Chinese Hepatolgy ›› 2016, Vol. 21 ›› Issue (3): 183-190.

• Original Articles • Previous Articles     Next Articles

Effects of salvianolic acid B and hawthorn flavone on lipid deposition and apoptosis of hepatocytes in rats induced by free fatty acids

XUE Dong-ying, XI Bo-ren, ZHANG Jie, YE Jun   

  1. Department of Infectious Disease, PuTuo Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200062, China
  • Received:2015-01-05 Published:2020-07-10
  • Contact: YE Jun, ZHANG Jie, Email: pzxgrk@163.com

Abstract: Objective To investigate the effects of salvianolic acid B and hawthorn flavone on free fatty acids (FFA)-induced lipid deposition and apoptosis of hepatocytes in rats, and its potential mechanism.Methods HepG2 cells and primary cultured hepatocytes isolated from male SD rats by collagenase perfusion were subjected for this study. Oleic acid and palmitic acid (O∶P=2∶1) were used to induce nonalcoholic steatohepatitis (NASH) in primary cultured hepatocytes and HepG2 cells, respectively. The cell models were divided into 11 groups, including normal control group, FFA group, FFA + salvianolic acid B group, FFA+ hawthorn flavone group, FFA + salvianolic acid B + hawthorn flavone group, FFA+ SP600125 group, salvianolic acid B group, hawthorn flavone group, hawthorn flavone + salvianolic acid B group, SP600125 group and DMSO control group, respectively. Correspondingly, the cell models of NASH in primary cultured hepatocytes and HepG2 cells were stimulated with different doses of FFA (250 μmol/L and 500 μmol/L ), salvianolic acid B (10-6mol/L), hawthorn flavone (5 μg/mL) and SP600125 (10 μmol/L), respectively. In order to evaluate the therapy effects of salvianolic acid B and hawthorn flavone against NASH, the following tests were performed: Oil Red O staining for lipid deposition, and enzyme-linked immune-sorbent assay (ELISA), chromatin dye and Hoechst 33258 staining for apoptosis. In addition, it was speculated that the therapy effect was associated with c-Jun N-terminal kinase (c-JNK), which was verified by western blot to assess the expression of phosphorylated-JNK (p-JNK). Results After 24h of incubation with FFA alone, lipids in primary cultured hepatocytes and HepG2 cells were significantly increased (P<0.01). Moreover, salvianolic acid B, hawthorn flavoneor and SP600125 sharply reduced lipids deposition, respectively (P<0.01,P<0.05). The intracellular lipids in DMSO control group had no significant difference than that in normal control group. ELISA showed that FFA induced apoptosis in primary hepatocytes and HepG2 cells (P<0.01). Meanwhile, salvianolic acid B, hawthorn flavone or SP600125 could significantly protect liver from FFA-induced apoptosis (P<0.05, P<0.01). Through chromatin dye and Hoechst 33258 staining, FFA-induced apoptosis in primary cultured hepatocytes and HepG2 cells was also observed under high content screening (HSC) (P<0.01). Salvianolic acid B and hawthorn flavone could significantly reduce the apoptosis (P<0.05, P<0.01), respectively. Western blot analysis showed that FFA treatment led to a significant increase in c-JNK activity (P<0.01) in primary cultured hepatocytes and HepG2 cells, while total JNK levels remained unchanged. Meanwhile, salvianolic acid B and hawthorn flavone could significantly reduce the JNK activity.Conclusion Salvianolic acid B and hawthorn flavone could alleviate FFA-induced apoptosis and lipid metabolism disorders, respectively. Furthermore, they could also inhibit JNK activity, which reveals an important molecular mechanism for treatment of NASH.

Key words: Nonalcoholic steatohepatitis, Salvia, Salvianolic acid B, Free fatty acid, Hepatocyte apoptosis, c-Jun N-terminal kinase