AIM: TO investigate the effects of anti-Fas ribozyme on Fas expression and apoptosis in primary cultured mouse hepatocytes.METHODS: Mouse hepatocytes were isolated by using collagenase irrigation. A hammerhead ribozym...AIM: TO investigate the effects of anti-Fas ribozyme on Fas expression and apoptosis in primary cultured mouse hepatocytes.METHODS: Mouse hepatocytes were isolated by using collagenase irrigation. A hammerhead ribozyme targeting the Fas mRNA was constructed, and transfected into mouse hepatocytes via Effectene. Then Fas expression in mouse hepatocytes was detected by RT-PCR and western blotting.After being treated with anti-Fas antibody (JO2), hepatocytes viability was measured with MTI- assay. Caspase-3 proteolytic activity was detected, and cell apoptosis was measured according to Annexin V-FITC apoptosis detection kit.RESULTS: Fas expressed in primary mouse hepatocytes.Fas expression in hepatocytes transfected with anti-Fas ribozyme was decreased remarkably and correlated with the resistance to Fas-mediated apoptosis as determined by flow cytometry and caspase-3 proteolytic activity.CONCLUSION: AnU-Fas ribozyme can remarkably decrease the Fas expression in mouse hepatocytes, thus inhibit Fasmediated apoptosis in hepatocytes, It is suggested that anti-Fas ribozyme could significantly increase the resistance of transplanted hepatocytes to apoptosis and improve the survival of transplanted hepatocytes,展开更多
基金Supported by the National Natural ScienceFoundation of China,No.30240022
文摘AIM: TO investigate the effects of anti-Fas ribozyme on Fas expression and apoptosis in primary cultured mouse hepatocytes.METHODS: Mouse hepatocytes were isolated by using collagenase irrigation. A hammerhead ribozyme targeting the Fas mRNA was constructed, and transfected into mouse hepatocytes via Effectene. Then Fas expression in mouse hepatocytes was detected by RT-PCR and western blotting.After being treated with anti-Fas antibody (JO2), hepatocytes viability was measured with MTI- assay. Caspase-3 proteolytic activity was detected, and cell apoptosis was measured according to Annexin V-FITC apoptosis detection kit.RESULTS: Fas expressed in primary mouse hepatocytes.Fas expression in hepatocytes transfected with anti-Fas ribozyme was decreased remarkably and correlated with the resistance to Fas-mediated apoptosis as determined by flow cytometry and caspase-3 proteolytic activity.CONCLUSION: AnU-Fas ribozyme can remarkably decrease the Fas expression in mouse hepatocytes, thus inhibit Fasmediated apoptosis in hepatocytes, It is suggested that anti-Fas ribozyme could significantly increase the resistance of transplanted hepatocytes to apoptosis and improve the survival of transplanted hepatocytes,
