Inhibition of Fas-mediated Apoptosis in Yac-1 Cell via Anti-Fas Ribozyme

• 来源期刊: 生物化学与生物物理学报（英文版）
• 学科: 生物学
• 关键词: transplantation immunology ribozyme Fas Yac-1 apoptosis
• 年，卷（期）: 2004,（3）
• 页码范围: 199-205
• 页数: 7页
• 分类号: Q5
• 语种: 英文

摘要

To detect a new and more effective way against apoptosis mouse lymphomatic cell lineYac-1 in which fas gene was expressed highly was used as a model for studying the effects of anti-Fas ribozyme on Fas-mediated apoptosis.A hammerhead ribozyme gene targeting the fas mRNA was synthesized and its in vitro transcription vector was constructed,which was transfected into Yac-1 cells using electroporation.Rz596 expression was detected using RT-PCR,and Fas expression in Yac-1 cells was detected using RT-PCR,Western blot and flow cytometry.After treated with anti-Fas antibody(JO2),Yac-1cell viability was measured with MTT assay,caspase-3 proteolytic activity was detected,and cell apoptosis was measured according to annexin V apoptosis detecting kit.Anti-Fas ribozyme could cleave fas mRNA efficiently in vivo and in vitro.Fas expression in Yac-1 cells transfected with anti-Fas ribozyme was decreased remarkably and correlated with resistance to Fas-mediated apoptosis as determined by flow cytometry and caspase-3 proteolytic activity.Anti-Fas ribozyme was detected in cells transfected with pU6-RZ596 and pU6-dRZ596 and could remarkably decrease the Fas expression in Yac-1 cells,which made Yac-1 cells get rid of Fas-mediated apoptosis.Because of wide expression of fas in organs and tissues,our research was very useful for studying the inhibition of apoptosis of many organs and tissues in the future.