Icariin promotes osteogenic differentiation of BMSCs by upregulating BMAL1 expression via BMP signaling

Huang,Zengfa; Wei,Hui; Wang,Xiang; Xiao,Jianwei; Li,Zuoqin; Xie,Yuanliang; Hu,Yun; Li,Xiang; Wang,Zheng; Zhang,Shutong

doi:10.3892/mmr.2020.10954

Icariin promotes osteogenic differentiation of BMSCs by upregulating BMAL1 expression via BMP signaling

Authors:
- Zengfa Huang
- Hui Wei
- Xiang Wang
- Jianwei Xiao
- Zuoqin Li
- Yuanliang Xie
- Yun Hu
- Xiang Li
- Zheng Wang
- Shutong Zhang
View Affiliations

Affiliations: Department of Radiology, The Central Hospital of Wuhan, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430014, P.R. China, Department of Orthopedics, The Affiliated Changzhou No. 2 People's Hospital of Nanjing Medical University, Changzhou, Jiangsu 213003, P.R. China
Published online on: January 20, 2020 https://doi.org/10.3892/mmr.2020.10954
Pages: 1590-1596
Copyright: © Huang et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

Increasing research has demonstrated that expression of brain and muscle ARNT‑like 1 (BMAL1) and other circadian clock genes can be regulated by drugs and toxicants. We previously demonstrated that icariin, extracted from Herba Epimedii, sromotes osteogenic differentiation. However, the mechanism underlying the association between icariin and BMAL1 in osteogenic differentiation of bone marrow‑derived mesenchymal stem cells (BMSCs) remains unclear. The present study was designed with an aim to clarify the association between icariin and BMAL1 in osteogenic differentiation of BMSCs. The Cell Counting Kit‑8 assay was used to evaluate cell proliferation. The expression of bone morphogenetic protein 2 (BMP2), RUNX family transcription factor 2 (RUNX2), alkaline phosphatase (ALP), osteocalcin (OC) and BMAL1 in BMSCs was evaluated by reverse transcription‑quantitative PCR and western blotting. ALP and Alizarin red S (ARS) staining were also performed. Icariin promoted BMSC proliferation, and upregulated expression of osteogenic genes and BMAL1. In addition, expression of the osteogenic genes BMP2, RUNX2, ALP and OC were upregulated by BMAL1 overexpression. Furthermore, we confirmed that BMAL1 deficiency suppressed osteogenic differentiation in BMSCs. Finally, ARS staining of BMAL1‑/‑ BMSCs revealed that BMAL1 was an essential intermediary in matrix mineralization during osteogenic differentiation. In conclusion, these results demonstrated that icariin promoted osteogenic differentiation through BMAL1‑BMP2 signaling in BMSCs. The present study thus described a novel target of icariin that has potential applications in the treatment of osteogenic disorders.

View Figures

View References

1	Mankin HJ: Nontraumatic necrosis of bone (osteonecrosis). N Engl J Med. 326:1473–1479. 1992. View Article : Google Scholar : PubMed/NCBI
2	Yu Y, Zhang Y, Wu J, Sun Y, Xiong Z, Niu F, Lei L, Du S, Chen P and Yang Z: Genetic polymorphisms in IL1B predict susceptibility to steroid-induced osteonecrosis of the femoral head in Chinese Han population. Osteoporos Int. 30:871–877. 2019. View Article : Google Scholar : PubMed/NCBI
3	Mont MA, Cherian JJ, Sierra RJ, Jones LC and Lieberman JR: Nontraumatic osteonecrosis of the femoral head: Where do we stand today? A ten-year update. J Bone Joint Surg Am. 97:1604–1627. 2015. View Article : Google Scholar : PubMed/NCBI
4	Shui C, Spelsberg TC, Riggs BL and Khosla S: Changes in Runx2/Cbfa1 expression and activity during osteoblastic differentiation of human bone marrow stromal cells. J Bone Miner Res. 18:213–221. 2003. View Article : Google Scholar : PubMed/NCBI
5	Samsa WE, Vasanji A, Midura RJ and Kondratov RV: Deficiency of circadian clock protein BMAL1 in mice results in a low bone mass phenotype. Bone. 84:194–203. 2016. View Article : Google Scholar : PubMed/NCBI
6	He Y, Chen Y, Zhao Q and Tan Z: Roles of brain and muscle ARNT-like 1 and Wnt antagonist Dkk1 during osteogenesis of bone marrow stromal cells. Cell Prolif. 46:644–653. 2013. View Article : Google Scholar : PubMed/NCBI
7	Mohawk JA, Green CB and Takahashi JS: Central and peripheral circadian clocks in mammals. Annu Rev Neurosci. 35:445–462. 2012. View Article : Google Scholar : PubMed/NCBI
8	Richards J and Gumz ML: Mechanism of the circadian clock in physiology. Am J Physiol Regul Integr Comp Physiol. 304:R1053–R1064. 2013. View Article : Google Scholar : PubMed/NCBI
9	DeBruyne JP, Weaver DR and Dallmann R: The hepatic circadian clock modulates xenobiotic metabolism in mice. J Biol Rhythms. 29:277–287. 2014. View Article : Google Scholar : PubMed/NCBI
10	Zmrzljak UP and Rozman D: Circadian regulation of the hepatic endobiotic and xenobitoic detoxification pathways: The time matters. Chem Res Toxicol. 25:811–824. 2012. View Article : Google Scholar : PubMed/NCBI
11	Bailey SM, Udoh US and Young ME: Circadian regulation of metabolism. J Endocrinol. 222:R75–R96. 2014. View Article : Google Scholar : PubMed/NCBI
12	Miranda J, Portillo MP, Madrid JA, Arias N, Macarulla MT and Garaulet M: Effects of resveratrol on changes induced by high-fat feeding on clock genes in rats. Br J Nutr. 110:1421–1428. 2013. View Article : Google Scholar : PubMed/NCBI
13	Chen P, Kakan X and Zhang J: Altered circadian rhythm of the clock genes in fibrotic livers induced by carbon tetrachloride. FEBS Lett. 584:1597–1601. 2010. View Article : Google Scholar : PubMed/NCBI
14	Gabás-Rivera C, Martínez-Beamonte R, Ríos JL, Navarro MA, Surra JC, Arnal C, Rodríguez-Yoldi MJ and Osada J: Dietary oleanolic acid mediates circadian clock gene expression in liver independently of diet and animal model but requires apolipoprotein A1. J Nutr Biochem. 24:2100–2109. 2013. View Article : Google Scholar : PubMed/NCBI
15	Huang Z, Cheng C, Wang J, Liu X, Wei H, Han Y, Yang S and Wang X: Icariin regulates the osteoblast differentiation and cell proliferation of MC3T3-E1 cells through microRNA-153 by targeting Runt-related transcription factor 2. Exp Ther Med. 15:5159–5166. 2018.PubMed/NCBI
16	Huang Z, Cheng C, Cao B, Wang J, Wei H, Liu X, Han Y, Yang S and Wang X: Icariin protects against glucocorticoid-induced osteonecrosis of the femoral head in rats. Cell Physiol Biochem. 47:694–706. 2018. View Article : Google Scholar : PubMed/NCBI
17	Gong H, Wang X, Wang L, Liu Y, Wang QL, Pang H, Zhang Q and Wang Z: Inhibition of IGF-1 receptor kinase blocks the differentiation into cardiomyocyte-like cells of BMSCs induced by IGF-1. Molecular medicine reports. 16:787–793. 2017. View Article : Google Scholar : PubMed/NCBI
18	Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods. 25:402–408. 2001. View Article : Google Scholar : PubMed/NCBI
19	Dierickx P, Van Laake LW and Geijsen N: Circadian clocks: From stem cells to tissue homeostasis and regeneration. EMBO Rep. 19:18–28. 2018. View Article : Google Scholar : PubMed/NCBI
20	Murray IR, West CC, Hardy WR, James AW, Park TS, Nguyen A, Tawonsawatruk T, Lazzari L, Soo C and Péault B: Natural history of mesenchymal stem cells, from vessel walls to culture vessels. Cell Mol Life Sci. 71:1353–1374. 2014. View Article : Google Scholar : PubMed/NCBI
21	Weger M, Diotel N, Dorsemans AC, Dickmeis T and Weger BD: Stem cells and the circadian clock. Dev Biol. 431:111–123. 2017. View Article : Google Scholar : PubMed/NCBI
22	Fu L, Patel MS, Bradley A, Wagner EF and Karsenty G: The molecular clock mediates leptin-regulated bone formation. Cell. 122:803–815. 2005. View Article : Google Scholar : PubMed/NCBI
23	Gosselet FP, Magnaldo T, Culerrier RM, Sarasin A and Ehrhart JC: BMP2 and BMP6 control p57(Kip2) expression and cell growth arrest/terminal differentiation in normal primary human epidermal keratinocytes. Cell Signal. 19:731–739. 2007. View Article : Google Scholar : PubMed/NCBI
24	Rogers MB, Shah TA and Shaikh NN: Turning bone morphogenetic protein 2 (BMP2) on and off in mesenchymal cells. J Cell Biochem. 116:2127–2138. 2015. View Article : Google Scholar : PubMed/NCBI
25	Shu B, Zhang M, Xie R, Wang M, Jin H, Hou W, Tang D, Harris SE, Mishina Y, O'Keefe RJ, et al: BMP2, but not BMP4, is crucial for chondrocyte proliferation and maturation during endochondral bone development. J Cell Sci. 124:3428–3440. 2011. View Article : Google Scholar : PubMed/NCBI
26	Hirai T, Tanaka K and Togari A: α1-adrenergic receptor signaling in osteoblasts regulates clock genes and bone morphogenetic protein 4 expression through up-regulation of the transcriptional factor nuclear factor IL-3 (Nfil3)/E4 promoter-binding protein 4 (E4BP4). J Biol Chem. 289:17174–17183. 2014. View Article : Google Scholar : PubMed/NCBI
27	Nam D, Guo B, Chatterjee S, Chen MH, Nelson D, Yechoor VK and Ma K: The adipocyte clock controls brown adipogenesis through the TGF-β and BMP signaling pathways. J Cell Sci. 128:1835–1847. 2015. View Article : Google Scholar : PubMed/NCBI
28	Tasaki H, Zhao L, Isayama K, Chen H, Yamauchi N, Shigeyoshi Y, Hashimoto S and Hattori MA: Inhibitory role of REV-ERBα in the expression of bone morphogenetic protein gene family in rat uterus endometrium stromal cells. Am J Physiol Cell Physiol. 308:C528–C538. 2015. View Article : Google Scholar : PubMed/NCBI
29	Gloston GF, Yoo SH and Chen ZJ: Clock-enhancing small molecules and potential applications in chronic diseases and aging. Front Neurol. 8:1002017. View Article : Google Scholar : PubMed/NCBI
30	Sun L, Wang Y, Song Y, Cheng XR, Xia S, Rahman MR, Shi Y and Le G: Resveratrol restores the circadian rhythmic disorder of lipid metabolism induced by high-fat diet in mice. Biochem Biophys Res Commun. 458:86–91. 2015. View Article : Google Scholar : PubMed/NCBI
31	Tsuchiya S, Sugiyama K and Van Gelder RN: Adrenal and glucocorticoid effects on the circadian rhythm of murine intraocular pressure. Invest Ophthalmol Vis Sci. 59:5641–5647. 2018. View Article : Google Scholar : PubMed/NCBI
32	Yang P, Guan YQ, Li YL, Zhang L, Zhang L and Li L: Icariin promotes cell proliferation and regulates gene expression in human neural stem cells in vitro. Mol Med Rep. 14:1316–1322. 2016. View Article : Google Scholar : PubMed/NCBI
33	Mok SK, Chen WF, Lai WP, Leung PC, Wang XL, Yao XS and Wong MS: Icariin protects against bone loss induced by oestrogen deficiency and activates oestrogen receptor-dependent osteoblastic functions in UMR 106 cells. Br J Pharmacol. 159:939–949. 2010. View Article : Google Scholar : PubMed/NCBI
34	Chen SR, Xu XZ, Wang YH, Chen JW, Xu SW, Gu LQ and Liu PQ: Icariin derivative inhibits inflammation through suppression of p38 mitogen-activated protein kinase and nuclear factor-kappaB pathways. Biol Pharm Bull. 33:1307–1313. 2010. View Article : Google Scholar : PubMed/NCBI
35	Xu CQ, Liu BJ, Wu JF, Xu YC, Duan XH, Cao YX and Dong JC: Icariin attenuates LPS-induced acute inflammatory responses: Involvement of PI3K/Akt and NF-kappaB signaling pathway. Eur J Pharmacol. 642:146–153. 2010. View Article : Google Scholar : PubMed/NCBI
36	Zhou J, Wu J, Chen X, Fortenbery N, Eksioglu E, Kodumudi KN, Pk EB, Dong J, Djeu JY and Wei S: Icariin and its derivative, ICT, exert anti-inflammatory, anti-tumor effects, and modulate myeloid derived suppressive cells (MDSCs) functions. Int Immunopharmacol. 11:890–898. 2011. View Article : Google Scholar : PubMed/NCBI