Bone Abstracts

Type 2 diabetes mellitus impairs bone quality and increases fracture risk. We showed that diabetic ZDF rats have low bone mass due to impaired osteoblastogenesis, which can be partially reversed with an intermittent parathyroid hormone 1–84 (PTH) therapy. It remains unclear, why PTH treatment does not fully restore osteoblast (OB) function in diabetic conditions. Here, we tested if high glucose (HG) conditions lead to a partial PTH resistance in osteoblasts. Pre-osteoblastic MC3T3-E1 cells were cultured in osteogenic medium in HG concentrations for 21d. HG decreased the production of mineralized matrix by −69% as compared to control cells. Intermittent PTH treatment reversed this effect (+115%), but did not reach the level of control cells. PTH dose-dependently increased cAMP levels up to 40-fold, whereas this increase was blunted by HG. Similarly, PTH-induced activation of protein kinase A, the downstream target of cAMP, was decreased in HG conditions. Gene expression of IGF1, alkaline phosphatase, osteopontin, connexin43 and RANKL significantly increased after intermittent treatment of PTH in control and HG conditions. To determine whether the blunted cAMP activation was due to a decreased expression of the PTH receptor 1 (PTHR1), we determined protein levels of PTHR1 using western blot. However, PTHR1 expression was not affected by HG or PTH. Instead N-linked glycosylation was increased in HG. Combined treatment of PTH with tunicamycin, blocking N-linked glycoprotein synthesis, elevated the amount of mineralization in HG treated cells similarly to PTH treatment (fourfold). Interestingly, tunicamycin alone was able to reverse the inhibition of mineralization of HG treated OB. These results suggest that increased N-linked glycosylation is a critical mechanism of OB function in HG conditions in MC3T3-E1 cells. Thus, preventing these posttranslational modifications may improve OB function and augment the response to PTH treatment in type 2 diabetes mellitus.