异丙肾上腺素对NCX1.1基因编码钠钙交换体电流的影响  

Effect of isoproterenol on sodium-calcium exchanger current encoded by NCX1.1 gene

作　　者：项国剑 张建成[1,2] 李泱[3] 陈茜[2,4] Xiang Guojian;Zhang Jiancheng;Li Yang;Chen Qian(Department of Cardiology,Fujian Provincial Hospital,Fuzhou 350001,China;Provincial Clinical Medicine College of Fujian Medical University,Fuzhou 350001,China;Institute of Geriatric Cardiology,Chinese PLA General Hospital,Beijing 100853,China;Department of Intensive Care Medicine,Fujian Provincial Hospital,Fuzhou 350001,China)

机构地区：[1]福建省立医院心血管内科,福州350001 [2]福建医科大学省立临床医学院,福州350001 [3]中国人民解放军总医院老年心血管病研究所,北京100853 [4]福建省立医院重症医学科,福州350001

出　　处：《中华老年多器官疾病杂志》2023年第2期124-129,共6页Chinese Journal of Multiple Organ Diseases in the Elderly

基　　金：国家自然科学基金(82070341)。

摘　　要：目的探讨β肾上腺素受体对钠钙交换电流(I_(NCX))的调控及可能的信号转导途径。方法利用免疫磁珠阳性标记表达系统将NCX1.1质粒转染人胚肾293(HEK-293)细胞,选取103个转染阳性的HEK-293细胞,随机分成空白对照组(n=20)、灌流异丙肾上腺素(ISO)组(n=20)、灌流Gi蛋白抑制剂百日咳毒素及ISO组(n=20)、灌流环磷酸腺苷(cAMP)合成激动剂毛喉素(forskolin)及ISO组(n=22),灌流蛋白激酶A(protein kinase A,PKA)抑制剂H89及ISO组(n=21)。运用全细胞膜片钳技术记录各组I_(NCX)的变化。采用SPSS 21.0统计软件进行数据分析。根据数据类型,分别采用t检验或ANOVA方差分析进行组间比较。结果以灌流用药前I_(NCX)为空白对照组,ISO组可使内向I_(NCX)密度从(-6.07±1.53)pA/pF增加至(-7.89±1.61)pA/pF(n=20,P<0.05),平均增加约30%。而预先使用G蛋白-cAMP-PKA通路关键分子激动剂或抑制剂后,ISO对I_(NCX)的效应均发生明显改变。其中,PTX及forskolin可使ISO对I_(NCX)密度增加效应更加显著,电流密度分别增至(-10.02±1.99)pA/pF及(-10.78±1.77)pA/pF,与单纯使用ISO相比,差异有统计学意义(n=20,P<0.01),提示Gi蛋白抑制和环磷酸腺苷激动均可增加ISO的作用。而预先应用H89,ISO增加I_(NCX)的效应几乎消失,电流密度约为(-6.22±1.70)pA/pF,与对照组相比无显著差异(n=20,P>0.05),提示抑制PKA可基本阻断ISO对I_(NCX)的刺激效应。结论β肾上腺素受体激动可增加NCX1.1内向电流,其可能通过G蛋白-cAMP-PKA信号通路参与调节。Objective To investigate the regulation ofβ-adrenoceptor on sodium-calcium exchange current(I_(NCX))and its possible signal transduction pathway.Methods The NCX1.1 plasmid was transfected into human embryonic kidney-293(HEK-293)cell by immunomagnetic-bead-based positive expression system.Transfected HEK-293 cells totaling 103 with positive transfection were selected and randomly divided into blank control group(n=20),isoproterenol(ISO)group(n=20),pertussis toxin(PTX)plus ISO group(n=20),forskolin plus ISO group(n=22),and H89[protein kinase A(PKA)inhibitor]plus ISO group(n=21).I_(NCX)changes in each group were recorded with whole-cell patch clamp technique.SPSS statistics 21.0 was used for statistical analysis.According to data type,t-test was used for comparison between two groups,and ANOVA was used for comparison between multiple groups.Results Against control group,ISO increased the inward I_(NCX)density from(-6.07±1.53)pA/pF to(-7.89±1.61)pA/pF(n=20,P<0.05),with an average increase of about 30%.However,the effect of ISO on I_(NCX)current was significantly changed after pre-administration of key molecular agonists or inhibitors of Gi-cAMP-PKA pathway.PTX and forskolin,significantly enhanced the effect of ISO,had a more significant effect on I_(NCX)density increase,the current density increased to(-10.02±1.99)pA/pF and(-10.78±1.77)pA/pF,respectively.Compared with ISO alone,there was a significant difference(n=20,P<0.01),suggesting that both the inhibition of Gi protein and activation of cAMP can enhance the effect of ISO.However,the effect of pre-treatment with PKA inhibitor H89 showed no enhancement on I_(NCX),and the current density was about(-6.22±1.70)pA/pF,which was not significantly different from that of the control group(n=20,P>0.05).It suggested that PKA inhibition can basically block the effect of ISO on I_(NCX).Conclusionβ-adrenoceptor activation may enhance the NCX 1.1 inward current,probably through stimulating G protein-cAMP-PKA signaling pathway.

关 键 词：Β肾上腺素受体 钠钙交换体电流 钙超载 G蛋白信号通路 

分 类 号：R318.04[医药卫生—生物医学工程]