title: Roles of TNF-α, GSK-3β and RANKL in the occurrence and development of diabetic osteoporosis.
abstract: OBJECTIVE
  To investigate the roles of TNF-α, GSK-3β and RANKL in the occurrence and development of diabetic osteoporosis.
  
  
  METHODS
  Diabetic rat model was established; tissue section technology was used to observe the situation of osteoporosis in diabetic rats; rat serum levels of OC, RANKL, GSK-3β, P38mapk, TNF-α and INS were detected by Elisa assay; osteoblasts and osteoclasts were primarily cultured and identified by immunohistochemistry and tartrate-resistant acid phosphatase (TRAP) staining respectively. The effects of GSK-3β inhibitors, lithium chloride, TNF-α antagonists and RANKL antagonists on the proliferation of osteoblasts and osteoclasts were evaluated; quantitative PCR was used to assess the effects of GSK-3β inhibitors, lithium chloride, on TNF-α and RANKL gene expression in osteoblasts and osteoclasts, and the effects of TNF-α and RANKL antagonists on GSK-3β gene expression in osteoblasts and osteoclasts.
  
  
  RESULTS
  Diabetic rat model was successfully established; osteoblasts and osteoclasts were successfully isolated and cultured. Elisa experiments showed that in diabetic model group, the levels of RANKL, GSK-3β, P38mapk and TNF-α were significantly increased, while the levels of osteocalcin (OC) and insulin (INS) were significantly reduced; MTT results showed that osteoclast proliferation in GSK-3β inhibitor and lithium chloride groups were weaker than the untreated group, while osteoclast proliferation in TNF-α antagonist group and RANKL antagonist Group was very close to the untreated group. Osteoblast proliferation in GSK-3β inhibitor and lithium chloride groups were weaker than the untreated group, while osteoblast proliferation in TNF-α antagonist group and RANKL antagonist group was higher than the untreated group. In all of the corresponding groups, cell proliferation in the diabetic group was stronger than the untreated group. In GSK-3β inhibitor and lithium oxide groups, TNF-α and RANKL gene expression levels were elevated, but TNF-α and RANKL gene expression levels in the diabetic group were slightly lower than the control group. GSK-3β gene expression level in TNF-α antagonist group and RANKL antagonist group was reduced; GSK-3β gene expression level in diabetic group was lower than the control group.
  
  
  CONCLUSIONS
  In diabetic rats, TNF-α, GSK-3β and RANKL levels were elevated; GSK-3β could promote the proliferation of osteoblasts and osteoclasts, and inhibit the expression of TNF-α and RANKL; TNF-α and RANKL can suppress the proliferation of osteoblasts while had little effect on osteoclast proliferation; they also can promote the GSK-3β gene expression; interactions between the three broke the balance between osteoblasts and osteoclasts, leading to osteoporosis.
authors.affiliation: The First Affiliated Hospital of Soochow University Suzhou 215006, Jiangsu, China.
authors.forename: Jun
authors.initials: J
authors.lastname: Qi
authors.affiliation: Affiliated Hospital of Nantong University Nantong 226001, Jiangsu, China.
authors.forename: Ke-Su
authors.initials: KS
authors.lastname: Hu
authors.affiliation: The First Affiliated Hospital of Soochow University Suzhou 215006, Jiangsu, China.
authors.forename: Hui-Lin
authors.initials: HL
authors.lastname: Yang
mesh_terms: D000818:Animals; D001842:Bone and Bones / Q000378:metabolism / Q000473:pathology; D002478:Cells, Cultured; D003921:Diabetes Mellitus, Experimental / Q000150:complications* / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism*; D000071679:Glycogen Synthase Kinase 3 beta; D007150:Immunohistochemistry; D008297:Male; D010006:Osteoblasts / Q000378:metabolism; D010010:Osteoclasts / Q000378:metabolism; D010024:Osteoporosis / Q000209:etiology* / Q000378:metabolism* / Q000473:pathology; D053245:RANK Ligand / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*
chemical_list: D053245:RANK Ligand; D014409:Tumor Necrosis Factor-alpha; D000071679:Glycogen Synthase Kinase 3 beta; C000605508:Gsk3b protein, rat; D038362:Glycogen Synthase Kinase 3
keywords: GSK-3β; RANKL; TNF-α; diabetes; osteoporosis