Objective: The present study was designed to examine whether Liangxue Huayu recipe (LHR) had the protective effects on D-galactosamine(D-GalN) and tumor necrosis factor-α (TNF-α)-treated human L02 hepatocytes and its possible association through the mitochondrial pathway. Method: Hepatocytes L02 injured by D-GalN and TNF-α was treated by LHR. The morphological changes of the cells were observed under inverted phase-contrast microscopy. The percentage of cell viability was evaluated by MTT. Apoptosis and mitochondrial membrane potential (MMP) were determined by high content screening (HCS) assay. In addition

the expression levels of apoptosis protease activation factor-1(Apaf-1)

apoptosis induced factor (AIF)

cleaved Caspase-3 and cleaved Caspase-9 were detected by Western-blot analysis in L02 cells. Result: It revealed that LHR pretreatment improved the morphology of TNF-α/GalN-treated human L02 hepatocytes. The study also showed that incubation with LHR caused significant increase in the viability of L02 cell and decrease of cell apoptosis. Furthermore

LHR prevented the loss of MMP

inhibited the upregulation of Apaf-1 and AIF protein expression

which was accompanied with a decrease in cleaved caspase-3 and cleaved caspase-9 activation in this cell model. Conclusion: These results indicate that LHR is effective in attenuating hepatocyte apoptosis in human L02 cells

and this effect is partly mediated through prevention on the loss of MMP and subsequent regulation of particular proapoptotic gene expression.