Relationship Of Advanced Glycation End Products(AGEs) And Receptor Of Advanced Glycation End Products(RAGE) And The Effect Of High Glucose And AGEs On Vessel Cell

With the extensive research on chronic diabetes complications, the role of advanced glycation end products on these diseases has been attached great importance. At home and abroad, research on the advanced glycation end products (AGEs) and receptor of advanced glycation end products (RAGE) are carried out intensively, especially in diabetes vessel complications. However, the relationship between AGEs accumulation and RAGE expression, as well as the combined damaging effect of high glucose and AGEs on vessel cells has not been reported yet. In this study, the effects of advanced glycation end products(AGEs) and the receptor of advanced glycation end products(RAGE) have been investigated on three levels.(1)In vitro: To supply experimentation evidence for nonenzymatic glycation in vivo, we set up nonenzymatic glycation system in vitro , prepared advanced glycation end products and detected the occur and development of nonenzymatic glycation during the time extending. 50g/l bovine serum albumin(BSA) was incubated with lOOmmol/1 D-glucose at 37℃ for 0, 2, 4, 6, 8, lOand 12 weeks, respectively, and stored in -20℃. At same time, 50g/l bovine serum albumin(BSA) was incubated with 100mmol/l D-glucose at 37℃ for 52 weeks is positive control. After incubation, their results were obtained by brown reaction, fluorescence reaction, fructosamine reaction, enzyme-linked immunosorbent assay(ELISA) and sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE). The results indicate that a positive correlation exists between AGE and the time of incubation.(2) Animal level: To detect the accumulation of advanced glycation end products in mice and rats and investigate the relationship of AGEs and RAGE. Alloxan was used to obtain experimental diabetic mice. We detected the AGEs accumulation in each viscera in mice and rats by using ELISA and anti-AGEs antiserum. And we also detected RAGE mRNA level in each viscera in rats by using RT-PCR. The results indicate that AGEs are distributed in mice widely. Compared with control group, in experimental diabetic mice, the content of accumulated AGEs in several visceras raised evidently. In normal rat, the more RGAE expressed, the fewer the AGEs accumulated, andvice versa.(3) Cell level: To explore the mechanism of advanced glycation end products, high glucose and combined effects of both involved in the process of damaging vessel endothelial cell and vessel smooth muscle cell as well as their influence on the expression of AGEs receptor. The bovine vessel endothelial cell and vessel smooth muscle cell in cell culture were affected by AGEs, glucose and combined of both. The effects on damaging of cells were investigated by MTT, while we detected the contents of LDH, GSH and NO. The expression of RAGE mRNA was analysed by RT-PCR and Cell-ELISA. The results indicate that compared with control group, high glucose group, AGEs group and combined group were found to inhibit the growth of vessel endothelial cell and increase the proliferation of vessel smooth muscle cell significantly. Moreover, they can increase the content of LDH and decrease that of GSH and NO for both cells. In addition, the expression of RAGE was much higher in AGEs group and combined group than those in normal group and high glucose group.Based on the above results, it is concluded that during the process of incubation, cross-link between protein and glucose increase gradually to form AGEs ultimately; AGEs are widely distributed in all visceras of animals, especially in diabetic ones. On the other hand, AGEs can somehow be eliminated given the combination of RGAE and AGEs; AGEs and high glucose have visible effects on damaging vessel cell. And furthermore, AGEs and combined of AGRs and high glucose damage more evidently on vessel cell, which should be mediated by RAGE through certain pathway.