Effect And Mechanism Of RKIP/p38-MAPK Signal Pathway On DR

ObjectiveRaf-1 kinase inhibitor protein(RKIP)can inhibit the expression and activation of p38 mitogen-activated protein kinase(p38-MAPK).However,the effect of RKIP / p38-MAPK signaling pathway on retinal ganglion cell(RGC)in DR state is not clear.This study intends to explore the effect and mechanism of this pathway on RGC in DR state.MethodsForty-eight male Sprague-Dawley rats were randomly divided into the control group,the diabetic group,the negative treatment group,and the RKIP group.Except for the control group,the other groups were intraperitoneally injected with streptozotocin(60mg/kg)to prepare a diabetes model.Established diabetes model 2 weeks,the RKIP gene fragment recombinant lentivirus(LV-RKIP)and recombinant lentivirus carrying a nonsense nucleotide sequence were injected into the vitreous body of the RKIP group and the negative treatment group,respectively.After 12 weeks,Western blot analysis RKIP,p38-MAPK and caspase-3 expression in the retina;Immunofluorescence assay for expression of glial fibrillary acidic protein(GFAP),RKIP,p38-MAPK,glutamine synthetase(GS)and glial glutamate/L-aspartate transporter(GLAST)in retinal Müller cells;Glutamate content detection kit detects retinal glutamate content;HE staining was used to detect the density of RGC.ResultsThe immunofluorescence results showed that the expression of GFAP in the diabetes group and the negative treatment group was significantly increased compared with the control group(P<0.01);while in the RKIP group,the GFAP decreased significantly compared with the diabetic group(P<0.01).Compared with the control group,the expression of RKIP in the diabetic group and the negative treatment group was significantly reduced(P<0.01);while the expression of RKIP was significantly increased in the RKIP group compared with the diabetic group(P<0.05).Compared with the control group,the expression of p38-MAPK in the diabetic group and the negative treatment group was significantly increased(P<0.05);the expression of p38-MAPK was significantly reduced in the RKIP group compared with the diabetes group(P<0.01).Compared with the control group,the expression of GS in the diabetic group and the negative treatment group was significantly reduced(P<0.01);the expression of GS was significantly increased in the RKIP group compared with the diabetic group(P<0.01).Compared with the control group,the expression of GLAST in the diabetic group and the negative treatment group was significantly reduced(P<0.01);the expression of GLAST was significantly increased in the RKIP group compared with the diabetic group(P<0.01).The above results indicate that upregulation of RKIP reduces the activation of retinal Müller cells.Further analysis of the degree of retinal damage,Western blot results showed that the diabetic group and the negative treatment group compared with the control group,caspase-3 and p38-MAPK expression in the retina increased(P<0.05),RKIP expression was significantly reduced(P<0.01);compared with the diabetic group,the expression of caspase-3 and p38-MAPK decreased in the RKIP group(P<0.01),and the expression of RKIP increased significantly(P<0.05).At the same time,the glutamate content of retinal synapses showed that compared with the control group,the content of synaptic glutamate in the diabetic group and the negative treatment group was significantly increased(P<0.05);the content of synaptic glutamate in the RKIP group was reduced compared with the diabetic group(P<0.01).Further analysis of retinal RGC damage,HE staining showed that the number of retinal RGC decreased in the diabetic group and the negative treatment group compared with the control group(P<0.01);the number of retinal RGC increased in the RKIP group compared with the diabetes group(P<0.05).ConclusionsRKIP can inhibit the p38-MAPK signal pathway,restore the expression of GLAST and GS in diabetic retina Müller cells,inhibit the activation of glial cells,and reduce the loss of RGC.