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Liraglutide reduces hepatosteatosis in NAFLD and drug-induced fatty liver cell culture models through PPARγ signaling pathway

Background: With the increasing prevalence of obesity, fatty liver injury, and drug use, appropriate treatment of non-alcoholic (NAFLD) and drug-induced fatty liver disease (DIFLD) is becoming more important. Numerous studies have proposed and tested a number of potential hepatoprotective agents. The aim of this study was to establish reliable cell culture models for NAFLD and DIFLD and to determine the effect of the GLP-1 agonist liraglutide in these models. Methods: Huh7 cells were grown overnight to reach 90% confluency, followed by 24h of incubation according to further described protocols. Experiments were performed in triplicates, with the following cell subgroups: Huh7 cells incubated in medium only as a negative control, cells incubated with increasing concentrations of liraglutide (5nM, 10nM, 20nM), cells treated with 0, 5 mM oleic acid (OA) (NAFLD model) , and/or cotreated with liraglutide, and cells treated with 20μM amiodarone (DIFLD model), and/or cotreated with liraglutide. The antiproliferative and cytotoxic effects were determined using a colorimetric MTT assay. Cells were stained with Oil-Red-O dye and Fluorescent mounting medium with DAPI, and visualized under a microscope. ImageJ software was used to count the cell nuclei and measure the integrated density relative to the cell count. After extraction of total RNA with NucleoZol, cDNA synthesis, PCR method and gel electrophoresis were performed, and finally the signals were analyzed by ImageJ software. Results: Both oleic acid and amiodarone reduced cell viability by 30% (p<0, 001, p<0, 05) compared to the control. Cells cotreated with lower concentrations of liraglutide (5 and 10nM) showed a slight increase in viability (p<0, 05), while higher concentration had an opposite effect. Both amiodarone and oleic acid significantly increased integrated density (p<0, 001), while 5nM liraglutide decreased it in cells cotreated with amiodarone (p<0, 05). PPARγ gene expression was decreased in both oleic acid and amiodarone treated cells, and 10nM liraglutide exerted greatest increase in PPARγ gene expression. Conclusions: Hepatoprotective effect of liraglutide in both models explored in this study is mediated through PPARγ signaling pathway.

Liraglutide ; non-alcoholic fatty liver disease ; amiodarone ; cell culture

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