Abstract
Aim:
Our previous studies show that salvianolic acid B (Sal B) promotes osteoblast differentiation and matrix mineralization. In this study, we evaluated the protective effects of Sal B on the osteogenesis in dexamethasone (Dex)-treated larval zebrafish, and elucidated the underlying mechanisms.
Methods:
At 3 d post fertilization, wild-type AB zebrafish larvae or bone transgenic tg (sp7:egfp) zebrafish larvae were exposed to Sal B, Dex, or a mixture of Dex+Sal B for 6 d. Bone mineralization in AB strain larval zebrafish was assessed with alizarin red staining, and osteoblast differentiation in tg (sp7:egfp) larval zebrafish was examined with fluorescence scanning. The expression of osteoblast-specific genes in the larvae was detected using qRT-PCR assay. The levels of oxidative stress markers (ROS and MDA) in the larvae were also measured.
Results:
Exposure to Dex (5–20 μmol/L) dose-dependently decreased the bone mineralization area and integral optical density (IOD) in wild-type AB zebrafish larvae and the osteoblast fluorescence area and IOD in tg (sp7:egfp) zebrafish larvae. Exposure to Dex (10 μmol/L) significantly reduced the expression of osteoblast-specific genes, including runx2a, osteocalcin (OC), alkaline phosphatase (ALP) and osterix (sp7), and increased the accumulation of ROS and MDA in the larvae. Co-exposure to Sal B (0.2–2 μmol/L) dose-dependently increased the bone mineralization area and IOD in AB zebafish larvae and osteoblast fluorescence in tg (sp7:egfp) zebrafish larvae. Co-exposure to Sal B (2 μmol/L) significantly attenuated deleterious alterations in bony tissue and oxidative stress in both Dex-treated AB zebafish larvae and tg (sp7:egfp) zebrafish larvae.
Conclusion:
Sal B stimulates bone formation and rescues GC-caused inhibition on osteogenesis in larval zebrafish by counteracting oxidative stress and increasing the expression of osteoblast-specific genes. Thus, Sal B may have protective effects on bone loss trigged by GC.
Similar content being viewed by others
Log in or create a free account to read this content
Gain free access to this article, as well as selected content from this journal and more on nature.com
or
References
Frenkel B, White W, Tuckermann J . Glucocorticoid-induced osteoporosis. Adv Exp Med Biol 2015; 872: 179–215.
Kaneko K, Kawai S . Mechanisms and therapeutics of glucocorticoid-induced osteoporosis. Nihon Rinsho Meneki Gakkai Kaishi 2011; 34: 138–48. Japanese.
Sharma T, Islam N, Ahmad J, Akhtar N, Beg M . Correlation between bone mineral density and oxidative stress in postmenopausal women. Indian J Endocrinol Metab 2015; 19: 491–7.
Cervellati C, Bonaccorsi G, Cremonini E, Bergamini CM, Patella A, Castaldini C, et al. Bone mass density selectively correlates with serum markers of oxidative damage in post-menopausal women. Clin Chem Lab Med 2013; 51: 333–8.
Arslan A, Orkun S, Aydin G, Keles I, Tosun A, Arslan M, et al. Effects of ovariectomy and ascorbic acid supplement on oxidative stress parameters and bone mineral density in rats. Libyan J Med 2011; 6: doi: 10.3402/ljm.v6i0.5965.
Muller FL, Lustgarten MS, Jang Y, Richardson A, Van Remmen H . Trends in oxidative aging theories. Free Radic Biol Med 2007; 43: 477–503.
Huang CX, Lv B, Wang Y . Protein phosphatase 2A mediates oxidative stress induced apoptosis in osteoblasts. Mediators Inflamm 2015; 2015: 804260.
Sato AY, Tu X, McAhdrews KA, Plotkin LI, Bellido T . Prevention of glucocorticoid induced-apoptosis of osteoblasts and osteocytes by protecting against endoplasmic reticulum (ER) stress in vitro and in vivo female mice. Bone 2015; 73: 60–8.
Shi J, Wang L, Zhang H, Jie Q, Li X, Shi Q, et al. Glucocorticoids: Dose-related effects on osteoclast formation and function via reactive oxygen species and autophagy. Bone 2015; 79: 222–32.
Suda N, Morita I, Kuroda T, Murota S . Participation of oxidative stress in the process of osteoclast differentiation. Biochim Biophys Acta 1993; 1157: 318–23.
Hurson CJ, Butler JS, Keating DT, Murray DW, Sadlier DM, O'Byrne JM, et al. Gene expression analysis in human osteoblasts exposed to dexamethasone identifies altered developmental pathways as putative drivers of osteoporosis. BMC Musculoskelet Disord 2007; 8: 12.
Feng YL, Tang XL . Effect of glucocorticoid-induced oxidative stress on the expression of Cbfa1. Chem Biol Interact 2014; 207: 26–31.
Almeida M, Han L, Ambrogini E, Weinstein RS, Manolagas SC . Glucocorticoids and tumor necrosis factor alpha increase oxidative stress and suppress Wnt protein signaling in osteoblasts. J Biol Chem 2011; 286: 44326–35.
Chen CY, Li H, Yuan YN, Dai HQ, Yang B . Antioxidant activity and components of a traditional Chinese medicine formula consisting of Crataegus pinnatifida and Salvia miltiorrhiza. BMC Complement Altern Med 2013; 13: 99.
Tang Y, Jacobi A, Vater C, Zou X, Stiehler M . Salvianolic acid B protects human endothelial progenitor cells against oxidative stress-mediated dysfunction by modulating Akt/mTOR/4EBP1, p38 MAPK/ATF2, and ERK1/2 signaling pathways. Biochem Pharmacol 2014; 90: 34–49.
Cui L, Li T, Liu Y, Zhou L, Li P, Xu B, et al. Salvianolic acid B prevents bone loss in prednisone-treated rats through stimulation of osteogenesis and bone marrow angiogenesis. PLoS One 2012; 7: e34647.
Cui L, Liu YY, Wu T, Ai CM, Chen HQ . 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 2009; 30: 321–32.
Xu D, Xu L, Zhou C, Lee WY, Wu T, Cui L, et al. Salvianolic acid B promotes osteogenesis of human mesenchymal stem cells through activating ERK signaling pathway. Int J Biochem Cell Biol 2014; 51: 1–9.
Goldsmith P . Zebrafish as a pharmacological tool: the how, why and when. Curr Opin Pharmacol 2004; 4: 504–12.
Barros TP, Alderton WK, Reynolds HM, Roach AG, Berghmans S . Zebrafish: an emerging technology for in vivo pharmacological assessment to identify potential safety liabilities in early drug discovery. Br J Pharmacol 2008; 154: 1400–13.
Barrett R, Chappell C, Quick M, Fleming A . A rapid, high content, in vivo model of glucocorticoid-induced osteoporosis. Biotechnol J 2006; 1: 651–5.
Chen B, Yan YL, Liu C, Bo L, Li GF, Wang H, et al. Therapeutic effect of deferoxamine on iron overload-induced inhibition of osteogenesis in a zebrafish model. Calcif Tissue Int 2014; 94: 353–60.
Jing LJ, Wang CM, Wei YJ, Jia XB, Xie L . Screening on antiosteoporotic active parts of dipsacus radix based on zebrafish model. Zhong Yao Cai 2014; 37: 635–40.
Li N, Felber K, Elks P, Croucher P, Roehl HH . Tracking gene expression during zebrafish osteoblast differentiation. Dev Dyn 2009; 238: 459–66.
DeLaurier A, Eames BF, Blanco-Sanchez B, Peng G, He X, Swartz ME, et al. Zebrafish sp7:EGFP: a transgenic for studying otic vesicle formation, skeletogenesis, and bone regeneration. Genesis 2010; 48: 505–11.
Knopf F, Hammond C, Chekuru A, Hans S, Weber CW, Mahatma G, et al. Bone regenerates via dedifferentiation of osteoblasts in the zebrafish fin. Dev Cell 2011; 20: 713–24.
Westerfield M . The zebrafish book. Oregon: University of Oregon Press; 2000.
Walker MB, Kimmel CB . A two-color acid-free cartilage and bone stain for zebrafish larvae. Biotech Histochem 2007; 82: 23–8.
Fleming A, Sato M, Goldsmith P . High-throughput in vivo screening for bone anabolic compounds with zebrafish. J Biomol Screen 2005; 10: 823–31.
Ali S, Champagne DL, Spaink HP, Richardson MK . Zebrafish embryos and larvae: a new generation of disease models and drug screens. Birth Defects Res C Embryo Today 2011; 93: 115–33.
Haga Y, Dominique VR, Du SJ . Analyzing notochord segmentation and intervertebral disc formation using the twhh:gfp transgenic zebrafish model. Transgenic Res 2009; 18: 669–83.
Mork L, Crump G . Zebrafish craniofacial development: a window into early patterning. Curr Top Dev Biol 2015; 115: 235–69.
Witten PE, Hansen A, Hall BK . Features of mono- and multinucleated bone resorbing cells of the zebrafish Danio rerio and their contribution to skeletal development, remodeling, and growth. J Morphol 2001; 250: 197–207.
Sharif F, de Bakker MA, Richardson MK . Osteoclast-like cells in early Zebrafish embryos. Cell J 2014; 16: 211–24.
Hui R, Liang D, Jiang X, Tang J, Cui J, Wei Q, et al. Variance of spinal osteoporosis induced by dexamethasone and methylprednisolone and its associated mechanism. Steroids 2015; 102: 65–75.
Zheng G, Zhang X, Meng Q, Gong W, Wen X, Xie H . Protective effect of total isoflavones from Pueraria lobata on secondary osteoporosis induced by dexamethasone in rats. Zhong Yao Cai 2002; 25: 643–6.
Thamamongood TA, Furuya R, Fukubas M, Nakamura M, Suzuki N, Hattori A . Expression of osteoblastic and osteoclastic genes during spontaneous regeneration and autotransplantation of goldfish scale: a new tool to study intramembranous bone regeneration. Bone 2012; 50: 1240–9.
Matsumoto T, Endo I . Eldecalcitol for the treatment of osteoporosis. Drugs Today 2012; 48: 189–96.
Yang Y, Su Y, Wang D, Chen Y, Wu T, Li G, et al. Tanshinol attenuates the deleterious effects of oxidative stress on osteoblastic differentiation via Wnt/FoxO3a signaling. Oxid Med Cell Longev 2013; 2013: 351895.
Acknowledgements
The authors are thankful to Dr Chung-der HSIAO's group from Chung Yuan Christian University, Taiwan, China for use of the tg(sp7:egfp) zebrafish. This research was supported by grants from the National Natural Science Foundation of China (81273518 and 31370824), the Science and Technology Planning Project of Guangdong Province (No 2012B060300027) and the Scientific Research Subject of Traditional Chinese Medicine Bureau of Guangdong Province (No 20152152).
Author information
Authors and Affiliations
Corresponding authors
Rights and permissions
About this article
Cite this article
Luo, Sy., Chen, Jf., Zhong, Zg. et al. Salvianolic acid B stimulates osteogenesis in dexamethasone-treated zebrafish larvae. Acta Pharmacol Sin 37, 1370–1380 (2016). https://doi.org/10.1038/aps.2016.62
Received:
Accepted:
Published:
Issue date:
DOI: https://doi.org/10.1038/aps.2016.62
Keywords
This article is cited by
-
Dynamic network biomarker identifies cdkn1a-mediated bone mineralization in the triggering phase of osteoporosis
Experimental & Molecular Medicine (2023)
-
New sequence division method of shallow platform with natural gamma spectrometry data: implication for reservoir distribution—a case study from Majiagou Formation of Bozhong 21–22 structure, Bohai Bay Basin
Carbonates and Evaporites (2020)
