Study on the mechanism of apoptotic signal transduction of murine hepatocytes induced by TGF-β1

Objective: The aim of present study was to determine the relationships between apoptosis induced by TGF-β1 and caspase-3 as well as caspase-8.Methods: Liver cirrhosis was induced in BALB/c mice by means of injection with 50% carbon tetrachloride for 8W. Hepatocytes were isolated from normal and cirrhotic mice by means of an in situ perfusion method with collagenase solution with minor modification. The induction of apoptosis by TGF-β1 was determined in these hepatocytes by electrophoresis on 1.5% agarose gel to observe DNA ladder. To detect apoptosis after treated with TGF-β1 （5ng/ml）, normal hepatocytes were stained with fluorescent DNA-binding dyes Hoechst 33342 and viewed under fluorescent microscope to investigate the effects of caspase-3 and caspases-8 on TGF-β1-induced apoptosis. Results: The viability of hepatocytes isolated from mice by in situ two step collagenase digestion was 95.2%. Internucleosomal fragmented DNA ladder was observed using a 1.5% agarose gel in TGF-β1 treated normal hepatocytes. In comparison, no obvious DNA ladder from cirrhotic hepatocytes was observed. The percentage of apoptotic rate in TGF-β1 treated normal hepatocytes stained with Hoechst 33342 （58.76%） were much higher than hepatocytes untreated with TGF-β1 （18.03%） and could be blocked by Z-IETD-FMK and DEVD-CHO. Conclusion: It was revealed that cirrhotic hepatocytes showed poor response to apoptosis induced by TGF-β1, indicating that inhibitory regulation of cell growth has been impaired in hepatocytes of cirrhotic liver. TGF-β1 can induce apoptosis of murine hepatocytes via caspase-8 and caspase-3-transducted signal pathway.