Education of Masters of Science in Medical Biochemistry at the Faculty of Pharmacy and Biochemistry, University of Zagreb: How did we fit in the Bologne process

Reforma visokog obrazovanja na sveučilištima u RH temelji se na prihvaćanju Bolonjske deklaracije, koja kao osnovnu zadaću postavlja stvaranje cjelovitog europskog prostora visokog obrazovanja i istraživanja. Ovaj cilj zahtijeva da sve zemlje potpisnice izrade nacionalni okvir komparabilnih i kompatibilnih kvalifikacija u vlastitim sustavima visokog obrazovanja. Nova koncepcija Studija medicinske biokemije posebnu važnost daje multidis-ciplinarnom karakteru struke da bi se osigurali temelji za kvalitetnu primjenu znanstvenih spoznaja u kliničkom okruženju. Program je usklađen s preporukama koje su prihvaćene u većini zemalja Europe za rad u medicinsko-biokemijskim laboratorijima, a predviđa jedan petogodišnji ciklus edukacije koji završava titulom magistra medicinske biokemije. Multidisciplinarnost se postiže tako da se kroz studij stječu: temeljna znanja iz prirodnih znanosti, biomedicinska znanja kao što su anatomija, fiziologija, patofiziologija, histologija i citologija, imunologija, mikrobiologija i parazitologija, te stručna medicinsko biokemijska znanja - klinička biokemija, hematologija s koagulacijom, transfu-ziologija, analitička toksikologija, molekularna dijagnostika, racionalna laboratorijska dijagnostika i sl., te znanja i vještine iz komunikacijskih disciplina, laboratorijskog menadžmenta, te informatizacije laboratorijskog sustava. U ukupnom petogodišnjem studiju ima 28 % temeljnih, 12% biomedicinskih, 45% stručnih i 15% izbornih predmeta. Program predviđa maksimalno povezivanje osnovnih i stručnih predmeta, stručna praksa se provodi već od prve godine. Predmeti su ocijenjeni prema ECTS sustavu tako da student može dio studija obaviti i na nekom drugom sveučilištu. Nastavni plan se izvodi u suradnji s kliničkim bazama Fakulteta, posebno se to odnosi na Stručnu praksu i Integrirani kolegij laboratorijske dijagnostike koji se u cijelosti organizira u kliničkim laboratorijima. Novi program studija omogućava diplomiranom stručnjaku stjecanje cjelovitog znanja iz medicinske biokemije te stjecanje kompetencija da organizira rad i primjeni suvremene tehnologije u laboratorijsku praksu, da kompetentno interpretira laboratorijske nalaze, čineći ga bitnim članom stručnog medicinskog tima koji zbrinjava bolesnika, ili kompetentnim znanstvenim radnikom. Suvremeno obrazovanje medicinskih biokemičara omogućuje uspješno sudjelovanje u razvoju struke koja se širi prema sveobuhvatnim laboratorijskim znanostima - medicinskoj biokemiji i laboratorijskoj medicini.The recent reform of the university education in Republic of Croatia is undertaken to attain the main objectives set up by Bologna Declaration on promoting global competitiveness of European Higher Education. The Declaration that was signed by 29 countries is the common commitment to reform the structures of respective higher education system in a convergent way and to produce the national framework of readable and comparable degrees. New concept for the university degree in Medical Biochemistry is particularly emphasising the multidisciplinary character of the profession in order to enable application of scientific knowledge in the medical context. Curriculum is harmonised with the recommendations that are accepted in the majority of European countries, and consists of an integral five years programme leading to the degree of Master in Medical Biochemistry. The multidisciplinary approach is achieved by introducing various disciplines into curriculum: 1. Fundamental natural sciences; 2. Biomedical disciplines (anatomy, physiology, pathophysiology, histology, cytology, immunology, microbiology and parasitology, etc.); 3. Professional medical biochemistry disciplines (clinical biochemistry, haematology, coagulation, clinical immunology, blood banking, analytical toxicology, molecular diagnostics, etc.; and knowledge, competences and skills in communication, laboratory management, automation, and informatization of laboratory system. The relative proportion of the subjects during the five year study is: 28% fundamental natural sciences, 12% of biomedical, 45% professional and 15% elective subjects. The fundamental and professional subjects would be intensively correlated, and training in the hospital laboratory will start already from the first year. The ECTS credits will be assigned to the each subject, and student mobility would be encouraged. There will be great cooperation with the teaching hospitals related to Faculty, in particular for the courses Integral laboratory diagnostics and Professional laboratory practice. At completion of the new Medical Biochemistry curriculum the graduate would have a through knowledge of all aspects of medical biochemistry science relevant to the discipline, and competences to organize laboratory practice and apply current techniques, to evaluate the diagnostic data and to provide an expert opinion in the medical team, or to pursue a career in the fundamental and applied scientific research. The new concept comprises relevant knowledge in clinical cytology, microbiology, clinical immunology, blood banking, analytical toxicology, molecular diagnostics thus concurring with the current trends in Medical Biochemistry and Laboratory Medicine

Utjecaj različitih koncentracija peroksovanadijevog spoja bpV(phen) na preživljavanje PC12 stanica

Peroxovanadium compounds are potent insulinomimetic agents and protein tyrosine phosphatase inhibitors. In this study, the potential toxicity of peroxovanadium complex bpV(phen) on rat pheochromocytoma PC12 cells was examined, and the mechanism by which this compound influences cell survival and/or death was explored. BpV(phen) exerted a bimodal effect on PC12 cells: survival enhancing effect at lower and death-inducing effect at higher micromolar concentrations. 1 and 3 μmol dm–3 bpV(phen) intensely induced ERK activation. In contrast, 10 and 100 μmol dm–3 bp (phen) stimulated strong and sustained JNK and p38 MAPK activation as well as caspase-3 activation that preceded bpV(phen)-induced apoptotic cell death. It is suggested that bpV(phen) might exert its action on PC12 cell survival by modulation of MAPKs and caspase-3 activation.Spojevi peroksovanadija snažni su inzulinomimetički agensi i inhibitori tirozinskih fosfataza. U ovom istraživanju ispitivana je moguća toksičnost peroksovanadijevoga kompleksa bpV(phen) na PC12 stanicama feokromocitoma štakora te način na koji ovaj spoj utječe na preživljavanje i/ili umiranje stanica. BpV(phen) dvojako je djelovao na PC12 stanice: primijenjen u nižim mikromolarnim koncentracijama poticao je preživljavanje stanica, dok su više mikromolarne koncentracije poticale umiranje stanica. 1 i 3 μmol dm–3 bpV(phen) snažno je inducirao aktivaciju ERK kinaza. Suprotno tome, 10 i 100 μmol dm–3 bpV(phen) stimulirao je snažnu i dugotrajnu aktivaciju JNK i p38 MAPK kinaza te aktivaciju kaspaze-3, što je prethodilo umiranju stanica procesom apoptoze. Pretpostavlja se da bpV(phen) djeluje na preživljavanje PC12 stanica na način da modulira aktivaciju MAPK kinaza i kaspaze-3

TNF-alpha, CXCL8, big ET-1 and hsCRP in Patients with Chronic Obstructive Pulmonary Disease

COPD (chronic obstructive pulmonary disease) is a chronic progressive inflammatory disease characterised by limitations in lung airflow that is not fully reversible. The concentrations and correlations of TNF-á, CXCL8, big ET-1 and hsCRP were investigated in healthy non-smokers, healthy smokers and patients with COPD in order to study their possible role in the pathophysiology of COPD. The concentrations of TNF-á, CXCL8 and big ET-1 were not statistically different between the experimental groups. No significant differences for the measured analytes were found between smokers and the non-smokers in the control group. The Spearman coefficient of correlation between the concentrations of TNF-á and CXCL8 was r = 0.638 (p<0.0001). However, the concentration of hsCRP was significantly higher in patients with COPD than in the control group (p = 0.0004). hsCRP proved to be a more sensitive diagnostic parameter than TNF-á, CXCL8 and big ET-1 in the systemic circulation in patients with COPD

Hydrolysis of Extracellular Adenine Nucleotides by Human Spermatozoa: Regulatory Role of Ectonucleotidases in Sperm Function

Evidence is presented for the existence of ectonucleotidases on the membrane of intact human spermatozoa. Enzymes hydrolyze extracellular ATP, ADP and AMP and hence could be described as ecto-NTPDase and ecto-5\u27-nucleotidase. Suramin, Cibacron 3GA and DIDS, well known ecto-NTPDase inhibitors, caused inhibition of the observed enzyme activity. Enzymatic hydrolysis of ATP, ADP and AMP follows simple Michaelis-Menten kinetics with similar enzyme affinity for all three substrates (Km for ATP, ADP and AMP were (0.395 ± 0.027), (0.401 ± 0.031), (0.517 ± 0.038) mmole dm−3, respectively). Influence of extracellular ATP, AMP, adenosine and cAMP on the parameters of sperm velocity and acrosome reaction was also examined. In normozoospermic samples, ATP and cAMP induced an increase in the amplitude of lateral head displacement and number of acrosomally reacted cells, but not in sperm velocity. However, adenosine and AMP enhanced sperm velocity, without influencing the acrosome reaction. These results show that ATP and adenosine regulate sperm motility parameters in different ways

Okratoksin A izaziva apoptozu u LLC-PK1 stanicama aktivirajući JNK i p38 MAPK

Ochratoxin A (OTA) is a potential inducer of a tubular-interstitial nephropathy in humans and animals. In our study we addressed the question of involvement of apoptosis in the development of OTA-provoked nephrotoxicity. LLC-PK1 kidney cells were treated with nanomolar and micromolar concentrations of OTA for different lengths of time. The apoptotic process was estimated by morphological (haematoxylin/eosin staining, fluorescent staining of DNA free ends – TUNEL assay) and biochemical (MAPKs and Hsps) changes of cells. Forty-eight hours of treatment with 5 10–6 M OTA significantly decreased cell viability and induced apoptosis in 30.7 % of cells. In addition, a transient activation of ERK was observed as well as a strong and prolonged activation of stress kinases, JNK and p38 MAPK, after 12 and 48 hours of treatment. Expression of Hsp72 and Hsp27 was not affected by OTA. The results suggest that apoptosis mediated by activation of JNK and p38 MAPK might play an important role in OTA-induced nephrotoxicity in LLC-PK1 cells.Okratoksin A (OTA) može izazvati tubularno-intersticijsku nefropatiju u ljudi i životinja. Cilj našeg istra- živanja bio je ispitati ulogu apoptoze u nastanku nefrotoksičnosti koju uzrokuje OTA. Naš eksperimentalni model bile su LLC-PK1 bubrežne stanice izložene djelovanju OTA, od nanomolarnih do mikromolarnih koncentracija kroz različita vremenska razdoblja. Apoptoza je u stanicama utvrđena na temelju morfoloških (bojanje stanica hematoksilin/eozinom, fluorescentno bojanje slobodnih krajeva DNA tzv. TUNEL) i biokemijskih (MAP kinaze, Hsps) promjena. 48-satno izlaganje stanica 5 x 10–6 M OTA značajno je smanjilo staničnu vijabilnost i potaknulo apoptozu u 30,7 % stanica. Osim toga, utvrđena je kratkotrajna aktivacija ERK te snažna i dugotrajna aktivacija JNK i p38 MAPK nakon 6-, 12- i 48-satnoga tretiranja. OTA nije utjecao na razinu Hsp72 i Hsp27. Rezultati istraživanja ukazuju da je apoptoza posredovana aktivacijom JNK i p38 MAPK važan događaj u razvoju nefrotoksičnosti izazvanoj djelovanjem OTA na LLC-PK1 stanice

Reduction in Peripheral Blood Leukocyte Heat Shock Proteins 27 and 70 Expression in Chronic Obstructive Pulmonary Disease

Chronic obstructive pulmonary disease (COPD) is characterized by chronic local and systemic inflammation, and increased oxidative stress. Changes in peripheral blood leukocyte counts or in the fractions of leukocyte subsets might be implicated in the development of this disease. In this study, increased monocyte fraction was observed in COPD non-smokers. In light COPD smokers, the fraction of neutrophils was significantly increased along with significantly decreased lymphocyte fraction and lung function parameters. Heat shock proteins (Hsps) could stimulate antioxidant defence of cells by decreasing intracellular levels of reactive oxygen particles (ROS) and by neutralizing toxic effects of oxidized proteins. This study showed that the expression of Hsps in leukocytes was influenced by health and smoking status. Hsp70 and Hsp27 were significantly decreased in COPD ex-smokers and healthy smokers, but the most striking suppression in Hsps expression was detected in COPD smokers. It is suggested that this decline in Hsps intracellular levels could be implicated in the pathogenesis of COPD

Genetic Frequencies of Paraoxonase 1 Gene Polymorphisms in Croatian Population

Paraoxonase 1 (PON1), a HDL-associated enzyme, is believed to contribute to the protective effects of HDL by decreasing the generation of lipid peroxidation products during the process of LDL oxidation. It has been reported that polymorphisms in promoter and coding regions of the pon1 gene could affect PON1 activity. The aim of this study was to determine the frequencies of pon1 polymorphisms Q192R, L55M and –108C>T and their influence on PON1 activity in healthy population of Croatia. The following genotype frequencies were determined: 60 % QQ, 34 % QR, 6 % RR for Q192R; 44 % LL, 43 % LM, 13 % MM for L55M; 30 % CC, 48 % CT, 22 % TT for –108C>T polymorphism. Genotypes RR, LL and CC were found to be associated with higher PON1 activity. The most frequent genotypes in healthy Croatian population were QQ, CT and equally present LL and LM

Okratoksin A inducira apoptotičko i nekrotičko umiranje bubrežnih stanica

MDCK, LLC-PK1 and RK 13 renal cells were exposed to different OTA concentrations (2.5 to 25 μg OTA/mL of medium) for 24 hours. 2.5 μg OTA/mL significantly reduced the number of living RK 13 cells to 69.4 %, LLC-PK1 cells to 34.5 % and MDCK cells to 27.5 %, as compared to untreated control cells. Effects of OTA on cell morphology were examined by haematoxylin/ eosin or phalloidin-FITC/propidium iodide staining. 12 %, 11 % and 8.2 % apoptotic cells were observed in MDCK, LLC-PK1 and RK 13 cells treated with 2.5 μg OTA/mL, respectively. The structure of actin cytoskeleton showed significant changes in OTA-treated cells. Release of LDH into the culture medium was more abundant in MDCK and LLC-PK1 cells. OTA induced apoptotic and/or necrotic cell death in a cell type- and concentration-dependent manner. The results suggest that MDCK and LLC-PK1 cells are more sensitive to OTA than RK 13 cells.MDCK, LLC-PK1 i RK 13 bubrežne stanice tretirane su različitim koncentracijama OTA (od 2,5 do 25 μg OTA/mL medija) tijekom 24 sata. Koncentracije toksina od 2,5 μg/mL uzrokuju značajno smanjenje broja živih stanica u odnosu na netretirane kontrolne uzorke: kod RK 13 stanica broj je smanjen na 69,4 %, kod LLC-PK1 na 34,5 %, a kod MDCK na 27,5 %. Ispitan je učinak OTA na morfologiju stanica koje su bojane hematoksilinom/ eozinom odnosno faloidin-FITC/propidijevim jodidom. Kod MDCK, LLC-PK1 i RK 13 stanica tretiranih s 2,5 μg OTA/mL opaženo je 12 %, 11 % odnosno 8,2 % stanica u apoptozi. OTA je uzrokovao značajne promjene u strukturi aktinskoga citoskeleta u tretiranim stanicama. OTA je potaknuo istjecanje LDH iz stanica u okolni medij, što je osobito izraženo u MDCK i LLC-PK1 stanicama. Možemo zaključiti da način umiranja stanica (apoptoza ili nekroza) ovisi o vrsti samih stanica i o primijenjenoj koncentraciji toksina. Naši rezultati ukazuju da su MDCK i LLC-PK1 stanice osjetljivije na djelovanje OTA od RK 13 stanica