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目的观察糖尿病早期视网膜中蛋白激酶C(PKC)、内皮素(ET)系统与核因子-κB(NF-κB)的表达变化,并探讨PKC抑制剂对上述因子的影响。方法将SD大鼠分为正常对照(NC)组与糖尿病(DM)组,STZ诱导糖尿病大鼠模型。ELISA及Western blot检测视网膜PKC活性及亚型的表达,QRT-PCR检测不同时间点视网膜ETs和NF-κB p65基因表达及12周时PKC抑制剂对上述基因的影响。结果 12周时,DM组视网膜组织细胞膜PKC活性较NC组增加(t=6.36,P=0.00),细胞浆PKC活性无明显改变;PKC-α、PKC-β_1、PKC-β_2、PKC-δ蛋白表达均较NC组增多(t=5.69、4.71、4.10、3.753,P<0.05),以PKC-β_2增加最明显,PKC-ε无改变;ETs及NF-κB p65 mRNA在不同时间点出现表达水平的上调,其中,ET-1及NF-κB出现最早;PKC抑制剂使视网膜ETs及NF-κB p65 mRNA表达呈浓度依赖性下降。结论 ETs及NF-κB的异常激活可能是PKC影响DR发生发展的作用机制之一。
Objective To observe the expression changes of protein kinase C (PKC), endothelin (ET) and nuclear factor-κB (NF-κB) in the early retina of diabetic patients and explore the effect of PKC inhibitors on these factors. Methods SD rats were divided into normal control (NC) group and diabetic (DM) group, STZ-induced diabetic rats. ELISA and Western blot were used to detect the retinal PKC activity and subtype expression. The expression of ET-1 and NF-κB p65 at different time points was detected by QRT-PCR and the effect of PKC inhibitor on the above-mentioned genes at 12 weeks. Results After 12 weeks, the activity of PKC in the membrane of retinal tissue in DM group was higher than that in NC group (t = 6.36, P = 0.00), while the activity of PKC-β, PKC- (T = 5.69,4.71,4.10,3.753, P <0.05). The expression of PKC-β_2 was the most obvious and the expression of PKC-ε was unchanged. The expression levels of ETs and NF-κB p65 mRNA at different time points , And ET-1 and NF-κB were the earliest. PKC inhibitor decreased the expression of ETs and NF-κB p65 mRNA in a concentration-dependent manner. Conclusion The abnormal activation of ETs and NF-κB may be one of the mechanisms by which PKC affects the occurrence and development of DR.