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目的为了探索基质金属蛋白酶9(MMP9)在糖尿病视网膜病变(DR)进展中的潜在调节机制。方法利用Lipofectamine 2000将pc DNA-MMP9质粒和pc DNA-Ang2质粒分别转染至原代大鼠视网膜Müller细胞(RMCs)中。分别利用MTT法与流式细胞仪检测细胞生存率与凋亡情况。同时探索了MMP9与血管紧张素2(Ang2)之间的交互作用。另外,RMCs分别在正常血糖水平与高血糖水平下用MMP9处理2天。此外,通过Western印迹测定细胞凋亡蛋白MMP9、Ang2、Bax2、Bcl2、聚腺苷二磷酸核糖聚合酶(PARP)降解产物、天门冬氨酸特异性半胱氨酸蛋白酶-3(caspase3)降解产物的表达水平。结果与对照组相比,siRNA-MMP9组细胞生存率显著增长而MMP9过表达组下降。与对照组相比,MMP9过表达组中细胞凋亡显著增加,而si RNA-MMP9组中则下降。MMP9表达由Ang2显着调节,而当MMP9表达改变时,Ang2表达无明显变化。再者,高血糖组中MMP9的表达显著性增加,而正常血糖组与对照组比较差异无统计学意义。另外,高血糖组中Bax2,Bcl2,PARP降解产物、caspase3降解产物的表达也显著增加,相应地对照组与正常血糖组则没有显著性改变。结论我们的研究结果表明,MMP9通过由Ang2调控或定位细胞凋亡相关蛋白(如Bax2,Bcl2,PARP降解产物、caspase3降解产物)诱导细胞凋亡,在DR进展中扮演着一个重要角色。
Objective To explore the potential regulatory mechanism of matrix metalloproteinase 9 (MMP9) in the progression of diabetic retinopathy (DR). Methods pcDNA-MMP9 and pcDNA-Ang2 plasmids were transfected into primary rat retinal Müller cells (RMCs) using Lipofectamine 2000 respectively. MTT assay and flow cytometry were used to detect cell viability and apoptosis. The interaction between MMP9 and angiotensin 2 (Ang2) was also explored. In addition, RMCs were treated with MMP9 for 2 days at normal glucose and high blood glucose levels, respectively. In addition, the apoptotic proteins MMP9, Ang2, Bax2, Bcl2, poly-ADP ribose polymerase (PARP) degradation products, aspartate specific caspase 3 degradation products The level of expression. Results Compared with the control group, the survival rate of siRNA-MMP9 group increased significantly while that of MMP9-overexpression group decreased. Compared with the control group, apoptosis in MMP9 overexpression group was significantly increased, while si RNA-MMP9 group was decreased. The expression of MMP9 was significantly regulated by Ang2, while when the expression of MMP9 was changed, there was no significant change of Ang2 expression. In addition, the expression of MMP9 in hyperglycemia group was significantly increased, but there was no significant difference between normal glucose group and control group. In addition, hyperglycemia group Bax2, Bcl2, PARP degradation products, caspase3 degradation products were also significantly increased, corresponding to the control group and normal blood glucose group did not change significantly. Conclusions Our results indicate that MMP9 plays an important role in the progression of DR by regulating Ang-2 or targeting apoptosis-related proteins such as Bax2, Bcl2, PARP degradation products and caspase3 degradation products.