Studies On The Mechanisms For Hepatotoxicity Of Saikosaponin D In LO2Cells

Objective:Herbal hepatotoxicity has been increasingly reported in clinical context, but the underlying mechanisms are poorly understood. Saikosaponin D (SSD) is a major component of saikosaponins isolated from Bupleurum falactum, a herb that has been linked to hepatotoxicity. Our current study was to examine the toxic effect of SSD on human hepatocyte LO2cells and explore the possible mechanism.Methods:We study LO2liver cell lines in vitro, and mice fed SSD in vivo. MTS, HE staining, TUNEL staining, LDH detection, Hoechst staining, flow cytometry, immunofluorescence, Western blotting, RNA interference and other experimental techniques detected. Observations:1) the role of liver injury induced after administration SSD in mice.2) mitochondria] damage and apoptosis-related proteins of Bcl-2/Bax, caspase-3/c-caspase-3, caspase-8/c-caspase-8, c-caspase-9or other expression changes after SSD treated in LO2cells in vitro.3) the impact of SSD on PDGF-βR/p38MAPK signaling pathway, and the relationship between PDGF-βR/p38and the downstream apoptotic pathway.4) the impact on fas-mediated extrinsic apoptotic signaling pathway.Results:SSD reduced cell viability and led to dramatic morphological alterations in LO2cells. Hoechst staining and flow cytometry analyses showed that SSD stimulated hepatocyte apoptosis. SSD-treated cells exhibited apparent nuclear condensation and fragmentation, and the apoptotic cells were increased by SSD dose-dependently. Subsequent experiments showed that SSD decreased mitochondrial membrane potential and downregulated Bcl-2but upregulated Bax. Moreover, caspase-9and caspase-3were activated in SSD-treated LO2cells. These data consistently indicated that SSD stimulated mitochondrial apoptosis in hepatocytes. Mechanistic investigations showed that SSD disrupted p38signaling and that p38specific inhibitor SB203580mimicked the pro-apoptotic effect of SSD. In addition, platelet-derived growth factor-β receptor (PDGF-PR) blocker imatinib reduced p38phosphorylation and also mimicked the pro-apoptotic effect of SSD in LO2cells. SSD also activated fas signaling pathway and collaborative with mitochondrial apoptosis signaling pathway in LO2.Conclusion:These data collectively indicated that SSD induced apoptosis by interrupting PDGF-PR/p38pathway and fas signaling pathway in LO2hepatocytes.