Glucocorticoid (GC) induced osteoporosis (GIO) is caused by the long-term use of GC for treatment of autoimmune and inflammatory diseases. The GC related disruption of bone marrow microcirculation and increased adipogenesis contribute to GIO development. However, neither currently available anti-osteoporosis agent is completely addressed to microcirculation and bone marrow adipogenesis. Salvianolic acid B (Sal B) is a polyphenolic compound from a Chinese herbal medicine, Salvia miltiorrhiza Bunge. The aim of this study was to determine the effects of Sal B on osteoblast bone formation, angiogenesis and adipogenesis-associated GIO by performing marrow adipogenesis and microcirculation dilation and bone histomorphometry analyses. (1) In vivo study: Bone loss in GC treated rats was confirmed by significantly decreased BMD, bone strength, cancellous bone mass and architecture, osteoblast distribution, bone formation, marrow microvessel density and diameter along with down-regulation of marrow BMPs expression and increased adipogenesis. Daily treatment with Sal B (40 mg/kg/d) for 12 weeks in GC male rats prevented GC-induced cancellous bone loss and increased adipogenesis while increasing cancellous bone formation rate with improved local microcirculation by capillary dilation. Treatment with Sal B at a higher dose (80 mg/kg/d) not only prevented GC-induced osteopenia, but also increased cancellous bone mass and thickness, associated with increase of marrow BMPs expression, inhibited adipogenesis and further increased microvessel diameters. (2) In vitro study: In concentration from 10？6 mol/L to 10？7 mol/L, Sal B stimulated bone marrow stromal cell (MSC) differentiation to osteoblast and increased osteoblast activities, decreased GC associated adipogenic differentiation by down-regulation of PPARγ mRNA expression, increased Runx2 mRNA expression without osteoblast inducement, and, furthermore, Sal B decreased Dickkopf-1 and increased β-catenin mRNA expression with or without adipocyte inducement in MSC. We conclude that Sal B prevented bone loss in GC-treated rats through stimulation of osteogenesis, bone marrow angiogenesis and inhibition of adipogenesis.
References
[1]
Shah SK, Gecys GT (2006) Prednisone-Induced Osteoporosis: An Overlooked and Undertreated Adverse Effect. J Am Osteopath Assoc 106: 653–657.
[2]
Canalis E (2005) Mechanisms of glucocorticoid action in bone. Curr Osteoporos Rep 3: 98–102.
[3]
Ito S, Suzuki N, Kato S (2007) Glucocorticoids induce the differentiation of a mesenchymal progenitor cell line, ROB-C26 into adipocytes and osteoblasts, but fail to induce terminal osteoblast differentiation. Bone 40: 84–92.
[4]
Weinstein RS (2010) Glucocorticoids, osteocytes, and skeletal fragility: The role of bone vascularity. Bone 46: 564–570.
[5]
Kerachian MA, Séguin C, Harvey EJ (2009) Glucocorticoids in osteonecrosis of the femoral head: a new understanding of the mechanisms of action. J Steroid Biochem Mol Biol 114: 121–128.
[6]
Khan E, AbuAmer Y (2003) Activation of peroxisome proliferator 2 activated receptor 2 gamma inhibits differentiation of preosteoblasts. J Lab Clin Med 142: 292–234.
[7]
Luppen CA, Chandler RL, Noh T, Mortlock DP, Frenkel B (2008) BMP-2 vs. BMP-4 expression and activity in glucocorticoid-arrested MC3T3-E1 osteoblasts: Smad signaling, not alkaline phosphatase activity, predicts rescue of mineralization. Growth Factors 26: 226–237.
[8]
Yin L, Li YB, Wang YS (2006) Dexamethasone-induced adipogenesis in primary marrow stromal cell culture: mechanism of steroid-induced osteonecrosis. Clin Med J (Engl.) 119: 581–588.
[9]
Kitajima M, Shigematsu M, Ogawa K, Sugihara H, Hotokebuchi T (2007) Effects of glucocorticoid on adipocyte size in human bone marrow. Med Mol Morphol 40: 150–156.
[10]
Miyanishi K, Yamamoto T, Irisa T, Yamashita A, Jingushi S (2002) Bone marrow fat cell enlargement and a rise in intraosseous pressure in steroid-treated rabbits with osteonecrosis. Bone 30: 185–190.
[11]
Ersoy A, Kahvecioglu S, Ersoy C, Akdag I, Yurtsever I, et al. (2006) Is glucocorticoid-induced osteonecrosis after kidney transplantation related to osteoporosis? Nephrol Dial Transplant 21: 1452–1453.
[12]
Marx N, Schonbeck U, Lazar MA (1998) PPAR2 gamma activators inhibit gene expression and migration in human vascular smooth muscle. J Circ Res 83: 1097–l103.
[13]
Adams JD, Wang R, Yang J, Lien EJ (2006) Preclinical and clinical examinations of Salvia miltiorrhiza and its tanshinones in ischemic conditions. Chin Med 1: 3–7.
[14]
Li YG, Song L, Liu M, Hu ZB, Wang ZT (2009) Advancement in analysis of Salviae miltiorrhizae Radix et Rhizoma (Danshen). J Chromatogr A 1216: 1941–1953.
[15]
Ji XY, Tan BKH, Zhu YZ (2000) Salvia miltiorrhiza and ischemic diseases. Acta Pharmacol Sin 21: 1089–1094.
[16]
He HB, Yang XZ, Shi MQ, Zeng XW, Wu LM, et al. (2008) Comparison of cardioprotective effects of salvianolic acid B and benazepril on large myocardial infarction in rats. Pharmacol Rep 60: 369–381.
[17]
Zhong J, Tang MK, Zhang Y, Xu QP, Zhang JT (2007) Effect of salvianolic acid B on neural cells damage and neurogenesis after brain ischemia-reperfusion in rats. Acta Pharmaceut Sin 42: 716–721.
[18]
Lam FF, Yeung JH, Kwan YW, Chan KM, Or PM (2006) Salvianolic acid B, an aqueous component of danshen (Salvia miltiorrhiza), relaxes rat coronary artery by inhibition of calcium channels. Eur J Pharmacol 553: 240–245.
[19]
Kang DG, Oh H, Chung HT, Lee HS (2003) Inhibition of angiotensin converting enzyme by lithospermic acid B isolated from radix Salviae miltiorrhiza Bunge. Phytother Res 17: 917–920.
[20]
Zhou L, Zuo Z, Chow MS (2005) Danshen: an overview of its chemistry, pharmacology, pharmacokinetics, and clinical use. J Clin Pharmacol 45: 1345–1359.
[21]
Cui L, Liu YY, Wu T, Ai CM, Chen HQ (2009) Osteogenic effects of D+beta-3,4-dihydroxyphenyl lactic acid (salvianic acid A, SAA) on osteoblasts and bone marrow stromal cells of intact and prednisone-treated rats. Acta Pharmacol Sin 30: 321–332.
[22]
Chinese Pharmacopoeia Commission. (2005) Pharmacopoeia of People's Republic of China (Part I). Beijing: Chemical Industry Publishing Co. 668 p.
[23]
Sun Y, Cui L, Wu T (2008) Preparation of Salvia miltiorrhiza utility aqueous extract by hydrochloric acid method and evaluation on osteoblast. Chinese Pharmacological Bulletin 24: 377–381.
[24]
Wang H, Wang Q (2002) Quantitative determination of active constituents in complex prescription of salvia miltiorrhiza by HPLC. Journal of China Pharmaceutical University 33: 219–221.
[25]
Padro T, Ruiz S, Bieker R (2000) Increased angiogenesis in the bone marrow of patients with acute myeloid leukemia. Blood 95: 2637–2644.
[26]
Weidner N (1995) Current pathologic methods for measuring intratumoral microvessel density within breast carcinoma and other solid tumors. Breast Cancer Res Treat 36: 169–180.
[27]
Baron R, Vignery A, Neff L, Silvergate A, Santa-Marria A (1983) Processing of undecalcified bone speciments for bone histomorphometry. In: Recker RR, editor. Bone histomorphometry: techniques and interpretation. Boca Raton, Florida: CRC press. pp. 22–28.
[28]
Cui L, Wu T, Liu XQ, Li QN, Lin LS, et al. (2001) Preventive effects of ginsenoside on osteopenia of rats induced by ovariectomy. Acta Pharmacol Sin 22: 428–434.
[29]
Jee WSS, Li XJ, Inoue J, Jee KW, Haba T, et al. (1997) Histomorphometric assay of the growing long bone. In: Takahashi H, editor. Handbook of Bone Morphology. pp. 87–112. Nishimusa, Niigata City, Japan.
[30]
Parfitt AM, Drezner MK, Glorieux FH (1987) Bone histomorphometry: standardization of nomenclature, symbols and units. Report of the ASBMR Histomorphometry Committee. J Bone Miner Res 2: 595–610.
[31]
Blair HC, Sun L, Kohanski RA (2007) Balanced regulation of proliferation, growth, differentiation, and degradation in skeletal cells. Ann N Y Acad Sci 1116: 165–173.
[32]
Hu CG, Chen JC, Liu Q, He XJ (2004) Mechanism of femoral head necrosis induced by long term glucocorticoid treatment in rabbit model. Chin J Orthop 24: 359–363.
[33]
Parfitt AM (1995) Problems in the application of in vitro systems to the study of human bone remodeling. Proceedings from the workshop on Human Models of Skeletal Aging. NIH. Calcif Tissue Int 56(Suppl.1): S5–7.
[34]
Parfitt AM (1998) Mini-review – Osteoclast precursors as leucocytes: Importance of the area code. Bone 23: 491–494.
[35]
Brandi ML, Collin-Osdoby P (2006) Vascular biology and the skeleton. J Bone Miner Res 21: 183–192.
Hauge EM, Qvesel D, Eriksen EF, Mosekilde L, Melsen F (2001) Cancellous bone remodeling occurs in specialized compartments linked by cells expressing osteoblastic markers. J Bone Miner Res 16: 1575–1582.
[38]
Eghbali-Fatourechi GZ, Lamsan J, Fraser D, Nagel D, Riggs BL, et al. (2005) Circulating osteoblast-lineage cells in humans. N Engl J Med 352: 1959–1966.
[39]
Eriksen EF, Eghbali-Fatourechi GZ, Khosla S (2007) Remodeling and vascular spaces in bone. J Bone Miner Res 22: 1–6.
[40]
Branemark PI (1958) Vital microscopy of bone marrow in rabbit. Scand J Clin Lab Invest 11(suppl 38): 1–82.
[41]
Van Dyke D (1967) Similarity in distribution of skeletal blood flow and erythropoietic marrow. Clin Orthop Relat Res 52: 37–51.
[42]
Reeve J, Arlot M, Wootton R, Edouard C, Tellez M, Hesp R, et al. (1988) Skeleton blood flow iliac histomorphometry and strontium kinetics in osteoporosis: a relationship between blood flow and corrected apposition rate. J Clin Endocrinol Metab 66: 1124–1131.
[43]
Wronski TJ, Smith JM, Jee WSS (1980) The microdistribution and retention of injected 239Pu on trabecular bone surfaces of the beagle: implications for the induction of osteosarcoma. Radiat Res 83: 74–89.
[44]
Wronski TJ, Smith JM, Jee WSS (1981) Variations in mineral apposition rate of trabecular bone within the beagle skeleton. Calcif Tissue Int 33: 583–586.
[45]
Partfitt AM (2008) Skeletal histo genelty and purpose of bone remodeling. Implication for the understanding of osteoporosis. In: Marcus R, Feldman D, Nelson DA, Prosen CJ, editors. Osteoporosis. Third volume. Elever Amstedam. pp. 71–92.
[46]
Tian XY, Setterberg RB, Li X, Paszty C, Ke HZ, Jee WSS (2010) Treatment with a sclerostin antibody increases cancellous bone formation and bone mass regardless of marrow composition in adult female rats. Bone 47: 529–533.
[47]
Liebschner MAK, Keller TS (2005) Hydraulic strengthening affects the stiffness and strength of cortical bone. Ann Biomed Eng 33: 26–38.
[48]
Day TF, Guo X, Garrett-Beal L, Yang Y (2005) Wnt/beta-catenin signaling in mesenchymal progenitors controls osteoblast and chondrocyte differentiation during vertebrate skeletogenesis. Dev Cell 8: 739–750.
[49]
Ohnaka K, Taniguchi H, kawate H (2004) Glucocorticoid enhances the expression of Dickkopf-1 in human osteoblast: novel mechanism of glucocorticoid -induced osteiporosis. Biochem Biophys Res Commun 318: 259–264.
[50]
Ohnaka K (2006) Wnt signaling and glucocorticoid-induced osteoporosis. Clin Calcium 16: 1812–1816.
[51]
Hamidouche Z, Hay E, Vaudin P, Charbord P, Schüle R, et al. (2008) FHL2 mediates dexamethasone-induced mesenchymal cell differentiation into osteoblasts by activating Wnt/beta-catenin signaling-dependent Runx2 expression. FASEB J 22: 3813–3822.
[52]
Nakashima K, Zhou X, Kunkel G, Zhang Z, Deng J, et al. (2002) The novel zinc finger-containing transcription factor osterix is required for osteoblast differentiation and bone formation. Cell 108: 17–29.
[53]
Das A, Botchwey E (2011) Evaluation of angiogenesis and osteogenesis. Tissue Eng Part B Rev 17: 403–414.
[54]
Lu B, Ye Z, Deng Y, Wu H, Feng J (2010) MEK/ERK pathway mediates cytoprotection of salvianolic acid B against oxidative stress – induced apoptosis in rat bone marrow stem cells. Cell Biol Int 34: 1063–1068.
[55]
Li G, Gao Y, Li S, Li C, Zhu X, et al. (2009) Study on toxicity of danshensu in beagle dogs after 3-month continuous intravenous infusion. Toxicol Mech Methods 19: 441–446.
