Discussion on the Effects of Liangxue Huayu Fang on Autophagy and Apoptosis of Protein Associated Hepatocyte Injury and Its Molecular Mechanism

OBJECTIVE To study the effects of Liangxue Huayu Fang on pathways related to autophagy and apoptosis of hepatocyte injury induced by tumor necrosis factor alpha (TNF-α) combined with D- galactosamine(D-GalN)， and to explore the molecular regulative mechanism.METHODS The normal human liver cells LO2 were cultured in vitro. The hepatocyte injury model was induced by TNF- α (100 ng/mL) and D-GalN(44 μg/mL). Fluorescence microscope was used to observe the morphological changes of cells. The cell viability was detected by MTT; Western blot was adopted to detect the apoptosis and autophagy marker， such as proteins caspase-3， Beclin1， LC3 Ⅰ， LC3 Ⅱ; changes of mTOR protein expression level of apoptosis and autophagy regulating protein Bax， Bcl-2 and mTOR signal pathway were also detected.RESULTS Liangxue Huayu Fang can improve pathological injury of hepatocyte with a dose-dependence; cell apoptosis was decreased and the expression of autophagy markers， such as protein caspase-3， Beclin1， LC3 Ⅱof the treatment group showed decreased and the expression of negative correlative Bcl-2 protein was up-regulated while the positive correlative Bax expression was down-regulated. All the data showed a dose - response relationship to some degree and the expression of mTOR protein showed down-regulated with dose - response.CONCLUSION The protective effects of Liangxue Huayu Fang on hepatocyte injury induced by TNF-α and D-GalN may be related to inhibition of autophagy and apoptosis. Its main mechanism may relate to regulation of the expression of proteins participating in autophagy and apoptosis.