Effects Of Glimepiride On Rat Osteoblasts Cultured In Hyperglycemic Conditions: The Role Of The PI3-kinase/Akt Pathway

Pathologic changes induced by hyperglycemia in diabetic patients have been considered to contribute to implant failures.Therefore,it plays a key role to reduce the retarding effects of diabetes on bone healing through diabetic control for the placement of implants for those diabetic patients.Liu Hongchen raised a new medicine administration of dental implant.We hope that using new medicine administration could accelerate bone regeneration and implant osseointegration. Accordingly,it is very necessary to investigate the effects of widely used anti-diabetic drugs on the partial bone cells to the implants.Glimepiride is a third-generation sulfonylurea agent and can stimulate insulin release from pancreaticβ-cells.In addition to the stimulatory effects on pancreatic insulin secretion,glimepiride has also been shown to play extrapancreatic roles in tyrosine phosphorylation of insulin receptor substrate-1/2(IRS-1/2) and activation of the PI3K/Akt pathway in rat adipocytes and skeletal muscle.Moreover,in endothelial cells,it is suggested that glimepiride induces eNOS phosphorylation with a dependent mechanism PI3K/Akt.The eNOS isoform seems to play a key role in regulating osteoblast activity and bone formation since eNOS knockout mice have osteoporosis due to defective bone formation.This study investigated the actions of glimepiride on primary rat mandibular osteoblasts cultured in two concentrations of glucose and the role of PI3K/Akt pathway in the process.It was expected to provide the experimental evidence for anti-diabetic drug administration of dental implant.Partâ… Effects of glimepiride on proliferation,differentiation and mineralization of rat mandibular osteoblasts in hyperglycemiaObjective:To explore the effect of hyperglycemia and glimepiride on proliferation,differentiation and mineralization of rat mandibular osteoblasts to verify the hypothesis of dental implant administration.Methods:Primary osteoblasts were isolated and cultured.Then the cells were placed in an osteogenic medium,containing different containing two concentrations of glucose(5.5mM and 16.5mM),with or without glimepiride (10μM).Cell proliferation was valuated through MTT assay,Alkaline phosphatase (ALP) activity was determined by biochemistry method,Colâ… protein levels were determined by Western blot analysis,OCN mRNA levels were tested by RT-PCR.Results:Hyperglycemic conditions interfered with the proliferation,ALP activity,OCN mRNA expression of rat osteoblasts,but improved the expression of Colâ… on 14 day.Glimepiride stimulated rat osteoblast proliferation,ALP activity and OCN mRNA expresson,the addition of glimepiride to normoglycemic(5.5 mM) cultures registered a significant increase Colâ… expression for 7d and 14d.We also noticed that glimepiride markedly increased Colâ… expression in cell cultured with 16.5 mM glucose for 7d,but failed to increase for14d.Conclusions:Hyperglycemic conditions interfered with the proliferation, differentiation and mineralization of osteoblasts in rats,however,glimepiride improved the proliferation,differentiation and mineralization of osteoblasts in rats.Partâ…¡Effect of Glimepiride on glucose uptake of rat mandibular osteoblasts in hyperglycemiaObjective:To explore the effect of glimepiride on glucose uptake,GLUT-1 and GLUT-3 expression of rat mandibular osteoblasts in hyperglycemia.Methods:Primary osteoblasts were isolated and cultured.Then the cells were placed in an osteogenic medium,containing different containing two concentrations of glucose(5.5mM and 16.5mM),with or without glimepiride (10μM).Glucose uptake was detected by using ¹⁸F-FDG in the cells and GLUT-1, GLUT-3 expression levels were evaluated by Western blot analysis.Results:The 16.5mM of glucose significantly inhibited 18F-FDG uptake and downregulated GLUT-3 protein expression in osteoblasts.Hyperglycemia increased GLUT-1 protein expression.The addition of glimepiride to the culture medium containing 5.5mM and 16.5mM glucose concentrations marked increased glucose uptake and GLUT-1,3 expressions.Conclusions:Hyperglycemia inhibited the glucose uptake in osteoblasts and expression of GLUT-3 Protein,however,enhanced the GLUT-1 protein expression.The addition of glimepiride to the culture medium containing 5.5mM and 16.5mM glucose concentrations caused a marked increase in glucose uptake and GLUT-1,3 protein expressions.Partâ…¢the role of PI3K/Akt pathway in glimepiride-induced rat osteoblasts cultured in hyperglycemiaObjective:To observe the role of PI3K/Akt pathway in glimepiride-induced rat osteoblastic proliferation,differentiation,and glucose transport.Methods:Primary osteoblasts were isolated and cultured.Then the cells were placed in an osteogenic medium,containing different containing two concentrations of glucose(5.5mM and 16.5mM),with or without glimepiride (10μM) or LY294002(10μM).Cell proliferation was valuated through MTT assay, Alkaline phosphatase(ALP) activity was determined by biochemistry method, Col,PI3K P85,Akt/P-Akt,IRS-1/P-IRS-1,and GLUT-1,3 protein levels were determined by Western blot analysis,glucose uptake was detected by using ¹⁸F-FDG.Results:Osteoblastic cultures pretreated with the PI3K inhibitor inhibited both glimepiride-induced cell proliferation,ALP activity,Colâ… production,glucose uptake,and GLUT-1,3 protein expressions.Further,glimepiride can stimulate PI3K P85,Akt/P-Akt,and IRS-1/P-IRS-1 protein expression.Conclusion:glimepiride-induced cell proliferation,ALP activity,typeâ… collagen production,glucose uptake,and GLUT-1,3 protein expressions were mediated by PI3K/Akt signaling pathway.Partâ…£Glimepiride induces eNOS expression in rat osteoblasts via a PI3K/Akt dependent pathwayObjective:To explore the effect of glimepifide on eNOS expression in rat osteoblasts and its associated PI3K/Akt pathwayMethods:Primary osteoblasts were isolated and cultured.Then the cells were placed in an osteogenic medium,containing different containing two concentrations of glucose(5.5mM and 16.5mM).Osteoblasts were treated with glimepiride(10μM) for 120min in the absence or in the presence of LY294002. Akt/P-Akt and eNOS/P-eNOS protein levels were determined by Western blot analysisResults:Glimepiride activates eNOS expression in rat osteoblasts cultured with two concentrations of glucose.PI3K/Akt pathway plays a key role in the precessionConclusion:Glimepiride activates eNOS expression in rat osteoblasts cultured with two concentrations of glucose via PI3K/Akt pathway.