二甲双胍调节糖尿病大鼠心房小电导钙激活钾通道亚型蛋白表达的信号通路  

Metformin can regulate the signaling pathway of KCa2.2 and KCa2.3 protein expressions in atrial SKCa of type 2 DM rats

作　　者：刘长河 付茜 李斌[1] 潘一龙[1] 刘贞[2] 李晓东[1] Liu Changhe;Fu Xi;Li Bin;Pan Yilong;Liu Zhen;Li Xiaodong(Department of Cardiology,Affiliated Shengjing Hospital of China Medical University,Shenyang 116001,Liaoning Province,China)

机构地区：[1]中国医科大学附属盛京医院第一心血管内科,沈阳116001 [2]新汶矿业集团中心医院心血管内科

出　　处：《中华老年心脑血管病杂志》2020年第1期70-74,共5页Chinese Journal of Geriatric Heart,Brain and Vessel Diseases

基　　金：辽宁省自然科学基金（201602839）

摘　　要：目的探讨蛋白激酶A(PKA)/钙调蛋白Ⅱ(CaMKⅡ)信号通路在二甲双胍(Met)调节2型糖尿病(T2DM)大鼠心房小电导钙激活钾通道亚型KCa2.2和KCa2.3蛋白表达中的作用。方法健康雄性Wistar大鼠40只,随机选8只喂普通饲料为对照组(Con组),另32只喂高脂高糖饲料联合腹腔注射小剂量链脲佐菌素构建T2DM大鼠模型后,再随机分为DM组、Met组、H-89组(腹腔注射PKA抑制剂H-89)和KN-93组(腹腔注射CaMKⅡ抑制剂KN-93),每组8只。用ELISA检测大鼠心房组织PKA活性,qRT-PCR检测CaMKⅡmRNA表达,Western blot和免疫组织化学检测KCa2.2、KCa2.3和磷酸化CaMKⅡ(p-CaMKⅡ)蛋白表达。结果 Con组、DM组、Met组和H-89组PKA活性分别为0.74±0.04、0.50±0.05、0.69±0.03和0.48±0.03。与DM组比较,Met组PKA活性明显提升(P<0.01);与Met组比较,H-89组显著抑制PKA活性(P<0.01)。Con组、DM组及Met组CaMKⅡmRNA分别为1.00±0.07、0.61±0.03和0.92±0.09。与Con组比较,DM组CaMKⅡmRNA表达明显降低(P<0.01);与DM组比较,Met组CaMKⅡmRNA表达明显增加(P<0.01)。与Con组比较,DM组心房组织p-CaMKⅡ和KCa2.2蛋白表达均明显降低,KCa2.3蛋白表达明显升高(P<0.01)。与DM组比较,Met组明显提升p-CaMKⅡ和KCa2.2蛋白表达,明显抑制KCa2.3蛋白表达(P<0.01)。与Met组比较,KN-93组和H-89组分别显著抑制p-CaMKⅡ蛋白表达和PKA活性,均显著下调KCa2.2蛋白表达,上调KCa2.3蛋白表达(P<0.01)。免疫组织化学染色显示,与Met组比较,KN-93组和H-89组均显著下调KCa2.2蛋白表达,上调KCa2.3蛋白表达(P<0.05,P<0.01),与Western blot检测结果一致。结论 Met通过激活PKA/CaMKⅡ信号通路部分修复T2DM大鼠心房KCa2.2蛋白下调和KCa2.3蛋白上调。Objective To study the role of PKA/CaMKⅡsignaling pathway in metformin-mediated expressions of KCa2.2 and KCa2.3 proteins in atrial SKCa of type 2 DM(T2 DM)rats.Methods Of the 40 healthy male Wistar rats,8 fed on a common diet served as a control group,the other 32 fed on a high fat and carbobhydrate diet were divided into T2 DM group,metformin group,H-89 group and KN-93 group(8 in each group)after a T2 DM model was established by intraperitoneal injection with a small dose of streptozotocin.The activity of PKA and expressions of CaMKⅡmRNA,KCa2.2,KCa2.3,p-CaMKⅡproteins in atrial tissue of rats were detected by ELISA,qRT-PCR,Western blot respectively with immunohistochemical staining.Results The activity of PKA was 0.74±0.04,0.50±0.05,0.69±0.03,0.48±0.03 respectively in control group,T2 DM group,metformin group,H-89 group,which was significantly higher in metformin group than in DM group(P<0.01)and significantly lower in H-89 group than in metformin group(P<0.01).The expression levels of CaMKⅡmRNA were 1.00±0.07,0.61±0.03 and 0.92±0.09 respectively in control group,T2 DM group and metformin group,which were significantly lower in DM group than in control group(P<0.01)and significantly higher in metformin group than in DM group(P<0.01).The expression levels of p-CaMKⅡand KCa2.2 proteins were significantly lower while those of KCa2.3 protein were significantly higher in DM group than in control group(P<0.01).The expression levels of p-CaMKⅡand KCa2.2 proteins were significantly higher while those of KCa2.3 were significantly lower in metformin group than in T2 DM group(P<0.01).KN-93 group and H-89 group significantly inhibited p-CaMKⅡexpression and PKA activity respectively,both downregulated KCa2.2 expression and upregulated KCa2.3 expression.The expressions of CaMKⅡmRNA,KCa2.2,KCa2.3 detected with immunohistochemical staining were consistent with those detected by Western blot(P<0.05,P<0.01).Conclusion Metformin can upregulate the expression of KCa2.2 and downregulate that of KCa2.3 in atria

关 键 词：二甲双胍 糖尿病 小电导钙激活钾通道 环AMP依赖性蛋白激酶类 钙调蛋白 

分 类 号：R965[医药卫生—药理学]