高糖及不同幅度高糖波动下牛视网膜毛细血管周细胞增生与凋亡的研究

Proliferation and apoptosis of cultured bovine retinal capillary pericytes in high glucose and high glucose fluctuation at different amplitude

目的分析不同浓度葡萄糖培养下的牛视网膜毛细血管周细胞增生、细胞周期改变及凋亡与葡萄糖浓度及高糖波动幅度的相关性,并对比同一水平下恒定高糖与高糖波动对周细胞作用的强度。方法以体外培养3代近融合的牛视网膜毛细血管周细胞为对象,分别在不同浓度高糖(15mmol.L-1、25mmol.L-1、35mmol.L-1)、不同幅度的高糖波动(5.5/15mmol.L-1、5.5/25mmol.L-1、5.5/35mmol.L-1)及对照组(5.5mmol.L-1)下孵育7d,采用MTT比色法观察周细胞增生情况;流式细胞术观察周细胞细胞周期变化情况;TUNEL法检测培养后周细胞的凋亡率。结果(1)高糖及高糖波动抑制周细胞增生,与葡萄糖浓度及高糖波动幅度呈正相关(P<0.01),同一水平下持续高糖组作用强于高糖波动组(P<0.05);(2)高糖及高糖波动阻滞周细胞周期,使G0/G1期细胞比例增加,S期细胞比例减少,G2/M细胞比例无明显改变;(3)高糖及高糖波动诱导周细胞凋亡,高糖各组凋亡率分别为23.78%、37.22%、48.73%,而高糖波动组分别为15.53%、28.73%、39.14%,与对照组的4.25%相比差异有显著性意义(P<0.01)。且周细胞凋亡率与高糖的浓度呈正相关(P<0.01);与高糖波动幅度呈正相关(P<0.01);同一水平下持续高糖组作用强于高糖波动组(P<0.05)。结论同一水平下恒定高糖较高糖波动可能具有更强的周细胞增生抑制、周期阻滞及诱导凋亡作用,这些作用可能与葡萄糖浓度及波动的幅度正相关。

ObjectJve To analyze the correlation of the proliferation, cell cycle change and apoptosis of preliminary cultured bovine retinal capillary pericytes in different concentrations of glucose and the concentration of glucose and the amplitude of glucose fluctuation, and to observe the strength of action in pericytes between high glucose and high glucose fluctuation in the same level. Methods Pericytes were cultured for 7 days respectively under different concentration glucose（ 15 mmol·L^-1] ,25 mmol ·L^-1 and 35 mmol ·L^-1 ） ,high glucose fluctuation of different amplitude （5.5/15 mmol ·L^-1,5.5/25 mmol ·L^-1 and 5.5/35 mmol ·L^-1 ） and control group（ 5.5 mmol ·L^-1）. The proliferation of pericytes were assayed with MTF assay ;The cell cycles of pericytes change were amalyzed by flow cytometry; The apoptosis of pericytes were studied by TUNEL method. Results （ 1 ） Both high glucose and high glucose fluctuation inhibited the proliferation of pericytes, the changes were correlated positively with the concentrations of glucose（P 〈 0.01 ） and the amplitude of fluctuation （ P 〈 0.01 ）. （2） Both high glucose and high glucose fluctuation blocked cell cycle ,increased the percentage of G0/G1 phase and decreased the percentage of S phase. But there was not any obvious change in the percentage of G2/M phase in each group. （3） Both high glucose and high glucose fluctuation induced cell apoptosis,the apoptotic rates of high glgh groups were 23.78% ,37.22% and 48. 73% respectively,and that of high glucose fluctuation groups were 15.53% ,28.73% and 39. 14% respectively. It was significantly difference compared with the 4. 25% of the control group. The changes were correlated positively with the concentrations of glucose （P 〈 0.01 ） and the amplitude of fluctuation （ P 〈 0.01 ） ; The changes were much more significant in the constant high glucose compared with the high glucose fluctuation in the same level （ P 〈0. 05 ）. Conclusion The constant high glucose may be more obviously than high glucose fluctuation to inhibit the proliferation of pericytes,block the cell cycle and induce apoptosis. All the changes may be correlated positively with the concentrations of glucose and the amplitude of fluctuation. [ Rec Adv Ophthaimol 2007 ;27（2） ：87-90 ]