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目的研究替米沙坦对高糖高胰岛素共同诱导的心肌成纤维细胞增殖的影响及其机制。方法培养Spargue Dawley(SD)大鼠乳鼠心肌成纤维细胞,随机分为对照组(5.5 mmol/L葡萄糖)、高糖高胰岛素组(25 mmol/L葡萄糖+1×10-7mol/L胰岛素)、细胞外调节蛋白激酶(ERK)抑制剂组(高糖高胰岛素+10μmol/L PD98059)和替米沙坦组(高糖高胰岛素+10μmol/L替米沙坦),培养48 h后,CCK8法检测细胞增殖;结晶紫染色法测定细胞数量;[3H]thymidine标记法测定细胞DNA合成;流式细胞仪分析细胞周期;ELISA试剂盒测定上清液中血管紧张素(Ang)Ⅱ、醛固酮(ALD)含量;Western blot测定磷酸化ERK1/2(p-ERK1/2)蛋白表达量。结果与对照组相比,高糖高胰岛素组细胞增殖(1.43±0.12 vs.0.50±0.10)、细胞计数(1.64±0.03 vs.1.06±0.02)、DNA合成量(26135.47±324.86 vs.14528.26±267.18)、S+G2+M百分比[(19.33±0.12)%vs.(8.56±0.07)%]、AngⅡ[(30.58±7.26)pg/ml vs.(18.26±2.64)pg/ml]与ALD含量[(19.62±1.25)pg/ml vs.(12.83±1.29)pg/ml]及p-ERK1/2蛋白表达量明显升高(P<0.05);与高糖高胰岛素组相比,ERK抑制剂组细胞增殖(0.72±0.06 vs.1.33±0.12)、细胞计数(1.35±0.01 vs.1.64±0.03)、DNA合成量(18643.76±192.52 vs.26135.47±324.86)、S+G2+M百分比[(12.84±0.36)%vs.(19.33±0.12)%]、AngⅡ[(23.17±5.31)pg/ml vs.(30.58±7.26)pg/ml]与ALD[(15.27±1.13)pg/ml vs.(19.62±1.25)pg/ml]含量及p-ERK1/2蛋白表达量明显降低(P<0.05);与高糖高胰岛素组相比,替米沙坦组细胞增殖(0.94±0.05 vs.1.33±0.12)、细胞计数(1.49±0.02 vs.1.64±0.03)、DNA合成量(21829.35±154.73 vs.26135.47±324.86)、S+G2+M百分比[(15.13±0.42)%vs.(19.33±0.12)%、AngⅡ[(26.84±4.36)pg/ml vs.(30.58±7.26)pg/ml]与ALD[(17.81±1.53)pg/ml vs.(19.62±1.25)pg/ml]含量及p-ERK1/2蛋白表达量明显降低(P<0.05);ERK抑制剂组与替米沙坦组各指标间没有统计学差异。结论血管紧张素1型受体AT1R阻滞剂替米沙坦通过部分抑制ERK1/2信号通路改善高糖高胰岛素共同诱导的心肌成纤维细胞增殖。
Objective To study the effect of telmisartan on the proliferation of cardiac fibroblasts induced by high glucose and high insulin and its mechanism. Methods Fibroblasts of Spargue Dawley rats were randomly divided into control group (5.5 mmol / L glucose), high glucose and high insulin group (25 mmol / L glucose + 1 × 10-7 mol / L insulin) , ERK inhibitor group (high glucose and high insulin + 10μmol / L PD98059) and telmisartan group (high glucose and high insulin + 10μmol / L telmisartan). After cultured for 48 h, CCK8 Cell viability was measured by crystal violet staining. [3H] thymidine labeling was used to determine the DNA synthesis. Cell cycle was analyzed by flow cytometry. Angiotensin II (Ang II), aldosterone ALD). The protein expression of phosphorylated ERK1 / 2 (p-ERK1 / 2) was detected by Western blot. Results Compared with the control group, the cell proliferation in high glucose and high insulin group (1.43 ± 0.12 vs.0.50 ± 0.10), cell count (1.64 ± 0.03 vs.1.06 ± 0.02), DNA synthesis (26135.47 ± 324.86 vs.14528.26 ± 267.18 (19.33 ± 0.12)% vs (8.56 ± 0.07)%], AngⅡ [(30.58 ± 7.26) pg / ml vs. (18.26 ± 2.64) pg / ml) (19.62 ± 1.25) pg / ml vs. (12.83 ± 1.29) pg / ml] and p-ERK1 / 2 protein expression (P <0.05). Compared with high glucose and high insulin group, ERK inhibitor group Cell proliferation (0.72 ± 0.06 vs. 1.33 ± 0.12), cell count (1.35 ± 0.01 vs.1.64 ± 0.03), DNA synthesis (18643.76 ± 192.52 vs.26135.47 ± 324.86), S + G2 + M percentage (19.33 ± 0.12)%], AngⅡ [(23.17 ± 5.31) pg / ml vs. (30.58 ± 7.26) pg / ml] and ALD [(15.27 ± 1.13) pg / ml vs. (19.62 ± 1.25) pg / ml] and the expression of p-ERK1 / 2 were significantly decreased (P <0.05). Compared with high glucose and high insulin group, the proliferation of cells in telmisartan group (0.94 ± 0.05 vs.1.33 ± 0.12) , Cell count (1.49 ± 0.02 vs 1.64 ± 0.03), DNA synthesis (21829.35 ± 154.73 vs.26135.47 ± 324.86), S + G2 + M percentage [(15.13 ± 0.42)% vs (19.33 ± 0.12)%, Ang II [( 26.84 ± 4.36 pg / ml vs. 30.58 ± 7.26 pg / ml] and ALD [(17.81 ± 1.53) pg / ml vs. (19.62 ± 1.25) pg / ml] and p-ERK1 / 2 protein expression (P <0.05). There was no significant difference between each index of ERK inhibitor group and telmisartan group. Conclusion Telmisartan, an angiotensin 1 type 1 receptor antagonist, ameliorates cardiac fibroblasts proliferation induced by high glucose and high insulin by partially inhibiting the ERK1 / 2 signaling pathway.