PRKAG2心脏综合征中 R302Q 突变对心肌细胞糖原和钙离子稳态的影响

摘要

目的：：探讨 PRKAG2心脏综合征中 R302Q 突变对心肌细胞糖原和钙离子稳态的影响。方法：用表达增强型绿色荧光蛋白(enhanced green fluorescent protein,EGFP)、PRKAG2-WT、PRKAG2-R302Q 的腺病毒感染大鼠 H9C2心肌细胞,同时设正常对照组。感染24 h 和48 h 后,采用过碘酸–雪夫(Periodic acid-Schiff,PAS)染色法进行糖原染色,观察心肌细胞的糖原贮积；采用酶联免疫吸附试验(ELISA)法检测细胞内糖原含量；同时用 Rohd-2/AM 活体染料进行钙离子染色,然后采用流式细胞术分析钙离子库信号差异。结果：EGFP 组与对照组糖原染色和含量结果无明显差别；PRKAG2-WT 组糖原染色和含量略微增强；PRKAG2-R302Q 组糖原染色和含量明显增强。各组的钙离子稳态并未受到影响。结论：PRKAG2心脏综合征中R302Q 突变导致心肌细胞内糖原贮积,而钙离子稳态并未受到影响,没有钙超载现象发生。%Objective:To investigate the effect of R302Q mutation on glycogen and calcium ion homeostasis of cardiomyocytes in PRKAG2 cardiac syndrome.Methods:The rat H9C2 cardiomyocytes were infected with adenovirus containing enhanced green fluorescent protein(EGFP),PRKAG2-WT,and PRKAG2-R302Q,meanwhile,normal control group was set up.After 24 h and 48 h,Periodic acid-Schiff(PAS)staining was used to observe cardiomyocyte glycogen storage,and the glycogen con-tent in cells was detected by enzyme-linked immunosorbent assay (ELISA).Meanwhile,Rohd-2/AM vital dye was used to car-ry out calcium ion staining,and then the signal differences among calcium stores were analyzed by flow cytometry.Results:There was no obvious difference regarding glycogen staining and content between EGFP group and control group.Glycogen staining and content was slightly enhanced in PRKAG2-WT group,while glycogen staining and content was obviously enhanced in PRKAG2-R302Q group.However,the calcium ion homeostasis in each group was not affected.Conclusions:R302Q muta-tion leads to intra cardiomyocyte glycogen accumulation in PRKAG2 cardiac syndrome.However,calcium ion homeostasis is not affected and there is no calcium overload.