Chondrogenic differentiation of bone marrow mesenchymal stem cells induced by different growth factors

doi:10.3969/j.issn.2095-4344.2015.45.004

Abstract

Abstract:

BACKGROUND: Articular cartilage is a highly differentiated tissue, which is very limited in its ability to repair after injury. Stem cell therapy for cartilage repair has completely solved this problem.

OBJECTIVE: To investigate the mechanism that different growth factors induce the differentiation of bone marrow mesenchymal stem cells into chondrocytes.

METHODS: Passage 5 rat bone marrow mesenchymal stem cells were induced by different growth factors and their combinations, including transforming growth factor beta 1 (TGF-β1) group, TGF-β1+insulin growth factor-1 (IGF-1) group, and bone morphogenetic protein 2 (BMP-2)+IGF-1 group, TGF-β1+BMP-2 group, TGF-β1+IGF-1+ BMP-2 group, and blank control group. At 21 days of induction, cells were stained with alcian blue and alizarin red; RT-PCR was employed to detect collagen II mRNA expression.

RESULTS AND CONCLUSION: Alcian blue staining showed metachromasia in the cytoplasm and mesenchyma, and proteoglycan was expressed green; alizarin red staining showed no orange calcium nodules. The bone marrow mesenchymal stem cells were preliminarily deduced to differentiate into chondrocytes, but could not express the cell phenotypes of bone cells. In the blank control group, the expression of collagen II mRNA was negative. Compared with the TGF-β1 group, the mRNA expression of collagen II was lower in the BMP-2+IGF-1 group, but higher in the TGF-β1+BMP-2 group and TGF-β1+IGF-1+BMP-2 group (P < 0.05). Moreover, the expression of collagen II mRNA was highest in the TGF-β1+IGF-1+BMP-2 group (P < 0.05). These findings indicate that TGF-β1 alone is able to induce the chondrogenic differentiation of bone marrow mesenchymal stem cells, and the combination of TGF-β1, IGF-1 and BMP-2 can play the biggest role to induce the chondrogenic differentiation of bone marrow mesenchymal stem cells.
中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

Key words: Bone Marrow, Mesenchymal Stem Cells, Chondrocytes, Transforming Growth Factor beta1, Insulin-Like Growth Factor I, Bone Morphogenetic Proteins, Tissue Engineering

Wang Shu-ying, Lu Xiao-bing, Ouyang Lin. Chondrogenic differentiation of bone marrow mesenchymal stem cells induced by different growth factors[J]. Chinese Journal of Tissue Engineering Research, 2015, 19(45): 7237-7241.

Figures/Tables 3

References

[1] Wu J, Wang F, Yang L, et al. Chondrocytic biomimetic matrix materials-induced differentiation of rat bone marrow mesenchymal stem cells into chondrocyte-like cells in vitro.Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2015;31(11): 1502-1505.

[2] Liu P, Sun L, Chen H, et al. Lentiviral-mediated multiple gene transfer to chondrocytes promotes chondrocyte differentiation and bone formation in rabbit bone marrow-derived mesenchymal stem cells. Oncol Rep. 2015;34(5):2618-2626.

[3] Zhang X, Hirai M, Cantero S, et al. Isolation and characterization of mesenchymal stem cells from human umbilical cord blood: reevaluation of critical factors for successful isolation and high ability to proliferate and differentiate to chondrocytes as compared to mesenchymal stem cells from bone marrow and adipose tissue. J Cell Biochem. 2011;112(4):1206-1218.

[4] Murphy MK, Huey DJ, Hu JC, et al. TGF-β1, GDF-5, and BMP-2 stimulation induces chondrogenesis in expanded human articular chondrocytes and marrow-derived stromal cells. Stem Cells. 201533(3):762-773.

[5] Wang H, Li Y, Chen J, et al. Chondrogenesis of bone marrow mesenchymal stem cells induced by transforming growth factor beta3 gene in Diannan small-ear pigs.Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2014;28(2):149-154.

[6] Wang F, Zhang YC, Zhou H, et al. Evaluation of in vitro and in vivo osteogenic differentiation of nano-hydroxyapatite/ chitosan/poly(lactide-co-glycolide) scaffolds with human umbilical cord mesenchymal stem cells. J Biomed Mater Res A. 2014;102(3):760-768.

[7] 甘凤英.生长因子联合诱导大鼠骨髓间充质干细胞向软骨分化的实验研究[D].福州:福建医科大学,2006.

[8] Bosnakovski D, Mizuno M, Kim G, et al. Chondrogenic differentiation of bovine bone marrow mesenchymal stem cells (MSCs) in different hydrogels: influence of collagen type II extracellular matrix on MSC chondrogenesis. Biotechnol Bioeng. 2006;93(6):1152-1163.

[9] 周瑜.慢病毒介导TGF-β3/BMP-2联合基因转染诱导兔骨髓间充质干细胞软骨分化的实验研究[D]. 青岛:青岛大学,2014.

[10] 周瑜,夏长所,王昌耀,等.慢病毒介导TGF-β3/BMP-2联合基因转染诱导兔骨髓间充质干细胞成软骨细胞分化[J].中华临床医师杂志:电子版,2014,8(4):688-694.

[11] Dahlin RL, Meretoja VV, Ni M, et al. Chondrogenic phenotype of articular chondrocytes in monoculture and co-culture with mesenchymal stem cells in flow perfusion. Tissue Eng Part A. 2014;20(21-22):2883-2891.

[12] Liu S, Zhang E, Yang M, et al. Overexpression of Wnt11 promotes chondrogenic differentiation of bone marrow-derived mesenchymal stem cells in synergism with TGF-β. Mol Cell Biochem. 2014;390(1-2):123-131.

[13] Moghadam FH, Tayebi T, Dehghan M, et al. Differentiation of bone marrow mesenchymal stem cells into chondrocytes after short term culture in alkaline medium. Int J Hematol Oncol Stem Cell Res. 2014;8(4):12-19.

[14] 刘吉斌,侯连东,于强,等.骨膜复合骨髓间充质干细胞和软骨细胞体内移植修复软骨缺损[J].中国组织工程研究,2015,19(3): 364-369.

[15] Zhang Z, Wang Y, Li M, et al. Fibroblast growth factor 18 increases the trophic effects of bone marrow mesenchymal stem cells on chondrocytes isolated from late stage osteoarthritic patients. Stem Cells Int. 2014;2014:125683.

[16] Leyh M, Seitz A, Dürselen L, et al. Subchondral bone influences chondrogenic differentiation and collagen production of human bone marrow-derived mesenchymal stem cells and articular chondrocytes.Arthritis Res Ther. 2014;16(5):453.

[17] Wang J, Li J, Deng N, et al. Transfection of hBMP-2 into mesenchymal stem cells derived from human umbilical cord blood and bone marrow induces cell differentiation into chondrocytes. Minerva Med. 2014;105(4):283-288.

[18] 陈伟,王丽娜,郝云甲,等.不同来源软骨细胞共培养对骨髓间充质干细胞早期分化的影响[J].中华老年医学杂志,2015,34(5): 557-560.

[19] 李斌,华永新,曲华正,等.软骨细胞和骨髓间充质干细胞相关基因转染的研究进展[J].医学与哲学,2014,35(24):58-61.

[20] 李斌,张伟,王健,等.体外微团培养兔骨髓间充质干细胞形成软骨细胞[J].中国矫形外科杂志,2009,17(23):1819-1822.

[21] Nixon AJ, Fortier LA, Williams J, et al. Enhanced repair of extensive articular defects by insulin-like growth factor-I-laden fibrin composites. J Orthop Res. 1999;17(4):475-487.

[22] Sekiya I, Larson BL, Vuoristo JT, et al. Comparison of effect of BMP-2, -4, and -6 on in vitro cartilage formation of human adult stem cells from bone marrow stroma. Cell Tissue Res. 2005;320(2):269-276.

[23] Olmez U, Ryan LM, Kurup IV, et al. Insulin-like growth factor-1 suppresses pyrophosphate elaboration by transforming growth factor beta1-stimulated chondrocytes and cartilage. Osteoarthritis Cartilage. 1994;2(3):149-154.

[24] Reyes R, Pec MK, Sánchez E, et al. Comparative, osteochondral defect repair: stem cells versus chondrocytes versus bone morphogenetic protein-2, solely or in combination Eur Cell Mater. 2013;25:351-365.

[25] Wu L, Leijten J, van Blitterswijk CA, et al. Fibroblast growth factor-1 is a mesenchymal stromal cell-secreted factor stimulating proliferation of osteoarthritic chondrocytes in co-culture. Stem Cells Dev. 2013;22(17):2356-2367.

[26] Sabatino MA, Santoro R, Gueven S, et al. Cartilage graft engineering by co-culturing primary human articular chondrocytes with human bone marrow stromal cells. J Tissue Eng Regen Med. 2012 Dec 6. [Epub ahead of print]

[27] Tang X, Sheng L, Xie F, et al. Differentiation of bone marrow-derived mesenchymal stem cells into chondrocytes using chondrocyte extract. Mol Med Rep. 2012;6(4):745-749.

[28] Liao X, Wu L, Fu M, et al. Chondrogenic phenotype differentiation of bone marrow mesenchymal stem cells induced by bone morphogenetic protein 2 under hypoxic microenvironment in vitro.Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2012;26(6):743-748.

[29] Kocamaz E, Gok D, Cetinkaya A, et al. Implication of C-type natriuretic peptide-3 signaling in glycosaminoglycan synthesis and chondrocyte hypertrophy during TGF-β1 induced chondrogenic differentiation of chicken bone marrow-derived mesenchymal stem cells. J Mol Histol. 2012;43(5):497-508.

[30] 冯万文,夏亚一,孙正义,等.骨髓间充质干细胞和软骨细胞共培养复合同种异体脱钙骨基质修复关节软骨全层缺损[J].第四军医大学学报,2006,27(18):1683-1686.

[31] 张文元,杨亚冬,房国坚,等.大鼠骨髓间充质干细胞的分离冻存及向成骨细胞和软骨细胞诱导分化[J].中国现代医学杂志,2006, 16(9):1306-1309.

[32] Gronthos S, McCarty R, Mrozik K, et al. Heat shock protein-90 beta is expressed at the surface of multipotential mesenchymal precursor cells: generation of a novel monoclonal antibody, STRO-4, with specificity for mesenchymal precursor cells from human and ovine tissues. Stem Cells Dev. 2009;18(9):1253-1262.