MiR-21协同BMP9促进间充质干细胞C3H10T1/2成骨分化

doi:10.13523/j.cb.20160204

中国生物工程杂志

2016, Vol. 36

Issue (2): 22-29 DOI: 10.13523/j.cb.20160204

研究报告

MiR-21协同BMP9促进间充质干细胞C3H10T1/2成骨分化

宋琪玲, 施琼, 陈楚, 唐祖川, 刘红霞, 周一青, 张汝益, 严树涓, 翁亚光

重庆医科大学检验医学院临床检验诊断学教育部重点实验室重庆 400016

MiR-21 Synergizes with BMP9 in Osteogenic Differentiation During Mesenchymal Stem Cells Line C3H10T1/2

SONG Qi-ling, SHI Qiong, CHEN Chu, TANG Zu-chuan, LIU Hong-xia, ZHOU Yi-qing, ZHANG Ru-yi, YAN Shu-juan, WENG Ya-guang

Key Laboratory of Clinical Laboratory Diagnostics of Ministry Education, Faculty of Laboratory Medicine, Chongqing Medicine University, Chongqing 400016, China

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摘要：

目的:探讨miR-21与BMP9之间的关系,明确miR-21在BMP9诱导间充质干细胞成骨分化中的作用。方法:(1)Ad-BMP9感染C3H10T1/2细胞,Real-time-PCR检测miR-21表达。RT-PCR检测ALP的表达。(2)MiR-21转染C3H10T1/2细胞,Real-time-PCR检测miR-21和BMP9表达。(3)MiR-21和BMP9-CM处理C3H10 T1/2细胞,ALP活性和染色实验检测C3H10 T1/2细胞早期成骨能力。茜素红S染色实验检测钙盐沉积情况。(4)MiR-21和BMP9-CM处理C3H10 T1/2细胞,Real-time-PCR检测成骨分化相关因子ALP,OCN的表达。(5)MiR-21和BMP9-CM处理C3H10T1/2细胞,Western blot检测p-Smad1/5蛋白水平的表达。结果:(1)BMP9暂时降低miR-21的表达。MiR-21也可以暂时降低BMP9的表达。(2)MiR-21可以协同BMP9增强ALP和钙盐沉积。(3)MiR-21协同BMP9增加了p-Smad1/5蛋白水平的表达。结论:MiR-21与BMP9存在相互关系,两者可以互相调节表达。MiR-21可以协同BMP9促进间充质干细胞C3H10T1/2细胞成骨分化,这一过程与增强BMP9/Smad信号的激活程度有关。

关键词： MiR-21; C3H10T1/2细胞; 骨形成蛋白9; 成骨分化

Abstract:

Objective:(1) To study if there is a crosstalk between bone morphogenetic protein 9(BMP9) and microRNA-21(miR-21);(2) to investigate the effect of miR-21 on BMP9-induced osteogenic differentiation of mesenchymal stem cells line C3H10T1/2.Methods:(1) Using Ad-BMP9 infecting C3H10 T1/2 cells, the expression of miR-21 were detected by Real-time-PCR. After transfecting miR-21 mimic 1 day and 5 days, the expression of BMP were tested by Real-time-PCR.(2) In vitro, after transfecting miR-21, BMP9-CM treating C3H10T1/2, ALP staining and ALP activity were detected and the calcium deposition were detected by Alizarin Red S staining.(3)The expression of BMP9 downstream factors ALP and OCN were detected by Real-time-PCR.(4) The protein expression of p-Smad1/5 were detected by Western blot. Results:BMP9 can decrease the expression of miR-21 temporarily and miR-21 also decreases the expression of BMP9 temporarily. After transfecting miR-21 and treating C3H10T1/2 cell with BMP9-CM, the early osteogenic differentiation marker ALP activity and ALP staining and later osteogenic differentiation marker calcium deposition were enhanced. What's more, the expression of ostengenic factors ALP and OCN also increase during miR-21 combined with BMP9-CM. The protein expression of p-Smad1/5 were upregulation during the same process. Conclusion:There is a crosswalk between BMP9 and miR-21. MiR-21 synergizes with BMP9 promoting mesenchymal stem cells C3H10T1/2 osteogenic differentiation by increasing BMP9/Smad signaling pathway activated.

Key words: Bone morphogenetic protein9 Osteogenic differentiation MiR-21 C3H10 T1/2 cells

收稿日期: 2015-07-27 出版日期: 2015-11-19

ZTFLH:

Q819

基金资助:

国家自然科学基金(NSFC 31200971)、教育部博士点基金(20115503110009)、重庆市教育委员会科学技术研究项目(KJ120327)资助项目

通讯作者: 翁亚光 E-mail: yaguangweng@126.com

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引用本文:

宋琪玲, 施琼, 陈楚, 唐祖川, 刘红霞, 周一青, 张汝益, 严树涓, 翁亚光. MiR-21协同BMP9促进间充质干细胞C3H10T1/2成骨分化[J]. 中国生物工程杂志, 2016, 36(2): 22-29.

SONG Qi-ling, SHI Qiong, CHEN Chu, TANG Zu-chuan, LIU Hong-xia, ZHOU Yi-qing, ZHANG Ru-yi, YAN Shu-juan, WENG Ya-guang. MiR-21 Synergizes with BMP9 in Osteogenic Differentiation During Mesenchymal Stem Cells Line C3H10T1/2. China Biotechnology, 2016, 36(2): 22-29.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/10.13523/j.cb.20160204 或 https://manu60.magtech.com.cn/biotech/CN/Y2016/V36/I2/22

[1] Lian J B,Stein G S,van Wijnen A J,et al. MicroRNA control of bone formation and homeostasis. Nat Rev Endocrinol,2012,8(4):212-227.
[2] Kim Y J,Hwang S J,Bae Y C,et al. MiR-21 regulates adipogenic differentiation through the modulation of TGF-beta signaling in mesenchymal stem cells derived from human adipose tissue. Stem Cells,2009,27(12):3093-3102.
[3] Yang N,Wang G,Hu C,et al. Tumor necrosis factor α suppresses the mesenchymal stem cell osteogenesis promoter miR-21 in estrogen deficiency-induced osteoporosis. J Bone Miner Res,2013,28(3):559-573.
[4] 杨楠,周威,王光,等.Spry1在miR-21调控人骨髓间充质干细胞成骨分化中的作用.中国组织工程研究,2014,18(32):5085-5090. Yang N,Zhou W,Wang G,et al. Role of Spry1 in osteogenic differentiation of human bone marrow mesenchymal stem cells under miR-21 regulation. Chinese Journal of Tissue Engineenng Research,2014,18(32):5085-5090.
[5] Mei Y,Bian C,Li J,et al. miR-21 modulates the ERK-MAPK signaling pathway by regulating SPRY2 expression during human mesenchymal stem cell differentiation. J Cell Biochem,2013,114(6):1374-1384.
[6] Kang Q,Sun M H,Cheng H,et al. Characterization of the distinct orthotopic bone-forming activity of 14 BMPs using recombinant adenovirus-mediated gene delivery. Gene Ther,2004,11(17):1312-1320.
[7] Lamplot J D,Qin J,Nan G,et al. BMP9 signaling in stem cell differentiation and osteogenesis. Am J Stem Cells,2013,2(1):1-21.
[8] Luther G,Wagner E R,Zhu G,et al. BMP-9 induced osteogenic differentiation of mesenchymal stem cells:molecular mechanism and therapeutic potential. Curr Gene Ther,2011,11(3):229-240.
[9] Cheng H,Jiang W,Phillips F M,et al. Osteogenic activity of the fourteen types of human bone morphogenetic proteins(BMPs). J Bone Joint Surg Am,2003,85-A(8):1544-1552.
[10] Davis B N,Hilyard A C,Lagna G,et al. SMAD proteins control DROSHA-mediated microRNA maturation. Nature,2008,454(7200):56-61.
[11] 李宝林,白慧丽,张汝益,等. miR-30a抑制BMP9诱导间充质干细胞C3H10T1/2的成骨分化. 中国生物工程杂志,2013,33(11):14-20. Li B L,Bai H L,Zhang R Y,et al.BMP9-induced osteogenic differentiationis partially inhibited by miR-30a in mesenchymal stem cells Line C3H10T1/2. China Biotechnology,2013,33(11):14-20.
[12] Wu J Y,Aarnisalo P,Bastepe M,et al. Gsa enhances commitment of mesenchymal progenitors to the osteoblast lineage but restrains osteoblast differentiation in mice. J Clin Invest,2011,121(9):3492-3504.
[13] Luo J,Deng Z L,Luo X,et al. A protocol for rapid generation of recombinant adenoviruses using the AdEasy system. Nat Protoc,2007,2(5):1236-1247.
[14] Wang X,Guo B,Li Q,et al. miR-214 targets ATF4 to inhibit bone formation. Nat Med,2013,19(1):93-100.
[15] Liu Y,Liu Y,Zhang R,et al. All-trans retinoic acid modulates bone morphogenic protein 9-induced osteogenesis and adipogenesis of preadipocytes through BMP/Smad and Wnt/β-catenin signaling pathways. Int J Biochem Cell Biol,2014,47:47-56.
[16] Krichevsky A M,Gabriely G. miR-21:a small multi-faceted RNA. J Cell Mol Med,2009,13(1):39-53.
[17] Naznin Akhtar,Hossen Mohammad Jamil,Rajat Suvra Banik,et al.TGF-β/BMP signaling and other molecular events:regulation of osteoblastogenesis and bone formation. Bone Research,2015,3:15005.

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