葛根总黄酮对高糖致血管内皮细胞损伤的保护作用及其机制

目的观察葛根总黄酮对高糖致脐静脉内皮细胞(HUVEC)增殖和细胞周期的影响,探讨葛根总黄酮保护高糖致HUVEC损伤的作用机制。方法采用MTT法观察不同浓度葡萄糖对HUVEC增殖的影响,并观察不同浓度葛根总黄酮对高糖环境下HUVEC增殖及用流式细胞仪检测细胞周期的变化,并测定细胞上清液中SOD活性、NO和ICAM-1的含量。结果30 mmol.L-1葡萄糖可明显抑制HUVEC的增殖反应;降低S期细胞的百分率,增加G0/G1期细胞百分率;并降低HUVEC上清液中SOD活性和NO的含量、升高ICAM-1的含量。葛根总黄酮干预后,随着药物浓度的增加,G0/G1期细胞百分率递减,而S期细胞百分率递增;并可提高细胞上清液中NO水平和SOD活性,降低ICAM-1水平,与30 mmol.L-1葡萄糖比较(P<0.05)。结论葛根总黄酮对高糖致HUVEC损伤具有一定的保护作用,该作用的产生可能与其抑制高浓度葡萄糖作用下HU-VEC的增殖,促使细胞由G0/G1进入S期,使DNA合成增加,使细胞上清液中SOD活性和NO含量增加,而ICAM-1的含量降低等有关。葛根总黄酮对防治糖尿病血管病变的发生发展具有积极的意义。

Preventing role and mechanisms of crude flavones of pueraria loblata on the damage of HUVEC cultured in high glucose medium

Aim To investigate the protecting role and mechanisms of crude flavones of pueraria loblata(FP) on human umbilical vein endothelial cells(HUVEC) in high glucose medium.Methods ①HUVECs were incubated in 20% FCS RPMI-1640 culture medium containing 0,5,10,30,60,90,120 mmol·L~(-1) glucose for 48 hours.The proliferation of HUVEC was analyzed with MTT method(490 nm).② HUVECs were incubated in 20% FCS RPMI-1640 culture medium containing 0 or 30 mmol·L~(-1) glucose,either alone or in the presence of FP(final concentration 10~(-3),10~(-4),10~(-5),10~(-6) mol·L~(-1) respectively) for 48 hours.At the same time,30 mmol·L~(-1) mannitol was added.The proliferation of HUVEC was analyzed using MTT method.The cell cycle of HUVEC was analyzed using flowcytometry.The contents of SOD,NO and ICAM-1 in HUVEC supernatant were measured using test kits.Results HUVEC in 5 mmol·L~(-1) glucose medium proliferated well,but the proliferation of HUVEC in 10,30,60,90 and 120 mmol·L~(-1) glucose media was disturbed,showing a negative correlation.Proliferation of HUVEC was inhibited in 30 mmol·L~(-1) glucose,compared with that in 0 mmol·L~(-1) glucose and 30 mmol·L~(-1) mannitol(P<0.01),FP(10~(-3) mol·L~(-1)).Proliferation of HUVEC in 0 mmol·L~(-1) glucose did not show fundamental FP(10~(-3),10~(-4) mol) mostly reversed the decreased proliferation of HUVEC in 30 mmol·L~(-1) glucose(P<0.01).Cell cycle analysis showed that cell percentage of G_0/G_1 phase increased,while that of S phase decreased in 30 mmol·L~(-1) glucose.But to the cells cultured with FP,the percentage of G_0/G_1 phase decreased,while that of S phase increased.The contents of SOD and NO in 30 mmol·L~(-1) glucose decreased,while ICAM-1 increased significantly(P<0.01).SOD,NO in the supernatant of that with FP or aminoguanidine increased and the level of ICAM-1 decreased with 30 mmol·L~(-1) glucose(P<0.05).Conclusion The results suggest that FP may prevent the cultured HUVEC from lesion and inhibit proliferation caused by high glucose.The mechanism of FP of protecting the HUVEC in high glucose is related with increasing the activity of SOD,restoring the equilibration of NO and ET and decreasing the level of ICAM-1.It is beneficial to preventing and curing diabetic vascular defects.