Inhibition of mouse hepatocyte apoptosis via anti-Fas ribozyme.
OBJECTIVE
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 MTT 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.


CONCLUSIONS
Anti-Fas ribozyme can remarkably decrease the Fas expression in mouse hepatocytes, thus inhibit Fas-mediated 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.
Institute of Hematology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, Hubei Province, China.
Min
M
Zhang
Wei
W
He
Fang
F
Liu
Ping
P
Zou
Juan
J
Xiao
Zhao-Dong
ZD
Zhong
Zhong-Bo
ZB
Hu
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D002470:Cell Survival / Q000502:physiology; D002478:Cells, Cultured; D015870:Gene Expression; D022781:Hepatocytes / Q000166:cytology*; D051379:Mice; D016337:RNA, Catalytic / Q000235:genetics*; D014162:Transfection; D019014:fas Receptor / Q000235:genetics*
D016337:RNA, Catalytic; D019014:fas Receptor; C427204:hammerhead ribozyme; C505409:Casp3 protein, mouse; D053148:Caspase 3; D020169:Caspases