CAJ | 학술논문

目的初步探讨缺血再灌注后心肌细胞胰岛素抵抗现象的分子机制.方法建立成年大鼠心肌细胞模拟缺血再灌注模型,应用同位素示踪技术评价心肌细胞的胰岛素敏感性.Westemblot分析检测心肌细胞葡萄糖转运蛋白4(GLUT4)的转位.荧光分光光度仪测定模拟缺血再灌注心肌细胞游离钙离子浓度([Ca2+]i).结果模拟缺血再灌注心肌细胞表现出明显的胰岛素抵抗.胰岛素刺激再灌注心肌细胞的GLUT4转位较对照组明显降低(P<0.05).同时,模拟缺血再灌注心肌细胞[Ca2+];显著高于对照组[(318.66±23.06)对(130.70 4±10.82)nmol/L,P<0.05],至再灌注60min仍高于对照组(P<0.05).偏相关分析显示,[Ca2+];与胰岛素刺激的葡萄糖摄取呈负相关(r=-0.557,P=0.006).结论胰岛素刺激的GLUT4转位障碍和内在活性的降低是模拟缺血再灌注心肌细胞胰岛素抵抗的重要分子机制,[Ca2+];超载可能是缺血再灌注心肌GLUT4内在活性降低的重要原因.
목적 초보탐토결혈재관주후심기세포이도소저항현상적분자궤제.방법 건립성년대서심기세포모의결혈재관주모형,응용동위소시종기술평개심기세포적이도소민감성.Westemblot분석검측심기세포포도당전운단백4(GLUT4)적전위.형광분광광도의측정모의결혈재관주심기세포유리개리자농도([Ca2+]i).결과 모의결혈재관주심기세포표현출명현적이도소저항.이도소자격재관주심기세포적GLUT4전위교대조조명현강저(P<0.05).동시,모의결혈재관주심기세포[Ca2+];현저고우대조조[(318.66±23.06)대(130.70 4±10.82)nmol/L,P<0.05],지재관주60min잉고우대조조(P<0.05).편상관분석현시,[Ca2+];여이도소자격적포도당섭취정부상관(r=-0.557,P=0.006).결론 이도소자격적GLUT4전위장애화내재활성적강저시모의결혈재관주심기세포이도소저항적중요분자궤제,[Ca2+];초재가능시결혈재관주심기GLUT4내재활성강저적중요원인.
Objective Recent.studies have found a strong association of insulin resistance, which might occur during ischemia reperfusion in vitro in the experimental dogs, with disturbed function of cardiomyocytes. Obvious acute insulin resistance, along with glucose dysmetabolism in the reperfused cardiomyocytes, was furher observed in the study performed with ischemia-reperfused ventric- ular myocytes of rats. We tried to investigate preliminarily the molecular mechanisms of insulin resistance in the cardiomyocytes after ischemia reperfusion. Methods An experimental model of insulin-stimulated ischemia reperfusion (SI/R) was created by isolating cardiomyocytes from adult rats. Glucose uptake of the cardiomyoctyes was evaluated with isotope-labeling technique. Glucose trans- porter 4 (GLUT4) translocation induced by insulin was investigated with Western blot analysis, and the intracellular level of free Ca2+ ([Ca2+]I) was measured quantitatively with Ca2+ indicator Fura-2. Results Insulin can stimulated glucose uptake by cardiomyo- cytes, indicating that these cells were insulin-sensitive. Cardiomyocytes were demonstrated notable acute insulin resistmce during reperfusion. Insulin-stimulated GLUT4 translocation in the cardiomyocytes 15 minutes after reperfusion was 72.2% of that in the con- trol group(P<0.05), in which the GLUT4 content in plasma membrane remained unchanged. The finding suggested that a disturbed GLUT4 translocation might happen in the cardiomyocytes during insulin-stimulated ischemia-reperfusion. Calcium overload was identi- fied in the cardiomyocytes with ischemia reperfusion. At 15 minutes of reperfusion, [Ca2+]I was significantly higher in the reperfused cardiomyocytes than that in the control cardiomyocytes[(318.66±23.06)vs(130.70±0.82) nmol/L, P<0.05], and kept at a higher level [(177.79±17.46) nmol/L] at 60 minutes of reperfusion (P<0.05, vs control). Partial correlation analysis revealed a negative correlation of[Ca2+]I with insulin-induced ghcose uptake in the cardiomyoctyes (r = -0.557,P=0.006). Conclusion Disturbed GLUT4 translocation and decreased intrinsic activity may be important molecular mechanisms for the development of insulin resistance in the cardiomyocytes of rat during insulin-simulated ischemia reperfusion,. [Ca2+]I overload may account for the de- creased intrinsic activity d GLUT4.