Mechanism Of Effects Of AGEs,GSK3β And TRPM2 On Osteoblast Proliferation In High Glucose Environment

Objective:Advanced glycation end products(AGEs),glycogen synthase kinase-3(Glycogensynthasekinase-3β,GSK3β),and transient receptor potential M2(TRPM2)have been reported in diabetes and osteoblast proliferation-limited mechanisms.It has an important role,but the research on the relationship between the three is still scarce.In vitro cell experiments were conducted to investigate the effects of AGEs,GSK3β and TRPM2 on the proliferation of osteoblasts in high glucose environment and the mechanisms by which the three affect osteoblast proliferation.Methods:The diabetic environment was simulated by culturing osteoblasts in high glucose medium.Western blot was used to detect differential expression of AGEs,GSK3β and TRPM2 in osteoblasts in high glucose environment,and AGEs inhibitor Aminoguanidine was added to high glucose medium.Aminoguanidine),GSK3β inhibitor AR-A014418,TRPM2 inhibitor Clotrimazole(clotrimazole),MTT assay for osteoblast activity,and western blot to detect the effect of each inhibitor on the expression of the other two substances,under light microscopy The morphology and number of osteoblasts in different inhibitor groups were observed.Results:The morphology and quantity of osteoblasts in different experimental groups were observed under light microscope.The results showed that the osteoblasts in the blank control group grew densely,and the morphology was spindle-shaped,fish-like,and pseudo-foot-like.In the high glucose group(High Glucose,HG group),the number of osteoblasts was significantly reduced,the cells were flat,and there was no obvious protuberance between the cells;while the AGEs inhibitor group,the GSK3β inhibitor group,and the TRPM2 inhibitor constituted the number of bone cells.The high-sugar group increased significantly,and the cells were full,with a small number of protuberances.However,compared with the blank control group,the number of cells and the relationship between the protuberances decreased,and the cell morphology was not full,indicating that the osteoblasts were proliferated in a high glucose environment.AGEs inhibitors,GSK3β inhibitors and TRPM2 inhibitors can partially restore the proliferative activity of osteoblasts.The expressions of AGEs,GSK3β and TRPM2 were detected by immunoblotting.The results showed that the expression of AGEs,GSK3β and TRPM2 in high glucose environment was significantly higher than that in the blank control group.MTT assay was used to detect the proliferation activity of osteoblasts.In the high glucose group,AGEs inhibitor,GSK3β inhibitor and TRPM2 inhibitor were added to detect osteoblast proliferation activity.The results showed that the proliferation activity of osteoblasts decreased in high glucose environment.The proliferation activity of osteoblasts was restored after the addition of AGEs inhibitor,TRPM2 inhibitor and GSK3β inhibitor.The expression of AGEs,GSK3β and TRPM2 in the inhibitor group was detected by immunoblotting.The results showed that AGEs inhibitor can inhibit the expression of TRPM2,GSK3β and P-GSK3β,and GSK3β inhibitor can only inhibit.The expression of TRPM2 did not affect the expression of AGEs.TRPM2 inhibitors did not inhibit the expression of AGEs,GSK3β and P-GSK3β.Conclusion:1.The proliferative activity of osteoblasts is limited in high glucose environment.2.The expression of AGEs,TRPM2 and GSK3β in osteoblasts was significantly increased in high glucose environment,indicating that these three factors play an important role in the proliferative activity of osteoblasts in high glucose environment.3.AGEs,GSK3β,and TRPM2 are involved in the mechanism of osteoblast proliferation inhibition in high glucose environment,which may be achieved through the AGEs-GSK3β-TRPM2 pathway.