanje je obuhvatilo tri studije. U prvoj studiji na 15 ispitanika istražene su promjene sljedećih parametara nakon
ronjenja s komprimiranim zrakom: vWf-a, PAI-1-a, hsCRP-a kao pokazatelja endotelne funkcije, te adenozina, TBARS i FRAP kao
pokazatelja oksidacijskog statusa uz istovremeno praćenje stupnja stvaranja mjehurića. U drugoj studiji na 10 ispitanika uspoređivan je
utjecaj ronjenja sa zrakom i ronjenja s nitrox-om na vaskularnu funkciju ispitivanjem promjena pokazatelja arterijske elastičnosti: brzine
pulsnog vala i augmentacijskog indeksa; pokazatelja endotelne funkcije mjerenjem protokom posredovane dilatacije (FMD); te koncentracije
nitrita u plazmi. Usporedba je uključivala i stupanj stvaranja mjehurića nakon zarona. U trećoj studiji na 16 ispitanika istraženo je dovodi li
arterijalizacija mjehurića do promjene koncentracije endotelina-1, S100-B i NSE u krvi odnosno do pojave znakova narušenog integriteta
endotela i središnjeg živčanog sustava.
Rezultati: Ronjenje je uzrokovalo značajno stvaranje plinskih mjehurića u sve tri studije. Arterijalizacije su zabilježene u sve tri studije i bile
su praćene visokim stupnjem stvaranja mjehurića. Značajno veći stupanj stvaranja mjehurića i češće arterijalizacije uslijedili su nakon
ronjenja sa zrakom u odnosu na nitrox. Indikatori arterijske elastičnosti pokazali su oprečne promjene nakon ronjenja s obje plinske smjese.
Brzina pulsnog vala je porasla sugerirajući porast krutosti arterija, dok se augmentacijski indeks smanjio nakon ronjenja. Ronjenje s nitroxom
pokazuje veći učinak na protokom posredovanu dilataciju u odnosu na ronjenje sa zrakom upućujući na veći utjecaj hiperoksije na
endotelnu funkciju od plinskih mjehurića. Ronjenje sa zrakom u prvoj studiji nije polučilo promjenu oksidacijskog statusa izuzev blagog
porasta antioksidacijskog kapaciteta nakon ronjenja. Mjerenje endotelnih parametara nije otkrilo promjene izuzev PAI-1 u prvoj studiji koji
pokazuje pad aktivnosti nakon ronjenja. Promjenu humoralnih biljega oštećenja SŽS-a u vidu porasta nakon ronjenja otkriva samo S-100B,
ali bez dokaza o oštećenju integriteta SŽS-a budući se sličan porast očitovao i nakon kontrolnog zarona bez stvaranja plinskih mjehurića i
arterijalizacije.
Zaključak: Ovim istraživanjem je pokazano da asimptomatski zaroni sa zrakom ne uzrokuju promjene biokemijskih pokazatelja endotelnog
oštećenja unatoč prisutnosti plinskih mjehurića u krvi, te da je potaknuta antioksidacijska obrana mogući mehanizam zaštite od endotelne
disfunkcije. Značajniji negativni učinak nitrox-a na endotel u odnosu na zrak, unatoč manjem stvaranju plinskih mjehurića i njihovoj rjeđoj
arterijalizaciji, upućuje na endotelno oštećenje uzrokovano hiperoksijom i na potrebu za dodatnim istraživanjima mehanizama endotelnih
promjena.
Background: Diving with compressed gases (SCUBA diving) is often associated with occurence of gas bubbles in the blood and hyperoxia,
which are possible causes of endothelial dysfunction after diving. Endothelial dysfunction is associated with asymptomatic cardiovascular
and neurological changes caused by SCUBA diving.
The aim of this study is to explore the physiological and biochemical changes of vascular and endothelial function, as well as to examine the
signs of impaired CNS integrity after diving with compressed gas mixtures. The following hypotheses were tested: asymptomatic dives with
compressed air will change the biochemical parameters of endothelial function and oxidative stress after dive; asymptomatic dives will have
different effects on vascular function considering different types of breathing gases; and, finally, asymptomatic dives will cause increased
levels of humoral markers of endothelial damage and CNS integrity due to arterializations of gas bubbles.
Methods: Three studies were performed. The first study including 15 participants examined changes of vWf, PAI-1 and hsCRP which
served as indicators of endothelial function, as well as adenosine, TBARS and FRAP as indicators of oxidative state while the bubble grades
were also monitored after dives with compressed air. In the second study that included 10 divers, the effects of diving with air and nitrox on
vascular function were compared by examining the indicators of arterial elasticity: pulse wave velocity and augmentation index; indicators
of endothelial function through assessment of flow-mediated dilation (FMD) and nitrite concentrations in plasma; and assessment of bubble
production. Third study which included 16 subjects examined whether bubble arterializations lead to increase in blood levels of endothelin-
1, S-100B and NSE, which would suggest an impaired endothelium and CNS integrity.
Results: Diving resulted in significant production of bubbles in all three studies. Arterializations were recorded in all three studies and were
accompanied by high bubble grade. Diving with air in the first study did not change the oxidative status with the exception of a slight
increase in antioxidant capacity after diving. Assessment of endothelial parameters did not reveal any changes, except PAI-1, activity of
which declined after diving . Air dives resulted in sign
Oralni antikoagulans varfarin se već desetljećima koristi u terapiji i/ili prevenciji tromboembolijskih stanja različite
etiologije. Varfarin ima uzak terapijski raspon i pokazuje veliku ...interindividualnu varijabilnost u odgovoru na terapiju,
što može potencijalno rezultirati i po život opasnim komplikacijama uslijed prekomjernog antikoagulacijskog učinka i
krvarenja.
Cilj ovoga istraživanja bio je ispitati utjecaj polimorfizama VKORC1 -1639G>A, VKORC1 1173C>T, CYP2C9*2,
CYP2C9*3, CYP4F2*3 i GGCX 12970C>G te kliničkih i okolišnih čimbenika na stabilnu dozu održavanja i nuspojave
terapije varfarinom. U istraživanju su sudjelovala 204 bolesnika na terapiji varfarinom i 420 zdravih ispitanika. Rezultati
genotipizacije na zdravim ispitanicima pokazuju kako je učestalost polimorfizama VKORC1 1639G>A i VKORC1
1173>T u populaciji Hrvatske u skladu s učestalošću u europskim populacijama. Rezultati istraživanja na skupini
bolesnika na terapiji varfarinom ukazuju na statistički značajnu povezanost stabilne doze održavanja varfarina sa
sljedećim čimbenicima: genotipovima CYP2C9, VKORC1 i CYP4F2, zatim s fizičkom aktivnošću, tjelesnom
površinom, te uzimanjem lijekova koji utječu na metabolizam varfarina. Algoritam za predviđanje doze dobiven
višestrukom linearnom regresijom u stanju je objasniti 49,8 % varijabilnosti doze varfarina. Dokazana je statistički
značajna povezanost vremena potrebnog za postizanje stabilne doze varfarina s polimorfizmom CYP4F2*3 te
povezanost udjela vremena provedenog unutar terapijskog raspona i pojave nuspojava terapije varfarinom u obliku
prekomjerne antikoagulacije s polimorfizmima VKORC1 -1639G>A i/ili VKORC1 1173C>T. Isto tako, utvrđena je
statistički značajna povezanost VKORC1 -1639G>A i/ili VKORC1 1173C>T te CYP4F2*3 s nuspojavama terapije
varfarinom u obliku krvarenja.
Warfarin is the most commonly used oral anticoagulant worldwide. It is used for over a half of century for prevention
of thromboembolic disorders. Dosage of warfarin is difficult due to a very narrow therapeutic index (with lifethreatening
overdose complications) and wide interindividual variability.
Aim of this study was to investigate association of VKORC1 -1639G>A, VKORC1 1173C>T, CYP2C9*2, CYP2C9*3,
CYP4F2*3 and GGCX 12970C>G genetic polymorphisms, along with some clinical and environmental factors, on
warfarin maintenance dose and adverse effects of warfarin therapy. This study included 204 patients on warfarin therapy
and 420 healthy volunteers. Results of genotyping study on healthy population confirms that allele frequencies for
VKORC1 -1639G>A and VKORC1 1173C>T polymorphisms in Croatian population are in good agreement with other
populations of European ancestry. Results obtained on warfarin patient population indicate that warfarin maintenance
dose was significantly associated with following variables: CYP2C9, VKORC1 and CYP4F2 genotype, physical activity,
body surface area and concomitant usage of drugs that influence warfarin metabolism. Dosage prediction algorithm
obtained with multiple regression could explain 49,8 % of dose variability. Time to warfarin stable dose was significantly
associated with CYP4F2*3 genetic polymorphism. Time in therapeutic range was significantly associated with VKORC1
-1639G>A and VKORC1 1173C>T polymorphisms. VKORC1 -1639G>A and VKORC1 1173C>T polymorphisms were
also significantly associated with time to first overanticoagulation in warfarin therapy. VKORC1 -1639G>A, VKORC1
1173C>T and CYP4F2*3 polymorphisms were significantly associated with bleedings encountered during warfarin
therapy.
Stresni proteini (HSP) pripadaju skupini unutarstaničnih proteina koji su eksprimirani konstitutivno i
kao odgovor na fizikalni odnosno biološki stres. U staničnom odgovoru na stres HSP imaju ulogu ...stabiliziranja
proteina i peptida čime promoviraju preživljenje stanice. Hsp27 i Hsp70 inhibitori su različitih čimbenika
apoptoze, dok je glavna uloga Hsp90 osiguravanje aktivnost čimbenika uključenih u proliferaciju stanica.
Sposobnost stresnih proteina da zaustave proces programirane stanične smrti i potaknu proliferaciiju predstavlja
bitan aspekt njihove povezanosti s malignom proliferacijom.
Glavni cilj ovog istraživanja bio je procijeniti ulogu Hsp70 u zaštiti od programirane stanične smrti u
tumorskim stanicama. U tom smislu ispitana je uloga inducibilnog oblika Hsp70 u fiziološkim i stresnim
uvjetima. Hsp70, induciran stresom, pokazao je značajnu ulogu u zaštiti tumorskih stanica od stanične smrti
izazvane staurosporinom. U svrhu utišavanja ekspresije Hsp70 pri fiziološkim uvjetima korištena je tehnologija
koja počiva na RNA interferenciji, specifičnoj razgradnji ciljne mRNA pomoću kratke siRNA uklopljene u
nanočestice. Dizajnirane su kitozanske nanočestice za dostavu Hsp70 sljedno-specifične siRNA. U uvjetima in
vitro kitozanske nanočestice pokazale su nisku toksičnost, učinkovito uklapanje Hsp70 siRNA te učinkovito
utišavanje Hsp70. U staničnim linijama Jurkat i U251N utišavanje Hsp70 omogućeno pomoću Hsp70 siRNA
uklopljene u kitozanske nanočestice, prouzročilo je smanjenje vijabilnosti ovih tumorskih staničnih linija.
U radu je ispitana i antitumorska učinkovitost istovremene inhibicije Hsp90, pomoću celastrola, i
Hsp70 pomoću kitozanskih nanočestica s uklopljenom Hsp70 siRNA. Učinkovitost ovog pristup bila je značajna
u dvodimenzionalnim modelima tumorskih staničnih linija, dok su u trodimenzionalnim modelima bile potrebne
povećane koncentracije oba spoja/sustava kako bi se postiglo značajno smanjenje vijabilnosti tumorskih stanica.
Rad predstavlja temelje za daljnja istraživanja optimalnih nanosustava za dostavu sljedno-specifičnih
siRNA za HSP te moguća ispitivanja istovremene inhibicije Hsp70 i Hsp90 u tumorskim modelima in vivo.
Stress proteins (HSP) are intracellular proteins expressed constitutively but can also be induced by
various types of stress including environmental changes and non-stress conditions such as cell cycle, growth
factors, development and differentiation. Under physiological conditions these proteins function as molecular
chaperones that enable the function of different proteins. Hsp27 and Hsp70 appear to function at key regulatory
control points in apoptotic process, whereas the major role of Hsp90 is protection from degradation for the major
factors responsible for cell proliferation. The promotion of tumor cell survival is closely related to the ability of
Hsp to inhibit programmed cell death during malignant proliferation.
The purpose and the main goal of the proposed study was to explore Hsp70's role in protection against
programmed cell death in tumor cells. The role of stress induced Hsp70 and the role of Hsp70 within
physiological levels was explored. Stress induced Hsp70 was found to efficiently protect tumor cells against cell
death promoted with staurosporin. Hsp70 silencing was accomplished using the technology based on RNA
interference, specific degradation of mRNA after binding with siRNA delivered into the cells with nanoparticles.
Chitosan based nanoparticles for Hsp70 siRNA delivery were prepared. These showed low toxicity, efficient
siRNA entrapment and efficient Hsp70 silencing. In Jurkat and U251N cell lines Hsp70 silencing with Hsp70
siRNA delivered in chitosan nanoparticles significantly decreased cell viability.
The antitumor efficiency of simultaneous inhibition of Hsp90 with celastrol and Hsp70 with chitosan
nanoparticles entrapping Hsp70 siRNA was also explored. Significant reduction in cell viability was observed in
two-dimensional cell cultures, while for the induction of comparable effect in three-dimensional cell cultures,
U251N spheroids, higher concentration of both celastrol and Hsp70 siRNA were necessary.
The results of this work present important findings that could lead into the development of optimal
delivery systems for siRNA and possible in vivo research of Hsp70/Hsp90 inhibition approach.
Alzheimerova bolest (engl. Alzheimer's disease, AD) je najčešća demencija karakterizirana stvaranjem plakova i
neurofibrilarnih spletova. Vaskularna demencija (VAD) je druga najučestalija vrsta ...demencije i nastaje uslijed
ishemijskih, hipoperfuzijskih ili hemoragičnih moždanih lezija. Ciljevi ovog rada su: ispitati diferencijalno
dijagnostičko značenje određivanja serumske koncentracije neurozina (humanog kalikreina 6, KLK6), klasterina
(CLU) i adiponektina (ADPN), te upalnog biljega interleukina-6 (IL-6) u razlikovanju AD i VAD; procijeniti
značenje ispitivanih biljega u razlikovanju kognitivno zdravih ispitanika iste dobi od oboljelih od demencije i
onih ispitanika iste dobi koji imaju blagi kognitivni poremećaj u odnosu na one s dijagnozom AD i VAD;
procijeniti korelaciju ispitivanih biljega sa standardnim pokazateljima kognitivnog deficita.
Ispitivanjem je obuhvaćeno 70 bolesnika s AD i 67 bolesnika s VAD koji su u neprekidnom slijedu pristizali na
redovnu neurološku obradu u Kliniku za neurologiju KBC-a Sestre milosrdnice u Zagrebu. Skupina kontrolnih
ispitanika iste dobi podijeljena je na kognitivno zdrave (N = 50) i one s blagim kognitivnim poremećajem (N =
48). U okviru rutinske neurološke obrade provedeni su neuropsihološki testovi Mini Mental State Examination
(MMSE) i Montreal Cognitive Assessment (MoCA). Korištene su slikovne tehnike: kompjuterizirana
tomografija mozga (MSCT), Colour Doppler Flow Imaging (CDFI) i transkranijska Doppler sonografija.
Rutinski biokemijski testovi izrađeni su na automatskom biokemijskom analizatoru. Koncentracije biljega KLK6
i CLU su određene ELISA metodom, a koncentracija ADPN je određena imunoturbidimetrijom na automatskom
biokemijskom analizatoru. Koncentracija IL-6 je određena na imunokemijskom analizatoru. Ovisno o vrsti
razdiobe dobivenih rezultata, za testiranje razlika korišteni su statistički testovi Kruskal Wallis i ANOVA. Za
analizu korelacije koristio se Spearmanov, odnosno Pearsonov test. Statistička analiza provedena je pomoću
programa MedCalc.
Kod ispitivanih skupina je nađena razlika za one testove koji se rutinski koriste u neurološkoj obradi dementnih
bolesnika. Koncentracije ispitivanih biljega KLK6, CLU i ADPN u serumu nisu se razlikovale između skupina
(P = 0,137, P = 0,178 i P = 0,268). Koncentracije upalnog biljega IL-6 značajno su se razlikovale između
ispitivanih skupina (P = 0,014), s najvećim medijanom koncentracije u skupini bolesnika s VAD (4,1 pg/mL) i
najmanjim medijanom koncentracije u skupini s MCI (2,3 pg/mL).
Koncentracije ispitivanih biljega KLK6, CLU i ADPN nisu se značajno razlikovale između oboljelih od AD i
VAD. Također, ispitivani biljezi ne pokazuju zadovoljavajuću mogućnost razlikovanja kognitivno zdravih
ispitanika iste dobi od bolesnika s demencijom, kao što ne pokazuju diskriminacijski potencijal kada je u pitanju
razlikovanje skupine iste dobi s dijagnozom blagog kognitivnog poremećaja od skupine s dijagnozom AD,
odnosno VAD. Određivanje koncentracije upalnog biljega IL-6 se pokazalo korisnim u razlikovanju ispitivanih
skupina. Koncentracije ispitivanih biljega nisu korelirale sa rezultatima standardnih pokazatelja kognitivnog
oštećenja.
Alzheimer's disease (AD) is the most frequent dementia characterized by formation of plaques and neurofibrilary
tangles. Vascular dementia (VAD) is the second most frequent type of dementia, which is caused by ischemic,
hypoperfusive or hemorrhagic brain lesions. The aims of this study are: assessment of potential serum
biomarkers neurosin (human kallikrein 6, KLK6), clusterin (CLU), adiponectin (ADPN) and inflammatory
marker interleukin – 6 (IL-6) in differential diagnostics of AD and VAD; assessment of potential of KLK6,
CLU, ADPN and IL-6 to separate age-matched cognitively healthy individuals from those who are demented and
to separate those individuals with symptoms of mild cognitive impairment (MCI) from those with overt AD or
VAD; assessment of correlation of KLK6, CLU, ADPN and IL-6 with results of parameters regularly used for
determination of cognitive deficit.
70 patients with diagnosis of AD and 67 patients with VAD were included in study in consecutive order during
their routine neurological follow-up in University Department of Neurology in Medical School University
Hospital Sestre milosrdnice, Zagreb. Control group of age-matched individuals consisted of cognitively healthy
individuals (N = 50) and those with mild cognitive impairment (MCI) (N = 48). Neuropsychological tests Mini
Mental State Examination (MMSE) and Montreal Cognitive Assessment (MoCA) were done as part of routine
neurological examination. Neuroimaging techniques multi slice computed tomography (MSCT), Colour Doppler
Flow Imaging (CDFI) and transcranial Doppler sonography were used. Routine biochemistry tests were
determined on automated biochemistry analyzer. Concentrations of potential biomarkers were measured using
ELISA method for KLK6 and CLU, and concentration of ADPN was measured using immunoturbidimetry on
automated biochemistry analyzer. Concentration of IL-6 was determined using immunochemistry analyzer.
Depending on data distribution, statistic tests Kruskal Wallis and ANOVA were used for difference testing. For
correlation analysis Spearman and Pearson tests were performed. Statistical analysis was done with MedCalc
program.
Difference between tested groups was found for those tests which are routinely used for neurological follow-up
of demented patients. Concentrations of potential biomarkers KLK6, CLU and ADPN did not differ between
tested participant groups (P = 0,137, P = 0,178 and P = 0,268). Concentrations of IL-6 were significantly
different for tested groups (P = 0,014), with highest median of concentration in VAD group (4,1 pg/mL) and
lowest median of concentration in MCI group (2,3 pg/mL).
Concentrations of investigated biomarkers KLK6, CLU and ADPN did not differ significantly between AD and
VAD patient group. Also, capability of tested biomarkers to differentiate age-matched cognitively healthy
participants from patients with diagnosis of dementia was not sufficient, as well as their discriminating potential
for age-matched participants with MCI compared to AD and VAD patient groups. Measurement of concentration
of inflammatory marker IL-6 proved to be useful in discriminating between tested groups. Concentrations of
potential biomarkers did not correlate with results of standard indicators of cognitive deficiency.
U fiziološkom odgovoru na stres i kontroli stresnog odgovora uglavnom sudjeluju dvije komponente neuroendokrinog sustava, simpatičko-medularna (SAM) os, koja regulira prvi odgovor na stres i os ...hipotalamus-adenohipofiza-kora nadbubrežne žlijezde (HPAC), čija se aktivacija uočava 15 do 20 minuta nakon stresnog podražaja. Cilj ovog istraživanja bilo je ispitati bazalnu, fiziološku aktivnost SAM i HPAC osi te njihovu povezanost s nizom kovarijabli (spol, tip škole, turnus pohađanja nastave, ritam spavanja i budnosti te demografska, akademska i društvena obilježja) u zdravih učenika koji pohađaju završne razrede srednjih škola. Specifični ciljevi uključivali su ispitivanje koncentracije kortizola i aktivnosti alfa-amilaze kao salivarnih biljega odgovora na stres te istraživanje njihove povezanosti sa subjektivnim procjenama razine stresa i sa strategijama suočavanja sa stresom, kao i s polimorfizmima jednog nukleotida (SNP) gena za mineralokortikoidni i glukokortikoidni receptor (rs5522, rs6189, rs6190) te za vezni protein 5 za FK506 (rs1360780). U istraživanje su bila uključena 903 maturanta gimnazija i strukovnih škola iz četiri najveća grada u Hrvatskoj (Zagreb, Split, Rijeka i Osijek). Uzorci sline prikupljeni kod kuće, tijekom jednog radnog dana u tri vremenske točke (po buđenju, 30 do 45 minuta nakon buđenja i neposredno prije lijeganja), uzeti su za analizu koncentracija kortizola dok se aktivnost alfa-amilaze mjerila samo u jednom uzorku, neposredno po buđenju. Kako bi se ispitao cirkadijalni ritam lučenja kortizola analizirani su i indeksi lučenja kortizola (CAR, DCD, AUCG). Učenice su imale statistički značajno veće koncentracije kortizola u jutarnjim satima, dok se koncentracije kortizola neposredno prije lijeganja nisu razlikovale među spolovima. Učenice imaju različite obrasce priprema za izazove koji ih očekuju tijekom nadolazećeg dana u odnosu na učenike, imaju značajno viši CAR, izraženiji DCD, veći AUCG u odnosu na učenike koji su imali statistički značajno veću aktivnost alfa-amilaze. Turnus pohađanja nastave, koji svakako posljedično utječe na ritam spavanja i trajanje budnosti, pokazao se kao prediktor lučenja kortizola. Statistički značajne razlike većine varijabli kortizola potvrđene su između učenika koji pohađaju različite turnuse nastave (SCC30-45, CAR, DCD i AUCG). Učenici koji su se budili ranije i imali duže trajanje budnosti imali su veći CAR, niži DCD i veći AUCG. Nisu potvrđene statistički značajne razlike niti jedne ispitivane varijable u učenika koji pohađaju različite tipove škola (gimnazije vs. strukovne škole). Također, statistički značajne razlike između učenika i učenica bile su uočene i u subjektivnoj procjeni stresa, suočavanju te njihovoj povezanosti s koncentracijama i indeksima kortizola. Zabrinutost za budućnost su oba spola procijenila kao najstresniju domenu života, dok su učenice imale statistički značajno veću razinu ukupnog stresa u odnosu na učenike te su više koristile aktivno suočavanje sa stresom. U ovom su istraživanju pronađene statistički značajne razlike povezanosti alela G polimorfizma rs 5522 s višom koncentracijom kortizola po buđenju (P=0,036), nižim CAR-om (P=0,021), dok je povezanost s DCD-om bila granična (P=0,050). Može se pretpostaviti kako učenici s dokazanim alelom G imaju niži CAR te zbog toga pokazuju lošiju pripremljenost za stresne izazove u nadolazećem danu, unatoč nešto izraženijim vrijednostima DCD-a. Ovo istraživanje ukazuje na potrebu definiranja protokola prikupljanja uzoraka sline u nekliničkim uvjetima i metodoloških smjernica za populacijska istraživanja, ponajprije zbog postojanja niza kovarijabli koje utječu na mjerenje koncentracije kortizola i aktivnosti alfa-amilaze. Dobiveni rezultati doprinose saznanjima o spolnim razlikama u funkciji HPAC osi, koje bi se trebale primijeniti i tijekom provedbe preventivnih mjera u adolescenata te ranijeg otkrivanja skupina koje su posebno osjetljive na stresne poticaje.
Physiological response to stress and the control of the stress response is mediated by two major components of the neuroendocrinological systems, sympathetic nervous (SAM) system and hypothalamic-pituitary-adrenal (HPA) axis. In the stress response, the SAM axis is first activated. About fifteen to twenty minutes following the SAM activation, the HPA axis is activated. The aim of the study was to examine basal, physiologically activity of the SAM and HPAC axis activity and to determine associations of various covariates (gender, school type, school shift, sleep-wake rhythm, demographic, academic and life style characteristics) with altered daily salivary cortisol profiles in healthy students attending finishing grades of secondary schools. Specific aims of the study were to analyse concentration of cortisol and activity of alpha- amylase, as salivary markers of stress reaction, to determine association of salivary markers of stress with stress perception and coping, and to determine associations of salivary markers of stress with single nucleotide polymorphisms (SNP) in the genes for the mineralocorticoid and glucocorticoid receptors (rs5522, rs6189, rs6190) and FK506 binding protein 5 (rs1360780). The study included 903 secondary school students enrolled in the finishing classes of gymnasiums and vocational schools from the four largest cities in Croatia (Zagreb, Split, Rijeka and Osijek). The saliva samples were collected at students’ homes, over the course of one weekday. Salivary cortisol was sampled at three time points: at awakening, 30 to 45 after wakening and at bedtime. Salivary alpha-amylase was sampled at awakening. In order to analyse the circadian rhythm of salivary cortisol secretion, three indexes were analysed (CAR, DCD and AUCG).Females had higher morning concentrations of salivary cortisol than males, whereas bedtime cortisol concentrations were not different in females and males. Females had different salivary cortisol profile and different mechanisms for preparing for the anticipated stress of the upcoming day, higher CAR, steeper DCD and larger AUCG than males. On the other side, males had higher activity of salivary alpha-amylase. School shift, and therefore the sleep-wake up rhythm, was an important predictor of the secretion of salivary cortisol. There were statistically significant differences in two school shifts with respect to the majority of salivary cortisol measures (SCC30-45, CAR, DCD and AUCG). Students who woke-up earlier and were longer awake had larger CAR, flatter DCD and larger AUCG. Students from two types of schools (gymnasiums vs. vocational schools) did not differ in any of the measured salivary variables. Furthermore, female and male students differed significantly in stress perception, coping and associations between stress perception, coping and salivary cortisol concentrations and indexes. Future was rated as the most stressful problem domain in both sexes. The perception of total stress was higher in females than males, and females used more active coping than males. Finally, we found statistically significant differences in associations between G allele of rs 5522 polymorphism and higher cortisol concentration at awakening (P=0.036), lower CAR (P=0.021) and higher DCD (P=0.050), but at borderline. It can be hypothesized that the G allele is associated with the lower CAR and therefore, decreased ability of preparing for the anticipated stress (despite borderline association with the steeper DCD). This study points to the need for defining the protocol for salivary sampling in non-clinical conditions and developing methodological guidelines for population-based research, primarily due to the number of co-variables that affect the concentration of salivary cortisol and the activity of salivary alpha-amylase. Obtained results contribute to better understanding of sex differences in the functioning of the HPA axis, which should be considered in the prevention of various psychopathological conditions in adolescence and used for earlier identification of vulnerable groups at high risk of stress-related health problems.
Kronična opstrukcijska plućna bolest (KOPB) je složeni poremećaj koji zahvaća pluća, ali
i sistemski odjeljak. Naše se istraživanje temeljilo na hipotezi da je u bolesnika s KOPB-om
prisutna ...sistemska upala i sistemski oksidacijski stres. Sukladno tome, cilj ovog istraživanja
bio je istražiti sistemske pokazatelje oksidacijskih i upalnih promjena u bolesnika sa stabilnim
KOPB-om te ispitati povezanost sistemskih antioksidansa s poremećajem funkcije pluća i
biljezima sistemske upale. Ispitali smo i dijagnostičku učinkovitost biljega sistemskog
oksidacijskog stresa u razlikovanju zdravih i oboljelih od KOPB-a.
Sistemski biljezi oksidacijskog stresa (albumin, transferin, ceruloplazmin, ukupni bilirubin,
slobodne tiolne skupine, malondialdehid (MDA), paraoksonazna i arilesterazna aktivnost
paraoksonaze 1 (PON1), te ekspresija i aktivacija unutarstaničnih signalnih molekula Hsp27,
Hsp70, ERK, JNK i p38) određeni su u 106 bolesnika sa stabilnim KOPB-om i 45 zdravih
ispitanika. Ispitane su i njihove povezanosti s biljezima sistemske upale (CRP, fibrinogen,
ukupni leukociti), pušenjem i pokazateljima funkcije pluća (FEV1 i FEV1/FVC).
Bolesnici s KOPB-om su imali povišene koncentracije ceruloplazmina i MDA, snižene
koncentracije albumina, transferina i tiola, te snižene obje ispitane aktivnosti PON1.
Koncentracije ukupnog bilirubina nisu se razlikovale usporedbom ispitivanih skupina.
Ceruloplazmin je pokazao pozitivnu korelaciju s CRP-om i fibrinogenom. Albumin i
transferin su pokazali negativnu korelaciju s CRP-om, te pozitivnu korelaciju sa slobodnim
tiolima. Transferin je negativno korelirao s fibrinogenom. Jedini pokazatelj povezan s
funkcijom pluća bio je MDA. Usporedbom pušača, bivših pušača i nepušača iz skupine
bolesnika s KOPB-om nisu nađene razlike u koncentracijama promatranih biljega
oksidacijskog stresa i upale. Povišene koncentracije ceruloplazmina su najsnažniji prediktor
prisutnosti KOPB-a. Model koji uključuje ceruloplazmin, albumin, MDA i arilesteraznu
aktivnost PON1 te biljege sistemske upale pokazao je najbolju dijagnostičku učinkovitost u
predviđanju KOPB-a (AUC (95%CI) = 0,96 (0,92 – 0,99)). Predloženi bi model mogao
ispravno predvidjeti prisutnost KOPB-a u 89% bolesnika. Također, naši su rezultati ukazali da
je razina ekspresije Hsp27 i Hsp70 u leukocitima periferne bila je najniža u pušača s KOPBom.
Ekspresija svih ispitivanih MAPK (ERK, JNK i p38) u perifernim leukocitima bolesnika
s KOPB-om nije se razlikovala u usporedbi sa zdravim ispitanicima. Aktivacija ERK bila je
značajnija kod zdravih i bolesnih nepušača, dok je aktivacija JNK i p38 bila najizraženija u
pušača s KOPB-om.
Rezultati su ukazali da prisutnost bolesti i pušenje utječu na ispitane unutarstanične
signalne molekule. Bolje razumijevanje ovih molekularnih mehanizama moglo bi pomoći u
dijagnozi i pronalaženju novih terapijskih meta za KOPB. Dijagnostičke karakteristike
predloženog modela koji kombinira koncentracije biljega sistemske upale i sistemskog
oksidacijskog stresa ukazuju da bi se on mogao koristiti kao vrijedan alat u razlikovanju
zdravih ispitanika i bolesnika s KOPB-om.
Background: Chronic obstructive pulmonary disease (COPD) is a complex disorder
affecting the lungs and the systemic compartment. We hypothesized that systemic
inflammation and systemic oxidative stress are present in patients with COPD. Therefore, we
aimed to investigate markers of systemic oxidative and inflammatory alterations in patients
with stable COPD, and to test the association of systemic antioxidants with indicators of lung
function and systemic inflammation. The diagnostic accuracy of systemic oxidative stress
parameters in distinguishing between healthy subjects and patients with COPD was also
evaluated.
Materials and methods: Systemic oxidative stress markers (albumin, transferrin,
ceruloplasmin, total bilirubin, thiols, malondialdehyde (MDA), paraoxonase and arylesterase
activity of paraoxonase 1 (PON1), and the expression and activation of intracellular signalling
molecules Hsp27, Hsp70, ERK, JNK i p38) were assessed in 106 stable COPD patients and
45 healthy subjects. Their association with systemic inflammatory markers (CRP, fibrinogen,
total leukocytes), smoking status and lung function parameters (FEV1 i FEV1/FVC) was
investigated.
Results: Higher ceruloplasmin and MDA concentrations, and lower albumin, transferrin,
thiols and PON1 activities (paraoxonase and arylesterase) were found in patients with COPD.
Total bilirubin concentrations were similar in the studied groups. Ceruloplasmin showed a
positive correlation with CRP and fibrinogen. Albumin and transferrin showed a negative
correlation with CRP, and a positive corelation with thiols. Transferrin negatively correlated
with fibrinogen. Only MDA showed an association with pulmonary function. No differences
were found comparing concentrations of oxidative stress and inflammatory markers between
COPD patients subdivided according to their smoking status. Ceruloplasmin was the strongest
single predictor of COPD. The model combining ceruloplasmin, albumin, MDA, arylesterase
PON1 activity, and markers of systemic inflammation demonstrated very good diagnostic
performances (AUC (95%CI) = 0,96 (0,92 – 0,99)). The proposed model correctly identifies
89% of patients with COPD. In addition, our results showed that the decrease in expression of
peripheral blood leukocytes' Hsp27 and Hsp70 was the most prominent in COPD smokers.
Expression levels of all three MAPKs investigated was not altered in leukocytes of COPD
patients compared to healthy subjects. However, ERK activation was stimulated in healthy and COPD non-smokers, while JNK and p38 activation was the most pronounced in COPD
smokers.
Conclusions: Our results showed that COPD and smoking affect the intracellular
signalling pathways investigated. Improved understanding of these molecular mechanisms
could help identify novel targets for diagnosis and therapeutic interventions in COPD.
Diagnostic characteristics of the proposed model, obtained by combining markers of systemic
inflammation and systemic oxidative stress, suggest its potential value as an additional tool in
COPD diagnosis.
Kosti su stalno u procesu trošenja i obnavljanja, a ravnoteža razgradnje i izgradnje
preduvjet je za zdravo koštano tkivo. Slabljenje kvalitete kosti posljedica je poremećaja te
ravnoteže. Za ...razgradnju su odgovori osteoklasti dok su za izgradnju odgovorni osteoblasti.
Osteopenija je stanje smanjene gustoće kosti, a do nje može doći i zbog dugotrajne primjene
glukokortikosteroida.
Glukokortikosteroidi su protuupalni lijekovi koji se, između ostalog, koriste za
sprečavanje upale dišnih puteva kod astme. Budući je astma česta kronična bolest u djece
potrebno je pratiti metaboličke i koštane učinke glukokortikosteroida tijekom terapije. Cilj
ovog rada je uspostava modela hipoplazije fize izazvane glukokortikosteroidima i odabir
optimalnih biljega njihovih metaboličkih i koštanih učinaka.
Istražen je učinak 3 glukokortikosteroida na rast i promjenu metabolizma kosti te na
ostale sustavne učinke u štakora. Beklometazon dipropionat, prednizolon i ciklezonid, davani
mladim muškim Sprague-Dawley štakorima 7 dana u dozama od 0,3-10 mg/kg dnevno, s.c.,
su ovisno o dozi inhibirali indeks tjelesne mase timusa (za 57%, 44% i 76% s 3 mg/kg).
Ciklezonid i manje učinkovit prednizolon su utjecali na ploču rasta glave femura inhibirajući
rast femura (za 41% i 18% s 10 mg/kg), značajno smanjujući povećanje tjelesne mase (oboje
za 100% s 10 mg/kg), te serumske koncentracije kisele fosfataze i tartarat rezistentne kisele
fosfataze (za >30% s 10 mg/kg); oba su povećala serumske razine glukoze i triglicerida.
Beklometazon dipropionat je imao slab učinak na ove dodatne varijable. Ciklezonid pokazuje
izraženo inhibirajuće djelovanje na rast kosti u štakora. Možemo zaključiti da je ovo dobar
model za ispitivanje utjecaja glukokortikosteroida na metabolizam kosti.
Bone in children is structurally different from adult bone. It is weaker but less brittle.
Bone growth starts with cartilage formation. Then vessels invade the cartilage, delivering
pluripotent stem cells, which initiate the formation of a primary center of ossification.
Secondary ossification centers are formed at each end of long bone, and between the primary
and secondary ossification centers the growth plate, or physis, develops. Bone grows as
secondary and primary ossification centers unite.
Bone tissue is in dinamic process of constant deteriorating and regeneration.
Weakening of bone quality is a result of imbalance in a process of bone remodelling. Bone
remodelling has two stages: bone resorption with the osteoclasts, bone cells which resorb the
bone, and bone formation with osteoblasts, bone cells which form the bone. The bone
remodelling cycle ends with bone mineralisation. The content of mineral in bones is defined
as bone density. Osteopenia is a condition with decreased bone density. Apart being a sign of
normal aging, osteopenia can be induced with prolonged use of glucocorticosteroids.
Glucocorticosteroids are antiinflammatory medications prescribed, among others, for
reducing and prevention inflammation of respiratory pathways in asthma. Since asthma is the
most common chronic disease in children, need for monitoring metabolic and bone effects of
glucocorticosteroids during therapy is appearing. Although inhaled glucocorticoids are known
to have systemic effects on bone metabolism, there is little comparative information on their
relative potencies.
The goal of this work is establishment of glucocorticosteroid induced hypoplasia of
the physis and finding the optimal markers of their metabolic and bone effects.
The effects of three standard glucocorticoids, beclomethasone dipropionate,
prednisolone and ciclesonide, in causing changes in bone metabolism and growth were
investigated in relation to other systemic effects in the rat.
Male, specific pathogen-free, Sprague-Dawley rats, 4,5–5,5 weeks old (at the
beginning of the experiments), were used in the study.
The rat femur model of glucocorticosteroid-induced hypoplasia of the physis was
established according to Belvisi et al., using subcutaneous (s.c.) drug administration to allow
for future parenteral comparison with novel compounds. Briefly, rats were randomly assigned
to experimental groups of 8 animals each. In total, three experiments were performed for each
glucocorticoid; beclomethasone dipropionate, prednisolone and ciclesonide, at doses of 0,3–
10 mg/kg daily for 7 days. Animals in the control groups received s.c. the volume of 10 ml/kg
of vehicle (4% DMSO in 0,125% CMC) daily, for 7 days. Twenty-four hours after the last
treatment, animals were anaesthetized with sodium thiopental and the blood was collected at
exsanguination in order to obtain serum. Also, thymus weights were recorded and the femoral
bones removed (for measurement of the thickness of the proliferating zone). Animal body
weights were correspondingly documented at the beginning and at the end of each
experiment. Body weight gain was calculated as the change in body weight from day 1 until
24 h after treatment on day 7.
Biochemical analyses were performed on rat sera. Serum concentrations of glucose
and triglycerides, alkaline and acid phosphatases, and tartrate-resistant acid phosphatase were
determined on the biochemical analyzer. Concentration of osteocalcin, as a biochemical
marker for bone formation, and TRACP 5b, as a biochemical marker for bone resorption,
were also determined using ELISA.
The thymus was dissected free of connective tissue and immediately weighed.
Thymus body mass index (BMI) was calculated according to the following formula: BMI
(thymus)=thymus weight (mg)/body mass (mg). The left femur was exposed and removed
with the head intact in the acetabulum by cutting through the pelvic girdle and through the
femur shaft above the knee joint. The tissue was then fixed in 10% neutral buffered formalin
for histological assessment.
For the purpose of quantitative histology of the femoral head proliferating zone femurs
were fixed, decalcified and processed to paraffin using the unit for tissue processing. Threemicrometer-
thick sections were cut in a way to include femoral head and stained. The femoral
head growth plate was examined under a light microscope. Images of the growth plate were
captured onto a computer. One image was captured from each tissue section, five
measurements of the growth plate width being obtained from each calibrated image.
Measurements involved drawing a line perpendicular to the growth plate between the edge of
the hypertrophic zone, distal to the articular cartilage and the end of the proliferating zone.
Daily treatment for 7 days with standard glucocorticoids resulted in significant
increases in serum glucose and triglycerides concentrations at the highest doses (10 mg/kg) of
prednisolone and ciclesonide. The most pronounced changes were observed with ciclesonide,
which also significantly increased serum triglycerides at a daily dose of 3 mg/kg.
None of the standard glucocorticoids had any significant effect on serum ALP, over
the tested dose range, given daily for 7 days. However, at the highest dose (10 mg/kg),
prednisolone and ciclesonide significantly inhibited both serum ACP and TRACP. The most
pronounced changes were observed with ciclesonide, which also significantly decreased
serum ACP and TRACP at a daily dose of 3 mg/kg. Beclomethasone dipropionate was less
effective, causing a slight but significant decrease in serum ACP (but not TRACP) at doses of
0,3 and 1 mg/kg.
The comparative potency of the three glucocorticoids in influencing non-specific
serum parameters of bone metabolism was also reflected in their effects on the proliferating
zone thickness of the femoral bone head. While beclomethasone dipropionate had no
significant effect, ciclesonide and prednisolong decreased physeal growth plate width.
Prednisolone reduced median bone growth by 18% at the highest dose (10 mg/kg) and
ciclesonide caused a dosedependent reduction in median bone growth, with significant
inhibition of up to 41% over the whole tested dose range (0,3–10 mg/kg).
All three standard glucocorticoids exerted significant, dose related inhibitory effects
on median body weight gain and thymus body mass indices after daily treatment for 7 days.
Beclomethasone dipropionate was the least growth inhibitory; although it caused statistically
significant inhibition of thymus BMIs by 50% at dose of 1 mg/kg per day, it caused
statistically significant 23% inhibition of median body weight gain only at 10 mg/kg per day.
Prednisolone affected thymus BMIs causing statistically significant inhibition by 44% at dose
of 3 mg/kg per day and body weight gain causing statistically significant inhibition by 33% at
dose of 1 mg/kg per day. Ciclesonide exerted the most pronounced inhibition of body weight
gain and thymus BMIs, causing statistically significant inhibition by 34% and 55%,
respectively, already at the lowest dose of 0,3 mg/kg per day.
In order to see how examined glucocorticoids effected bone markers results showed
that prednisolone had no statistical effect on bone formation while ciclesonide significantly
reduced osteocalcin concentration in doses of 3 and 10 mg/kg per day (157 ng/mL and 88
ng/mL, respectively) vs. control (453 ng/mL).
On the other hand, ciclesonid showed no significant effect on bone resorption while
prednisolone significantly reduced TRACP 5b concentration at the highest dose of 10 mg/kg
per day compared to negative control group (10,6±0,9 U/L vs. 19,7±3,1 U/L).
Ciclesonide, although a pro-drug, still has potent systemic activity in the rat, causing
typical glucocorticoid effects, including inhibition of bone growth. Prednisolone exhibits a
similar, though less potent, spectrum of systemic activity, while beclomethasone dipropionate
has weak activity in causing systemic metabolic effects, but retains thymus inhibiting
potency. However, although the distinction between the effect of glucocorticoids on bone
growth was observed in this study, the model can provide the toxic effect dose titration as
well as
Kardiovaskularne bolesti uzrokovane aterosklerozom predstavljaju značajan uzrok smrtnosti u hemodijaliziranih
(HD) bolesnika. Bolesnici koji se liječe HD imaju i do 30 puta veću stopu smrtnosti ...uzrokovanu kardiovaskularnim
bolestima. Rizični čimbenici uključeni u patogenezu ateroskleroze u HD bolesnika pacijenata uključuju dijabetes,
hipertenziju, dislipidemiju, pušenje, oksidativni stres, upalu te endotelnu disfunkciju. Budući da ateroskleroza ima
dugačku subkliničku fazu veoma je važno da patološki procesi budu otkriveni čim ranije, dok je bolest još u
asimptomatskoj fazi. Povećana debljina intime-medije (IMT) prepoznata je kao rani indikator subkliničke
ateroskleroze, kako u općoj populaciji tako i u HD-ih bolesnika te je povezana s tradicionalnim i novijim
kardiovaskularnim čimbenicima rizika. Nekoliko je studija dokazalo da je karotidna ateroskleroza povezana s
adhezijskim molekulama te omentin-1 proteinom. Ciljevi ovog rada bili su sljedeći: a) istražiti povezanost sICAM-1,
sVCAM-1, omentin-1 proteina te ostalih netradicionalnih rizičnih čimbenika sa subkliničkom aterosklerozom; b)
ispitati dijagnostičku vrijednost ovih specifičnih markera u otkrivanju subkliničke ateroskleroze i c) ispitati njihovu
ulogu kao prediktora smrtnosti u asimptomatskih bolesnika.
Počevši od studenog 2011. godine, kohorta od 210 HD-ih bolesnika sudjelovala je u trogodišnjem istraživanju.
Ispitanici su bili podijeljeni u tri skupine ovisno o prisutnosti ateroskleroze. Ateroskleroza je dijagnosticirana na
temelju mjerenja debljine intima-medija karotidne arterije. Uzorkovanje krvi izvršeno je na početku studije te svakih
12 mjeseci iza toga sve do kraja istraživanja. Ehokardiografska mjerenja učinjena su na početku i na kraju studije.
Dokazana je slaba korelacije između IMT i sICAM-1 (r=0,39, P=0,001), sVCAM-1 (r=0,27, P=0,015) i omentin-1
proteina (r=-0,25, P=0,020). Omentin-1 protein je pokazao dobru korelaciju s parametrima sistoličke i dijastoličke
funkcije lijeve klijetke (r=0,52, P=0,001 i r=0,51, P=0,001). Multivarijantna analiza je pokazala kako su koncentracije
sICAM-1 i omentin-1 proteina snažni neovisni pretkazatelji IMT (P=0,031 i P=0,010). Coxova analiza
proporcionalnih rizika pokazala je da su koncentracija sICAM-1 i omentin-1 proteina snažni pretkazatelji
kardiovaskularne smrti (HR=1,85, CI=1,18-2,32, P=0,021 i HR=4,14, CI=1,38-12,1, P=0,004,) te da uzastopno
mjerenje koncentracija sICAM-1 i omentin-1 proteina predstavlja jake prediktore IMT progresije (HR=1,98, 95%
CI=1,21-2,38, P<0,002 i HR=2,91, 95% CI=1,57-4,72, P<0,001).
Naša studija je pokazala kako su koncentracije ICAM-1 I omentin-1 proteina jaki pretkazatelji kardiovaskularne
smrti u asimptomatskih HD bolesnika. Porast koncentracije ICAM-1 te pad koncentracije omentin-1 proteina imaju
ključnu ulogu u ranoj progresiji ateroskleroze.
Atherosclerotic cardiovascular complications represent significant cause of mortality in hemodialysis (HD)
patients. Patients undergoing HD have up to 30 times higher incidence of cardiovascular disease mortality. Risk
factors involved in pathogenesis of atherosclerosis in patients undergoing HD include diabetes, hypertension,
dyslipidemia, smoking, oxidative stress, inflammation and endothelial dysfunction. Since atherosclerosis has a long
subclinical phase it is important that pathogenic processes are identified while disease is still asymptomatic. Increased
carotid intima-media thickness (cIMT) is well recognized as an early indicator of subclinical atherosclerosis both in
the general population and in HD patients and it is associated with traditional and emerging CV risk factors. Several
studies have shown that carotid atherosclerosis is associated with circulating concentrations of soluble adhesion
molecules (CAMs) and omentin-1 protein. The aims of this study were to: a) investigate association of sICAM-1,
sVCAM-1, omentin-1 and other non-traditional risk factors with subclinical atherosclerosis; b) examine the diagnostic
value of these specific markers in the early detection of subclinical atherosclerosis and c) examine their role as
predictors of mortality in group of patients with subclinical atherosclerosis on regular HD.
Starting from November 2011, a cohort of 210 HD patients participated in this three-year follow-up study. The
subjects were divided into three groups according to the presence of atherosclerosis. Atherosclerotic disease was
assessed by measuring carotid intima-media thickness (IMT). Samplings were withdrawn at baseline and thereafter
every 12 months until the end of follow-up. Echocardiography was performed at baseline and at the end of follow-up.
IMT showed weak correlation with sICAM-1 (r=0.39, P=0.001), sVCAM-1 (r=0.27, P=0.015), and omentin-1 (r=-
0.25, P=0.020), also omentin-1 showed good correlation with parameters of systolic and diastolic function (r=0.52,
P=0.001 and r=0.51, P=0.001). Multivariate analysis showed that sICAM-1 and sVCAM-1 concentrations were a
strong independent correlate of IMT (P=0.031 and P=0.010, respectively). The Cox proportional analysis showed that
sICAM-1 and omentin-1 concentrations were strong predictors of cardiovascular death (HR=1.85, CI=1.18-2.32,
P=0.021 and HR=4.14, CI=1.38-12.1, P=0.004, respectively) and that serial measurements of these markers predict
IMT progression (HR=1.98, 95% CI=1.21-2.38, P<0.002 and HR=2.91, 95% CI=1.57-4.72, P<0.001, respectively).
Our study demonstrated that sICAM-1 and omentin-1 levels are strong predictors of cardiovascular death in HD
patients with subclinical atherosclerosis. . We believe that an increase in sICAM-1 and decrease in omentin-1 levels
could play an important role in the early progression of atherosclerosis.
Moždani udar (MU) u djece heterogeni je poremećaj s višestrukom etiologijom, koja je još uvijek nepoznata u
približno 30% slučajeva. Genetički čimbenici nisu do kraja karakterizirani, a raspodjela ...gena kandidata za
nastanak MU različita je u pojedinim populacijama. U istraživanje je bilo uključeno 100 djece starosne dobi do
18 godina s potvrđenim MU i 100 zdrave djece podudarne po dobi i spolu djeci s MU. Napravljena je
genotipizacija 14 polimorfizama u 12 gena kandidata koji kodiraju proteine uključene u sustav zgrušavanja i
fibrinolize (FV Leiden, FV HR2, FII G20210A, beta-fibrinogen -455G>A, FXIII-A Val34Leu, PAI-1 4G/5G),
ljudskih trombocitnih antigena (HPA-1, -2, -3 i -5), metabolizma homocisteina (MTHFR C677T, MTHFR
A1298C) i intermedijernih rizicnih cimbenika (ACE I/D i ApoE 2-4). Utvrđena je cešća pojavnost MU u
dječaka, koja je uočena i u svim ispitivanim podskupinama. Najveći rizik za nastanak MU zabilježen je u prvoj
godini života, kao i 2,7 puta veći broj arterijskih ishemijskih MU (AIMU) u odnosu na hemoragijski MU
(HMU). Rezultati genotipizacije potvrdili su prije opisanu povezanost FV Leiden s nastankom (pojavom) MU.
Dokazana je i povezanost 4 polimorfizma (FV Leiden, FXIII-A Val34Leu, HPA-3 i apoE _2-4) s AIMU i to
različitih polimorfizama u pojedinim podskupinama prema spolu i dobi pojave AIMU, kao i povezanost
kombinacija genotipova polimorfizama MTHFR C677T i MTHFR A1298 (CC/AC) i dvostrukog heterozigotnog
oblika (GA/AG) polimorfizama FV Leiden i FV HR2 s AIMU, te kombinacije genotipova GA/AA s
perinatalnim AIMU. Utvrđen je i umjereno povećan rizik za pojavu HMU u nosioca haplotipa HA2 (HPA-
1a2a3a-ACED). Ovo istraživanje ukazalo je na povezanost pojedinih dosad nedovoljno istraženih polimorfizama
kao i kombinacija polimorfizama s etiologijom MU u djece u Hrvatskoj, što upotpunjuje i proširuje dosadašnje
spoznaje o etiološkim rizičnim čimbenicima.
Although stroke in children is a relatively rare and heterogeneous disorder with a wide range of identified risk
factors, the etiology of stroke is still undetermined in up to 30% of children. Taking into consideration that
genetic risk factors are incompletely characterized at present and the frequency of genetic factors may vary
among different populations, a genotype analysis was perforrmed of 14 polymorphisms in 12 candidate genes
encoding proteins of the coagulation and fibrinolysis systems (FV Leiden, FV HR2, FII G20210A, beta-fibrinogen -
455G>A, FXIII-A Val34Leu, PAI-1 4G/5G), human platelet alloantigens (HPA-1, -2, -3 and -5), homocysteine
metabolism (MTHFR C677T, MTHFR A1298C) and intermediate risk factors (ACE I/D and apoE 2-4) was
performed. The subject group comprised 100 children with a confirmed diagnosis of stroke aged £18 years and
100 age- and sex-matched control subjects. Obtained results were also analyzed in gender-specific stroke group
and subgroups according to the type of stroke: arterial ischemic (AIS) and hemorrhagic stroke (HS), and
according to the time of stroke onset (perinatal and childhood AIS). The predominance of boys was found
among (in) children with stroke and in all tested subgroups. The greatest risk for stroke was identified in the first
year of life, and AIS was found 2.7 times more frequently than HS. This case-control study has confirmed the
association between FV Leiden and stroke that was also observed in numerous studies so far, but it has also
shown that other previously not reported polymorhisms (FXIII-A Val34Leu, HPA-3 and the combination of FV
Leiden and FV HR2 polymorphisms) can be related to AIS in Croatian population. Analysis performed in
gender-specific stroke subgroups revealed the association of different polymorphisms in boys (FXII-A
Val34Leu, HPA-3 and combination of MTHFR polymorphisms) and girls (apoE 4 allele). The strongest
association (with OR>10.0) was found between FV Leiden and perinatal AIS for both genders whereas the
lowest risk for perinatal AIS was observed in girls who were carriers of at least one HPA-3b allele. Obtained
results have at least partially elucidated the role (impact) of new evaluated polymorphisms in the etiology of
stroke in children and their impact according to gender, type of stroke and time of stroke onset.
Oralni kontraceptivi imaju brojne učinke na organizam žene poput poželjnog smanjenja pojavnosti zloćudnih
bolesti jajnika i endometrija, ali i nepoželjne učinke od kojih je najpoznatiji povišeni ...trombotički rizik. Činjenica
je da su oralni kontraceptivi jedan od rizika za razvoj trombo-embolijskih bolesti, s relativnim rizikom od 1,15
do 8,5 ovisno o formulaciji pripravka i obliku trombo-embolijske bolesti. Razvoj laboratorijske dijagnostike,
molekularnih ispitivanja i koagulacijskih pretraga, dao je novi zamah istraživanjima povezujući epidemiološke
podatke o trombotičkim rizicima poput pušenja, debljine, dobi, tjelovježbe i drugih s biološkom osnovom.
Istovremeno, razvoj sintetičnih estrogena i progesterona, njihovih doza i kombinacija, dodatan su poticaj
studijama učinka pojedine formulacije na koagulacijski odgovor.
Cilj istraživanja bio je utvrđivanje utjecaja oralnih kontraceptiva na čimbenike zgrušavanja i fibrinolize u
lokalnoj populaciji, ovisno o: količini i hormonalnom sastavu korištenog pripravka, postojanju nasljednih
trombofilija te razlici među ispitanicama u dobivenim vrijednostima aktivnosti čimbenika hemostaze u odnosu
na dodatne čimbenike rizika za trombo-embolijske bolesti.
U istraživanje su uključeni rezultati 195 žena koje su uzimale 6 ciklusa oralnu kontracepciju. Određivani su
čimbenici nasljedne trombofilije (FV Leiden, FIIG20210A i PAI-1 5G/4G) i parametri hemostaze (PV, APTV,
fibrinogen, aPCR, protein C i S, FVIII, antitrombin, plazminogen, inhibitor plazmina, PAI-1 i d-dimeri) mjereni
u 3 vremenske točke: osnovni - prije uzimanja oralne kontracepcije, nakon 3 i nakon 6 ciklusa.
Rezultati istraživanja pokazuju da su pod utjecajem oralnih kontraceptiva promijenjeni svi koagulacijski
parametri osim aktivnosti FVIII. Analizom rezultata formulacija pripravaka, najveće razlike nađene su u
kombinacijama s 20μg EE2 GSD vs DRSP za koagulacijske pretrage protein C i S, plazminogen i PAI-1. U
istraživanju je potvrđena jednaka učestalost trombofilnih mutacija među ispitanicama kao u općoj populaciji.
Polimorfizam PAI-1 5G/4G nije imao utjecaja na aktivnost PAI-1 kod mladih žena, niti je uzimanje oralnih
kontraceptiva imalo učinka na aktivnost PAI-1.
Istraživanje je potvrdilo utjecaj oralne kontracepcije na koagulacijske pretrage, ali i nastojanje sustava
hemostaze za stjecanjem nove točke ravnoteže. Gestageni također utječu na koagulacijski odgovor žena.
Patološke vrijednosti pretraga aktivnosti proteina S i određivanja koncentracije d-dimera mogu pomoći
kliničarima u odluci o nastavku ili prekidu oralne kontracepcije kod žena s drugim visokim rizikom i/ili
sumnjom na trombo-embolijski događaj.
Oral contraceptives have numerous effects on women’s body like favorable decrease of prevalence risk for
ovarian and endometrial cancer, but also undesirable increase of thrombotic events. The fact is that oral
contraceptives are one of the reasons for increased risk for venous thromboembolism, with relative risk is 1.15-
8.5, depending on the drug formulation and type of thromboembolic disease. The development of laboratory
diagnostics, molecular tests and coagulation tests, gave a new momentum to the researches linking
epidemiological data on thrombotic risks such as smoking, obesity, age, exercise, and others with a biological
basis. Simultaneously, development of synthetic estrogen and progesterones are an additional stimulus for new
studies of the effect of oral contraceptive formulations on the coagulation response.
The aim of the study was to determine the influence of oral contraceptives on factors of coagulation and
fibrinolysis in the local population, depending on: the amount and composition of used drug, the presence of
hereditary thrombophilia and the difference among study subjects in the obtained values of factors of coagulation
and fibrinolysis in relation to the presence of additional risk factors for thromboembolic diseases.
The study included the data results of 195 women, who took six cycles of oral contraceptives. Tests for
hereditary thrombophilia (FV Leiden, FIIG20210A and PAI-1 5G/4G) were performed, as well as tests of
hemostasis (PT, APTT, fibrinogen, APCR, protein C and S, FVIII, antithrombin, plasminogen, plasmin inhibitor,
PAI-1 and d-dimer) measured at three time points: basic - before taking oral contraceptives after three and after
six cycles of their use.
The results showed that oral contraceptives have changed all measured parameters except activity of FVIII.
Analyzing the results by formulation of drug, the greatest differences were found in combination with 20μg EE2
GSD vs. DRSP for coagulation tests protein C and S, plasminogen and PAI-1. Data confirmed the same
frequency of FV Leiden, FII G2021A i PAI-1 5G/4G among study subjects as in general population.
Polymorphism of PAI-1 5G/4G had no influence on PAI-1 activity, nor the use of oral contraceptives.
This study confirmed impact of oral contraceptives on coagulation tests and tendency of hemostasis to
reach new equilibrium. Gestagens also have impact on coagulation response. Pathological test results of protein
S activity and d-dimers may help clinicians in the decision on continuation or discontinuation of oral
contraceptives in women with other high-risk and / or suspected thromboembolic event.
