二氢生物喋呤还原酶减轻脂肪酸诱导的细胞凋亡的机制研究  

The mechanism of QDPR on reducing apoptosis induced by fatty acids

作　　者：古艳婷[1] 冯茹[1] 孙斯凡 

机构地区：[1]航空总医院药剂科,北京100012

出　　处：《安徽医科大学学报》2018年第2期181-184,共4页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金(编号:81603091)

摘　　要：目的探讨二氢生物喋呤还原酶(QDPR)高表达对脂肪酸(PA)诱导的细胞凋亡的作用。方法 HEK293T细胞转染实验分为A、B、C 3组,A组为空载体转染组,B组为经过PA刺激的空载体转染组,C组为经过PA刺激的QDPR重组质粒转染组。首先将空载体及构建的QDPR重组质粒分别转染至HEK293T细胞,培养24 h后采用0.5 mmol/L PA刺激上述细胞,将没有经过PA刺激的空载体组细胞、PA刺激的空载体组细胞和PA刺激QDPR重组质粒组细胞三组细胞继续培养24 h后收集细胞。采用四氢生物喋呤(BH_4)和活性氧簇(ROS)检测试剂盒检测3组BH_4和ROS的生成量;采用Western blot法检测3组凋亡相关蛋白Beclin1、Caspase 3和Beclin 2的表达。结果 (1)转染细胞后,QDPR融合蛋白成功表达;(2)与A组相比,B组的BH_4生成量差异无统计学意义;与B组相比,C组BH_4生成量明显增多(P<0.05);(3)与A组相比,B组的ROS生成量明显增加(P<0.05);与B组相比,C组ROS生成量明显下降(P<0.05);(4)Western blot结果显示:与A组相比,B组经过PA刺激后细胞中Beclin 1和Caspase 3表达水平显著上调(P<0.05),Beclin 2表达水平下降(P<0.05);QDPR过表达后C组Beclin 1和Caspase 3表达水平显著下调(P<0.05),Beclin 2表达水平升高(P<0.05)。结论 QDPR高表达后,可以刺激BH_4的生成,同时能降低PA诱导的ROS的生成量,还可以降低细胞凋亡相关蛋白Beclin 1和Caspase 3的表达,增强抗凋亡蛋白Beclin 2的表达,提示其可能通过调节BH_4和ROS含量影响细胞凋亡。Objective To investigate the effects of dihydropteridine reductase （QDPR） on regulating apoptosis induced by plamitie acid（PA）. Methods The transfection of HEK293T cells experiment was divided into 3 groups. A group was the control vector group. B group was the control vector group induced by PA. C group was the recombinant plasmid QDPR group induced by PA. First, control vector and recombinant plasmid QDPR was respectively transfected into HEK293T cells. After 24 h, PA with concentration of 0. 5 mmol/L was added into the medium of above cells. The cells of control vector group, the cells of control vector group induced by PA and the cells of recombinant plasmid QDPR group induced by PA were cultured for another 24 hours. At last, cells were harvested to detect tetrahydrobiopterin （BH4） and reactive oxygen species（ROS） generation, Beclin 1, Caspase 3 and Beclin 2 expression. Results ①After transfection, the recombinant plasmid QDPR was successfully constructed and expressed in cells. ② There was no significant difference between A group and B group in BH4 generation. Compared with B group, BH4 generation increased in C group （P 〈 0. 05 ）. ③ROS generation was increased in B group compared with A group,and decreased ROS generation in C group compared with B group （P 〈 0. 05）. ④ Western blot analysis revealed that Beclin 1 and Caspase 3 increased （P 〈 0.05 ） while Beelin 2 decreased in B group compared with A group （P 〈 0. 05）. Compared to B group, Beclin 1 and Caspase 3 decreased while Beclin 2 increased in C group （P 〈 0. 05 ）. Conclusion QDPR may regulate apoptosis induced by fatty acids by decreasing the generation of ROS and increasing the level of BH4 and the expression of Beclin 2 associated with anti-apoptosis.

关 键 词：二氢生物喋呤还原酶 HEK293T细胞 凋亡 BECLIN1 活性氧簇 

分 类 号：R587.24[医药卫生—内分泌]