过表达磷脂酶Cβ1对葡萄糖刺激胰岛素分泌的影响.docVIP

过表达磷脂酶Cβ1对葡萄糖刺激胰岛素分泌的影响　　　　　　　　　 周恒宇1，邓华聪1*，郑宏庭2，蹇 锐2，阮燕2 （1 重庆医科大学附属第一医院内分泌科，重庆 400016；2第三军医大学微生物教研室，重庆 400038）　 提要：目的　在大鼠胰岛素瘤细胞系――INS-1细胞中过表达磷脂酶C（phospholipase C,PLC）β1亚型,观察INS-1细胞胰岛素分泌，探讨PLCβ1在葡萄糖刺激胰岛素分泌（glucose-stimulated insulin secretion GSIS）信息传递中的作用。方法　（1）确定PLCβ1与GSIS的联系。（2）构建PLCβ1真核表达载体（PCMV-HA--PLCβ1）转染INS-1细胞,设转染空载体的对照组。（3）Western-blot证实PLCβ1在INS-1细胞中表达水平上调。（4）用大鼠胰岛素ELISA试剂盒检测转染后的INS-1细胞培养上清液胰岛素含量。结果　获得过表达PCMV-HA--PLCβ1的INS-1细胞株与对照组相比，过表达PLCβ1的INS-1细胞株培养上清液的胰岛素含量增加。结论　过表达PLCβ1显著增加INS-1细胞的胰岛素分泌，提示PLCβ1可能是参与GSIS信息传递的主要亚型之一。　　 关键词：磷脂酶Cβ1；葡萄糖刺激胰岛素分泌；过表达；胰岛素；瞬时转染 Effects of overexpression of phospholipase cβ1 on glucose-stimulated insulin secretion ZHOU Heng-Yu1,DENG Hua-cong1*, ZHENG Hong-Ting2,JIAN Rui2,RUAN Yan2, (Department of Endocrinology, First Affiliated Hospital of Chongqing Medical University, Chongqing 400016; Department of Microbiology, Third Millitary Medical University, Chongqing 400038, China) Corresponding author:DENG Hua-cong Abstract:Objective To determine the role of PLCβ1 in GSIS.Methods (1)Make sure there is the relation between PLCβ1 and GSIS.(2)Eukaryotic expression vector PCMV-HA--PLCβ1 was constrcted and then transient transfected into INS-1.(3)The PLCβ1 protein was detected by western blot.(4)Using the rat insulin ELISA kit to detect the content of insulin in INS-1’s supernatant liquid.Results Cell strains of transient overexpression PLCβ1 INS-1-- PCMV-HA--PLCβ1 was obtained.As comparing to the control,the content of insulin in INS-1’s supernatant liquid was raised.Conclusion Transient overexpression PLCβ1 in INS-1 had effects on insulin secretion,so PLCβ1 probably participated the signal transduction pathway of GSIS in INS-1. Key words: PLCβ1,GSIS,overexpression,insulin,transient transfection 　 磷脂酶c(Phospholipase C，PLC)是磷脂酰肌醇信号通路的关键酶，在体内分布极为广泛。在哺乳动物中已鉴定出15种PLC亚型,根据其不同的氨基酸序列分为6大类: PLCβ、PLCγ、PLCδ、PLCε和新近发现的PLCζ[1]及PLC[2]，PLCβ1是15种PLCs亚型中的一种。 Sophia Thore[3]等提出PLC参与葡萄糖刺激的胰岛素分泌（GSIS）信息传递，随着对PLC分类的细化，何种PLC亚型参与GSIS成为亟待解决的问题。近期发现[4]大鼠胰