Protective Effects Of Medium-chain Fatty Acids On Drug-induced Liver Injury

The liver is a vital organ of mammals and primarily responsible not only for maintaining nutrient homeostasis by regulating protein,carbohydrate,and fat metabolism,but also exerting detoxification by decomposing and transforming various drugs,toxins and excreting certain metabolites in the body.Therefore,when some drugs were taken overdose,they can overwhelm the liver self-detoxification capacity,leading to liver injury.Drug-induced liver injury(DILI)refers to liver injury induced by the drug itself and its metabolites due to taking conventional or high-dose drugs.DILI is a common adverse drug reaction,and it can lead to liver failure and even death.Medium-chain fatty acids(MCFAs)are widely studied because of their short carbon chains and small molecular weights,which are absorbed easily and supply energy rapidly compared with Long-chain fatty acids(LCFAs).Nevertheless,these studies were hardly focusing on the relationship between MCFA with DILI.In this study,we first explore the conditions of AP and RFP to build DILI model in the normal human liver cell line HL7702(LO2)and further investigate the direct effects of fatty acids(FA)on hepatocellular injury.L02 cells were treated with AP and RFP in this study.respectively.Through MTT colorimetry,liver injury index detection method and inverted fluorescence microscope and flow cytometry methods to explore the best situation of drug concentration and incubation time in order to build the drug-induced liver injury model.In addition,MTT colorimetry,LDH method and Hoechst 33342 staining method were used to investigated the maximum concentration of FA on the premise of minimal toxicity to LO2 cells.and liver injury indicators were added to explore the protective effect of FA on drug-induced liver injury cells.Meantime,to explore the effects of fatty acids on apoptosis,oxidative damage and inflammatory response of drug-induced liver injury cells by the experient of cell apoptosis,detection of oxidative stress indicators and analysis of changes in transcription and translation levels of classic inflammatory factors using real-time fluorescent quantitative and Western Blotting methods.Our results showed that with the increase in the concentration of AP or RFP and treatment time,the survival rate of LO2 cells decreased gradually,the degree of apoptosis aggravated and the activities of both ASTand ALT significantly increased in a dose and time-dependent manner.The results showed that the optimal modeling concentration on the cells was 10 mM of AP or 600μM of RFP for 24 h.These results showed that less than 200μM FA did not cause significant differences on cell viability and LDH activity to LO2 cells.However,with the increase of FA concentration,the cell activity gradually decreased and the cytotoxicity gradually increased.Especially lauric acid and oleic acid at concentrations higher than 200 μM induced remarkable cytotoxic responses.Hoechst 33342 staining assay also showed that 400 μM FA could markedly aggravated apoptosis compared to 200μM FA.Therefore,200μM is the optimal concentration of FA on LO2 cells and be used to further explore the effect of FA on drug-induced liver injury.In this study,MCFA could markedly increased the proliferation of LO2 cells and decreased the leakage of ALT.AST and LDH compared with treatment with LCFA in AP or RFP-induced liver injury cell model(AP or RFP model).In addition,MCFA could reduced cell apoptosis rate and increased mitochondrial membrane potential(MMP).significantly increased the activities of superoxide dismutase(SOD)and superoxide dismutase(CAT),and the levels of Glutathione(GSH)and total antioxidant capacity(T-AOC).and significantly reduced the levels of malonaldehyde(MDA)compared to treatment with LCFA.The results also revealed that the mRNA expression levels of IL-1β,IL-6,IL-8,MCP-1 and TNF-α and the protein expression levels of IL-1β and TNF-α in LCFA groups were significantly higher than MCFA and NR group.The expression levels of IL-1β and TNF-α were significantly decreased in both C8:0 and C10:0 groups compared with the MR group in the RFP model.These results indicated that MCFA treatment may improve AP-or RFP-induced inflammation and cell apoptosis compared to treatment with LCFA.Taken together,we believe that compared with the NR group.LCFA will increase the degree of damage to the DILI cell model by inducing apoptosis,oxidative stress,destroying the integrity of the mitochondrial membrane,and triggering an inflammatory response,while MCFA will not increase its liver damage or even certain protection improvement effect.