The Relationgship Between Serine Palmitoyl Transferase And Oleic Acid Induced Hepatic Stellate Cell Apoptosis And Autophagy

Objective:To investigate The relationship between the de novo ceramide synthesis pathway and oleic acid induced hepatic stellate cell apoptosis and autophagy.Methods: Rat hepatic stellate cells, HSC-T6, were administrated with concentration gradient of oleic acid(0 umol/L ?25umol/L?50umol/L ?100 umol/L?150umol/L). After 24 h, cell viability was measured by cell counting MTT,Select three concentration, less effect on cell activity, as the best concentration of oleic acid induced hepatic stellate cells. Experiment is divided into blank control group and three different concentration of oleic acid induced group.Expression of Serine palmitoyltransferase(SPT) mRNA in HSC-T6 cells was quantified by reverse transcriptase polymerase chain react(RT-PCR). The autophagy lysosomal levels of HSC-T6 cells were measured using MDC staining method,flow cytometric detection of HSC apoptosis, real time PCR and Western blot detection of apoptosis protein Bcl-2, Bax; Western blot detection of apoptosis proteins Bcl-2, Bax and autophagy protein LC3 B, Beclin-1, and autophagy gene Atg5.Results:1.The inhibition rate of HSC-T6 cells in oleic acid induced group was significantly higher than that in control group(P<0.01), and the higher the concentration of oleic acid, the higher the inhibition rate.2. The apoptosis rate of HSC-T6 cells induced by oleic acid group increased significantly higher than that in control group(P<0.01),with increased oleic acid concentration, the apoptosis rate increased gradually.3. The HSC-T6 cells induced by oleic acid group autolysosome significantly more than the control group, and with the increase of the concentration of oleic acid, the formation of HSC-T6 cell autophagy lysosome increased gradually.4. There was a significant positive correlation between SPT mRNA expression and apoptosis rate in HSC-T6 cells induced by oleic acid at different concentrations(r=0.963, P<0.01).5. There was a significant positive correlation between SPT mRNA expression and the expression of Pro apoptotic protein Bax in HSC-T6 cells induced by oleic acid at different concentrations(r=0.958, P<0.01), there was a significant negative correlation between SPT expression and anti apoptotic protein Bcl-2 expression in HSC-T6 cells induced by different concentrations of oleic acid(r=-0.971, P<0.01).6.There was a significant positive correlation between SPT mRNA expression and the expression of autophagy mark protein lc3 b in HSC-T6 cells induced by oleic acid at different concentrations(r=0.993, P<0.01), there was a significant positive correlation between SPT expression and the expression of autophagy mark protein beclin-1 in HSC-T6 cells induced by different concentrations of oleic acid(r=0.986, P<0.01).Conclusions:1.Oleic acid can inhibit the proliferation of HSC-T6 cells, promote the autophagy and apoptosis, and has a concentration dependent.2. The relationship between ceramide de novo synthesis pathway key enzyme serine palm acyl coenzyme A transferase(SPT) and Autophagy and apoptosis of HSC-T6 cells induced by oleic acid was positively correlated.