The Correlation Research Of Protein Kinase C And Endothelin System, Nuclear Factor-kB In Retina Of Early Diabetic Rats

Objective: To investigate the activity and isoforms protein expression of protein kinase C(PKC) in the retina in early diabetes. And to continuous dynamic observe the m RNA expression of the endothelin(ET) systemand nuclear factor B-Kappa(NF-k B) in retina in diabetes. To observe the effect of pan PKC inhibitor, GF103209 X, on the m RNA expression of ETs and NF-k B in the retina.To explore the possible mechanism of PKC induced retinal cell injury in retina in early diabetic.Method: Sprague Dawley(SD) were used as experimental subjects. Diabetic animal model was prepared by peritoneally injecting STZ. The PKC activity in the retina of 12-weekdiabetic rats was measured by enzyme-linked immunosorbent assay(ELISA). The protein expression of PKC imforms in the retina of 12-weekdiabetic rats was detected by western blotting(WB). Quantitative Real-time polymerase chain reaction(QRT-PCR) was used to observe the m RNA expression of ETs and NF-k B p65 in the retina atdifferent duration of diabetes. QRT-PCR was also used to detect the alteration of the cell factors mentioned above after intravitreal injection of a general PKC inhibitor in 12-week diabetic rats in different concentration(10-5mol/L, 10-6mol/L, 10-7mol/L).Results: 1. At the 12-week diabetic rat retina, PKC activity was significantly increased in the membranous fraction compared to the normal rats, the difference was statistically significant(P<0.05), whereas PKC activity in the cytosolic fraction were incr eased without statistically significant(P>0.05). 2. At the 12-week diabetic rat retina, the expression of PKC-α,-β1,-β2 and-δ were increased compared with the normal rats, the difference was statistically significant(P<0.05), especially the PKC-β2 isoform, whereas the PKC-ε was unchanged (P>0.05). 3. Within the retina of early diabetic rats, the m RNA expression of ETs and NF-k B were significantly increased compared with the normal rats, the ET-1 and NF-k B p65 had the earliest change which were found after 2 weeks, and the rise of ET-3, ET-A were found after 4 weeks, and ET-B was the latest which was detected after 12 weeks, the differences were statistically significant(P<0.01). 4. After intravitreal injection of GF109203X(10-5, 10-6,10-7M) into the retina of 12-week diabetic rats, the m RNA expression of ETs and NF-k B in the retina had a different degrees decease, and the reduction was in a dose-dependent manner,the differences were statistically significant(P<0.01).Conclusion: 1. The increase of PKC activation and the expression of ETs and NF-k B had emerged in the retina of early diabetic rats. 2. PKC inhibitor, GF103209 X, can significantly make a different degree reduction of the expression of ETs and NF-k B in a dose-dependent way. 3. In the early stage of diabetes, PKC may induce cell injury by destroying the balance of ETs and NF-k B in the retina.