FasL基因转染对大鼠肥大细胞凋亡的影响

Fas Ligand gene transfection induces rat mast cells apoptosis

目的观察FasL基因转染对大鼠肥大细胞凋亡的影响。方法RTPCR法扩增大鼠FasL穿膜区和胞外区cDNA,成功构建pcDNA31FasL真核表达质粒,瞬时转染RBL2H3,RTPCR、免疫印迹法鉴定FasL在RBL2H3上的表达,AnnexinV流式细胞检测pcDNA31FasL转染RBL2H3后细胞的凋亡情况。结果获得FasL穿膜区和胞外区cDNA,构建pcDNA31FasL真核表达质粒,瞬时转染RBL2H3后在细胞膜和上清均有FasL的存在,瞬时转染RBL2H3后48小时,细胞发生凋亡。结论肥大细胞可以通过FasFasL途径凋亡,为FasL基因应用于过敏性疾病的治疗提供依据。

Objective:To show whether the Fas Ligand gene induces mast cells apoptosis.Methods:RT-PCR was used to amplify the gene of rat Fas ligand extracelluar domain and transmembrane domain and cloned it into eukaryotic expression plasmid pcDNA3.1.Transcent transfect RBL-2H3,the expression of Fas ligand RBL-2H3 was detected by RT-PCR、Western blot.The Annexin V FCM was used to detect the RBL-2H3 apoptosis after the transfection of Fas Ligand.Results:It is successful to obtain the gene of rat Fas Ligand extracellular domain and transmembrane segment,cloning it into pcDNA3.1,FasL was expressed on the surface of RBL-2H3 and it's supernatant after the transfection of pcDNA3.1/FasL.The cell start to be apoptosis.Conclusion:Our study reveals that Fas Ligand gene transfection in RBL-2H3 can effectively induced apoptosis.It is a promising strategy for Fas Ligand to be used in the therapy of allergic disease. [