| Abstract | Uvod i ciljevi istraživanja: Masna promjena jetre izazvana lijekovima (DIFLD) postala je čest uzrok nealkoholne masne promjene jetre (NAFLD). zbog starenja stanovništva i raširenosti polifarmacije u starijih osoba. Moguće je da NAFLD i DIFLD koegzistiraju, što čini njihovo razlikovanje još važnijim. Naš je cilj bio istražiti modele NAFLD i DIFLD steatoze hepatocita u staničnoj kulturi i utvrditi učinke liraglutida u tim modelima. Materijali i metode: Za izradu modela steatoze hepatocita NAFLD i DIFLD, Huh7 stanice su tretirane oleinskom kiselinom, amiodaronom ili tamoksifenom. Liraglutid je dodan kao kotretman u stanice, a MTT je korišten za mjerenje viabilnosti stanica, dok je test isključenja eritrozin B bojom korišten za potvrdu rezultata za preživljavanje stanica. Oil-Red-O bojenje i test triglicerida korišteni su za mjerenje nakupljanja lipida, GSH kit korišten je za mjerenje koncentracija GSH u sva tri modela, a RT-PCR je korišten za mjerenje intracelularnih signala uključenih u hepatosteatozu. Rezultati: Koncentracije oleinske kiseline, amiodarona i tamoksifena koje su postigle 80% vijabilnosti stanica u usporedbi s negativnom kontrolom korištene su u narednim eksperimentima za uspostavljanje NAFLD i DIFLD modela. Te su koncentracije bile: 0,5 mM oleinske kiseline, 20 µM amiodarona, odnosno 2 µM tamoksifena. Liraglutid je poboljšao vijabilnost stanica u svim modelima (p < 0,01). Svi modeli pokazali su povećanu akumulaciju lipida, pri čemu je DIFLD pokazao mikrosteatotsku sliku, a NAFLD makrosteatotsku sliku, od kojih potonji odgovara većoj akumulaciji triglicerida. Liraglutid je smanjio ove steatotične promjene (p < 0,001). Razine GSH bile su smanjene u sva tri modela steatoze. Amiodaron i tamoksifen smanjili su ga na 80% u usporedbi s DMEM negativnom kontrolom (p < 0,01, p < 0,05), dok je učinak OA bio manji. Nije bilo značajne razlike u koncentracijama GSH s kotretmanom LIRA-om. U NAFLD i DIFLD modelu steatoze hepatocita, liraglutid je smanjio ekspresiju lipogenih puteva ACSL1, PPARG i SREBF1 (p < 0,01). Zaključak: U NAFLD i DIFLD modelima steatoze hepatocita uspostavljenim s Huh7 staničnom kulturom, liraglutid je smanjio steatozu hepatocita kroz regulaciju lipogenih puteva ACSL1, PPARG i SREBF1. |
| Abstract (english) | Background and aims: Drug-induced fatty liver disease (DIFLD) has become a common cause of non-alcoholic fatty liver disease (NAFLD) due to the aging of the population and the prevalence of polypharmacy in the elderly. It is possible for NAFLD and DIFLD to coexist which makes it even more crucial to be able to differentiate them. Our aim was to investigate hepatocyte steatosis NAFLD and DIFLD cell culture models and ascertain the effects of liraglutide in those models. Materials and Methods: To create models of hepatocyte steatosis NAFLD and DIFLD, Huh7 cells were treated with oleic acid, amiodarone, or tamoxifen. Liraglutide was administered to the cells, and MTT was used to measure cell viability, while Erythrosin B color exclusion test was used to confirm results for cell survival. Oil-Red-O staining and a triglyceride assay were used to measure lipid accumulation, GSH assay kit was used for the measurement of GSH concentrations in all three models, and RT-PCR was used to measure intracellular signals involved in hepatosteatosis. Results: The concentrations of oleic acid, amiodarone, and tamoxifen that achieved 80% cell viability comparing to the negative control were used in subsequent experiments to establish the NAFLD and DIFLD models. Those were 0.5 mM oleic acid, 20 µM amiodarone, and 2 µM tamoxifen, respectively. Liraglutide improved cell viability in all models (p < 0,01). All models showed increased lipid accumulation, with DIFLD showing a microsteatotic pattern and NAFLD showing a macrosteatotic pattern, the latter of which corresponds to greater triglyceride accumulation. These steatotic changes were lessened by liraglutide (p < 0,001). GSH levels were reduced in all three models of steatosis. Amiodarone and tamoxifen reduced it to 80% compared to DMEM negative control (p < 0,01, p < 0,05), while effect of OA was smaller. There was no significant difference in GSH concentrations with liraglutide co-treatment. In the hepatocyte steatosis NAFLD and DIFLD models, liraglutide decreased the expression of the lipogenic ACSL1, PPARG, and SREBF1 pathways (p < 0,01). Conclusion: In hepatocyte steatosis NAFLD and DIFLD models established with Huh7 cell culture, liraglutide reduced hepatocyte steatosis through downregulation of lipogenic ACSL1, PPARG, and SREBF1 pathways. |
