Inhibition of the Mevalonate Pathway Rescues the Dexamethasone-induced Suppression of the Mineralization in Osteoblasts via Enhancing Bone Morphogenetic Protein-2 Signal

Abstract

We used dexamethasone (DEX)-treated osteoblastic MC3T3-E1 cells, and investigated the effects of an AMP-activated protein kinase activator, 5-aminoimidazole-4-carboxamide-1-β-D-ribonucleoside (AICAR), a Rho-associated protein kinase inhibitor, fasudil hydrochrolide, as well as HMG-CoA reductase inhibitors, simvastatin and pitavastatin, all of which inhibit the mevalonate pathway. DEX (10⁻⁸ M) significantly enhanced mRNA expression of bone morphogenetic protein (BMP)-2 antagonists, follistatin and Dan, and addition of each of 10⁻⁴ M AICAR, 10⁻⁵ M fasudil, 10⁻⁶ M simvastatin, and 10⁻⁶ M pitavastatin significantly reversed the enhancement in mRNA expression of follistatin and Dan and stimulated that of BMP-2 in the cells (p<0.05). DEX (10⁻⁸ M) also significantly suppressed mineralization in the cells, and addition of each of these agents significantly reversed the suppression of mineralization (p<0.05). These findings suggest that the mevalonate pathway was involved in glucocorticoid-induced osteoblast dysfunction, and that its inhibition might promote bone formation through BMP-2 and alleviate glucocorticoid-induced osteoporosis.

glucocorticoid - osteoblast - BMP-2 - mevalonate pathway

References

• 1 van Staa TP, Leufkens HG, Cooper C. The epidemiology of corticosteroid-induced osteoporosis: a meta-analysis.  Osteoporos Int. 2002;  13 777-787
• 2 Chen D, Zhao M, Mundy GR. Bone morphogenetic proteins.  Growth Factors. 2004;  22 233-241
• 3 Jones SE, Jomary C. Secreted Frizzled-related proteins: searching for relationships and patterns.  Bioessays. 2002;  24 811-820
• 4 Canalis E, Economides AN, Gazzerro E. Bone morphogenetic proteins, their antagonists, and the skeleton.  Endocr Rev. 2003;  24 218-235
• 5 Iemura S, Yamamoto TS, Takagi C, Uchiyama H, Natsume T, Shimasaki S, Sugino H, Ueno N. Direct binding of follistatin to a complex of bone-morphogenetic protein and its receptor inhibits ventral and epidermal cell fates in early Xenopus embryo.  Proc Natl Acad Sci USA. 1998;  95 9337-9342
• 6 Ozaki T, Sakiyama S. Molecular cloning and characterization of a cDNA showing negative regulation in v-src-transformed 3Y1 rat fibroblasts.  Proc Natl Acad Sci USA. 1993;  90 2593-2597
• 7 Hsu DR, Economides AN, Wang X, Eimon PM, Harland RM. The Xenopus dorsalizing factor Gremlin identifies a novel family of secreted proteins that antagonize BMP activities.  Mol Cell. 1998;  1 673-683
• 8 Hayashi K, Yamaguchi T, Yano S, Kanazawa I, Yamauchi M, Yamamoto M, Sugimoto T. BMP/Wnt antagonists are upregulated by dexamethasone in osteoblasts and reversed by alendronate and PTH: potential therapeutic targets for glucocorticoid-induced osteoporisis.  Biochem Biophys Res Commun. 2009;  379 261-266
• 9 Garret IR, Gutierrez G, Mundy GR. Statins and bone formation.  Curr Pharm Des. 2001;  7 715-736
• 10 Mundy G, Garrett R, Harris S, Chan J, Chen D, Rossini G, Boyce B, Zhao M, Gutierrez G. Stimulation of bone formation in vitro and in rodents by statins.  Science. 1999;  286 1946-1949
• 11 Maeda T, Matsunuma A, Kawane T, Horiuchi N. Simvastatin promotes osteoblast differentiation and mineralization in MC3T3-E1 cells.  Biochem Biophys Res Commun. 2001;  280 874-877
• 12 Maeda T, Matsunuma A, Kurahashi I, Yanagawa T, Yoshida H, Horiuchi N. Induction of osteoblast differentiation indices by statins in MC3T3-E1 cells.  J Cell Biochem. 2004;  92 458-471
• 13 Kaji H, Naito J, Inoue Y, Sowa H, Sugimoto T, Chihara K. Statin suppresses apoptosis in osteoblastic cells: role of transforming growth factor-beta-Smad3 pathway.  Horm Metab Res. 2008;  40 746-751
• 14 Fujii N, Aschenbach Musi WGN, Hirshman MF, Goodyear LJ. Regulation of glucose transport by the AMP-activated protein kinase.  Proc Nutr Soc. 2004;  63 205-210
• 15 Hardie DG, Carling D. The AMP-activated protein kinase: fuel gauge of the mammalian cell?.  Eur J Biochem. 1997;  246 259-273
• 16 Beg ZH, Allman DW, Gibson DM. Modulation of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity with cAMP and wth protein fractions of rat liver cytosol.  Biochem Biophys Res Commun. 1973;  54 1362-1369
• 17 Goldstein JL, Brown MS. Regulation of the mevalonate pathway.  Nature. 1990;  343 425-430
• 18 Kanazawa I, Yamaguchi T, Yano S, Yamauchi M, Yamamoto M, Sugimoto T. Adiponectin and AMP kinase activator stimulate proliferation, differentiation, and mineralization of osteoblastic MC3T3-E1 cells.  BMC Cell Biol. 2007;  8 51
• 19 Kanazawa I, Yamaguchi T, Yano S, Yamauchi M, Sugimoto T. Activation of AMP-kinase and inhibition of Rho-kinase induce the mineralization of osteoblastic MC3T3-E1 cells through endothelial NOS and BMP-2 expression.  Am J Physiol Endocrinol Metab. 2009;  296 E139-E146
• 20 Sudo H, Kodama HA, Amagai Y, Yamamoto S, Kasai S. In vitro differentiation and calcification in a new clonal osteogenic cell line derived from newborn mouse calvaria.  J Cell Biol. 1983;  96 191-198
• 21 Ogawa N, Yamaguchi T, Yano S, Yamauchi M, Yamamoto M, Sugimoto T. The combination of high glucose and advanced glycation end-products (AGEs) inhibits the mineralization of osteoblastic MC3T3-E1 cells through glucose-induced increase in the receptor for AGEs.  Horm Metab Res. 2007;  39 871-875
• 22 Harris SE, Bonewald LF, Harris MA, Sabatini M, Dallas S, Feng JQ, Ghosh-Choudhury N, Wozney J, Mundy GR. Effects of transforming growth factor beta on bone nodule formation and expression of bone morphogenetic protein 2, osteocalcin, osteopontin, alkaline phosphatase, and type I collagen mRNA in long-term cultures of fetal rat calvarial osteoblasts.  J Bone Miner Res. 1994;  9 855-863
• 23 Shimokawa H. Rho-kinase as a novel therapeutic target in treatment of cardiovascular diseases.  J Cardiovasc Pharmacol. 2002;  39 319-327
• 24 Leclerc N, Luppen CA, Ho VV, Nagpal S, Hacia JG, Smith E, Frenkel B. Gene expression profiling of glucocorticoid-inhibited osteoblasts.  J Mol Endocrinol. 2004;  33 175-193
• 25 Ohnaka K, Shimoda S, Nawata H, Shimokawa H, Kaibuchi K, Iwamoto Y, Takayanagi R. Pitavastatin enhanced BMP-2 and osteocalcin expression by inhibition of Rho-associated kinase in human osteoblasts.  Biochem Biophys Res Commun. 2001;  287 337-342
• 26 Saag KG, Enkey R, Schnitzer TJ, Brown JP, Hawkins F, Goemaere S, Thamsborg G, Liberman UA, Delmas PD, Malice MP, Czachur M, Daifotis AG. Alendronate for the prevention and treatment of glucocorticoid-induced osteoporosis. Glucocorticoid-induced osteoporosis intervention study group.  N Engl J Med. 1998;  339 292-299
• 27 Reid DM, Hughes RA, Laan RF, Sacco-Gibson NA, Wenderoth DH, Adami S, Eusebio RA, Devogelaer JP. Efficacy and safety of daily risedronate in the treatment of corticosteroid-induced osteoporosis in men and women: a randomized trial. European corticosteroid-induced osteoporosis treatment study.  J Bone Miner Res. 2000;  15 1006-1013
• 28 Jilka RL, Weinstein RS, Bellido T, Roberson P, Parfitt AM, Manolagas SC. Increased bone formation by prevention of osteoblast apoptosis with parathyroid hormone.  J Clin Invest. 1999;  104 439-446
• 29 Saag KG, Shane E, Boonen S, Marin F, Donley DW, Taylor KA, Dalsky GP, Marcus R. Teriparatide or alendronate in glucocorticoid-induced osteoporosis.  N Engl J Med. 2007;  357 2028-2039

Correspondence