Abstract
Poorly controlled or untreated type I diabetes mellitus is characterized by hyperglycemia and is associated with decreased bone mass and osteoporosis. We have demonstrated that osteoblasts are sensitive to hyperglycemia-associated osmotic stress and respond to elevated extracellular glucose or mannitol by increasing c-jun and collagen I expression. To determine whether MAPKs are involved in this response, MC3T3-E1 osteoblasts were treated with 16.5 mm glucose, mannitol, or contrast dye for 1 h. Immunoblotting of phosphorylated p38 demonstrated activation of p38 MAPK by hyperosmotic stress in vitro and in vivo. Activation peaked at 20 min, remained detectable after 24 h, and was protein kinase C-independent. Activating transcription factor-2 (ATF-2) activation followed the same pattern as phospho-p38. Transactivation of cAMP response element (CRE)- and c-jun promoter (containing a CRE-like element)-reporter constructs increased following hyperosmotic treatment. SB 203580 (a p38 MAPK inhibitor) blocked ATF-2 phosphorylation, CRE transactivation, and c-jun promoter activation. Hyperosmotic activation of collagen I promoter activity was also inhibited by SB 203580, consistent with the involvement of c-jun in collagen I up-regulation. Therefore, we propose that hyperglycemia-induced increases in p38 MAPK activity and ATF-2 phosphorylation contribute to CRE activation and modulation of c-jun and collagen I expression in osteoblasts.
Highlights
(9), and altered cellular metabolism [10]
We have demonstrated that osteoblasts are sensitive to hyperglycemia-associated osmotic stress and respond to elevated extracellular glucose or mannitol by increasing c-jun and collagen I expression
Given the important role of MAPK in the regulation of transcription factor activities and gene expression, we examined the effect of treating osteoblasts with 16.5 mM extracellular sugar on activation of MAPK
Summary
(9), and altered cellular metabolism [10]. These responses are thought to contribute to tissue pathology secondary to diabetes. It is not surprising that expression of a dominantnegative AP-1 fusion protein in osteoblasts suppresses glucose and mannitol effects on gene expression.1 This finding suggests that AP-1 member levels and activity are involved in osteoblast responsiveness to hyperglycemia and osmotic stress. We demonstrate that hyperosmotic stress caused by a physiologically relevant increase in extracellular glucose results in the specific activation of p38 MAPK and its downstream transcription factor, ATF-2. This activation is associated with an increase in CRE and c-jun promoter transactivation. Phosphorylation, CRE and c-jun promoter transactivation, and collagen I transactivation These results demonstrate that p38 MAPK is involved in osmotic stress-induced osteoblast phenotypic changes
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