Effects of insulin-like growth factor-induced Wharton jelly mesenchymal stem cells toward chondrogenesis in an osteoarthritis model

Document Type: Original Article


1 Medical Research Center, Faculty of Medicine, Maranatha Christian University, Bandung 40164, West Java, Indonesia

2 Biomolecular and Biomedical Research Center, Aretha Medika Utama, Bandung 40163, West Java, Indonesia

3 Research Center for Chemistry, Indonesian Institute of Sciences, Serpong, Indonesia

4 Department of Biochemistry and Molecular Biology, Faculty of Dentistry, Trisakti University, Jakarta, Indonesia

5 Stem Cell and Cancer Institute, Jakarta 13210, Indonesia


Objective(s): This study aimed to determine the collagen type II (COL2) and SOX9 expression in interleukin growth factor (IGF-1)-induced Wharton’s Jelly mesenchymal stem cells (WJMSCs) and the level of chondrogenic markers in co-culture IGF1-WJMSCs and IL1β-CHON002 as osteoarthritis (OA) cells model.
Materials and Methods: WJMSCs were induced with IGF1 (75, 150, and 300 ng/ml) to enhance their chondrogenesis capability. The gene expression of SOX9 and COL2 was evaluated with quantitative RT-PCR. Furthermore, IGF1-WJMSCs were co-cultured with IL1β-CHON002 cells in varied ratios (1:2, 1:1, 2:1). Chondrogenic markers ADAMTS1, ADAMTS5, MMP3, MMP1, and RANKL were measured with ELISA.
Results: The IGF1-WJMSCs had an increased expression of COL2 and SOX9. ADAMTS1, ADAMTS5, MMP1, MMP3, and RANKL levels were decreased in the co-culture IGF1-WJMSCs and IL1β-CHON002.
Conclusion: The IGF1-induced WJMSCs were capable to enhance chondrogenesis, indicated by increased expression of SOX9 and COL2 and decreased expression of ADAMTS1, ADAMTS5, MMP3, MMP1, and RANKL. These findings can be further used in the osteoarthritis treatment.


1. Bijlsma JW, Berenbaum F, Lafeber FP. Osteoarthritis: an update with relevance for clinical practice. Lancet 2010; 377: 2115-2126.
2. Anderson DD, Chubinskaya S, Guilak F. Post-traumatic osteoarthritis: improved understanding and opportunities for early intervention. J Orthop Res 2011; 29: 802-809.
3. Van den Berg WB. Osteoarthritis year 2010 in review: pathomechanisms. Osteoarthr Cartilage 2010; 19: 338-341.
4. Aigner T, Gebhard PM, Schmid E, Bau B, Harley V, Poschl E. SOX9 expression does no correlate with type II collagen expression in adult articular chondrocytes. Matrix Biol 2003; 22: 363-372.
5. Ashraf A, Cha BH, Kim JS, Ahn J, Han I, Park H, Lee SH. Osteoarthritis and Cartilage. Osteoarthritis  Cartilage 2016; 24:196-205.
6. Buckwalter JA, Saltzman C, Brown T. The impact of osteoarhtritis: implications for research. Clin Orthop Relat Res 2004; S6: 15.
7. Aigner T, McKenna L. Molecular pathology and pathobiology of osteoarthritic cartilage. Cell Mol Life Sci 2012; 59: 5-18.
8. Johnson K, Zhu S, Tremblay MS. A stem cell-based approach to cartilage repair. Science 2012; 336: 717-721.
9. Haleem-Smith H, Calderon R, Song Y. Cartilage oligomeric matrix protein enhances matrix assembly during chondrogenesis of human mesenchymal stem cells. J Cell Biochem 2012; 113: 1245-1252.
10. Hass R, Kasper C, Bohm S, Jacobs R. Different populations and sources of human mesenchymal stem cells (MSC): a comparison of adult and neonatal tissue-derived MSC. Cell Commun Signal 2011; 9: 12-25.
11. Widowati W, Laura W, Bachtiar I. Effect of oxygen tension towards proliferation and characteristics of wharton’s jelly-derived mesenchymal stem cells. BGM 2014; 7: 1-8.
12. Kim HJ, Im GI. Chondrogenic differentiation of adipose tissue-derived mesenchymal stem cells: Greater doses of growth factor are necessary. J Orthop Res 2009; 27: 612-619.
13. Guenther HL, Guenther HE, Froesch ER, Fleisch H. Effect of insulin-like growth factor on collagen and glycosaminoglycan synthesis by rabbit articular chondrocytes in culture. Experientia 1982; 38: 979-981.
14. McQuillan DJ, Handley CJ, Campbell MA, Bolis S, Milway VE, Herington AC. Stimulation of proteoglycan biosynthesis by serum and insulin-like growth factor-I in cultured bovine articular cartilage. Biochem J 1986; 240: 423-430.
15. Schoenle E, Zapf J, Humbel RE, Froesch ER. Insulin-like growthfactor I stimulates growth in hypophysectomized rats. Nature 1982; 296: 252-253.
16. Trippel SB, Corvol MT, Dumontier MF, Rappaport R, Hung HH, Mankin HJ. Effect of somatomedin-C/insulin-like growth factor I and growth hormone on cultured growth plate and articular chondrocytes. Pediatr Res 1989; 25: 76-82.
17. Bhaumick B, Bala RM. Differential effects of insulin-like growth factors I and II on growth, differentiation and glucoregulation in differentiating chondrocyte cells in culture. Acta Endocrinol 1991; 125: 201-211.
18. Bhaumick B. Insulin-like growth factor (IGF) binding proteins and insulin-like growth factor secretion by cultured chondrocyte cells: Identification, characterization and ontogeny during cell differentiation. Regul Pept 1993; 48: 113-122.
19. Madry H, Kaul G, Gucchiarini M, Stein U. Enhanced repair of articular cartilage defects in vivo by transplanted chondrocytes overexpressing insulin-like growth factor I (IGF-I). Gene Ther 2005; 12: 1171-1179.
20. Widowati W, Wijaya L, Murti H, Widyastuti H, Agustina D, Laksmitawati DR, et al. Conditioned medium from normoxia (WJMSCs-norCM) and hypoxia-treated WJMSCs (WJMSCs-hypoCM) in inhibiting cancer cell proliferation. Biomarkers and Genomic Medicine 2015; 7: 8-17.
21. Tsuchiya K, Chen G, Ushida T, Matsuno T, Tateishi T. The effect of coculture of chondrocytes with mesenchymal stem cells on their cartilaginous phenotype in vitro. Materials Science and Engineering: C 2004; 24: 391-396.
22. Ng LJ, Wheatley S, Muscat GE, Conway-Campbell J, Bowles J, Wright E. SOX9 binds DNA, activates transcription, and coexpresses with type II collagen during chondrogenesis in the mouse. Dev Biol 1997; 183: 108e21.
23. Zhao Q, Eberspaecher H, Lefebvre V, deCrombrugghe B. Parallel expression of Sox9 and Col2a1 in cells undergoing chondrogenesis. Dev Dyn 1997; 209: 377e86.
24. Kelwick R, Desanlis I, Wheeler GN, Edwards DR. The ADAMTS (A Disintegrin and Metalloproteinase with Thrombospondin motifs) family. Genome Biol 2015; 16: 113-128.
25. Gardiner MD, Vincent TL, Driscoll C, Burleigh A, Bou-Gharios G, Saklatvala J. Transcriptional analysis of micro-dissected articular cartilage in post-traumatic murine osteoarthritis. Osteoarthritis and Cartilage 2015; 23: 616-628.
26. Seo S, Na K. Mesenchymal stem cell-based tissue engineering for chondrogenesis. J Biomed Biotechnol 2011: 1-8.
27. Can A, Karahuseyinoglu S. Concise review: Human umbilical cord stroma with regard to the source of fetus – derived stem cells. Stem Cells 2007; 25: 2886-2895.
28. Sekiya I, Tsuji K, Koopman P, Watanabe H, Yamada Y, Shinomiya K, et al. SOX9 enhances aggrecan gene promoter/enhancer activity and is up-regulated by retinoic acid in a cartilage-derived cell line, TC6. J Biol Chem 2000; 275: 10738-10744.
29. Zhang P, Jimenez SA, Stokes DG. Regulation of human COL9A1 gene expression. Activation of the proximal promoter region by SOX9. J Biol Chem 2003; 278: 117-123.
30. Bridgewater LC, Lefebvre V, de Crombrugghe B. Chondrocyte-specific enhancer elements in the Col11a2 gene resemble the Col2a1 tissue-specific enhancer. J Biol Chem 1998; 273: 14998-15006.
31. Kou I, Ikegawa S. SOX9-dependent and -independent transcriptional regulation of human cartilage link protein. J Biol Chem 2004; 279: 50942-50948.
32. Wagner T, Wirth J, Meyer J, Zabel B, Held M, Zimmer J, et al. Autosomal sex reversal and campomelic dysplasia are caused by mutations in and around the SRY-related gene SOX9. Cell 1994; 79: 1111-1120.
33. Li Y, Tew SR, Russell AM, Gonzalez K, Hardingham TE, Hawkins RE. Transduction of human articular chondrocytes with adenoviral, retroviral and lentiviral vectors and the effects of enhanced expression of SOX9. Tissue Eng 2004; 10: 575-584.
34. Tew SR, Li Y, Pothacharoen P, Tweats LM, Hawkins RE, Hardingham TE. Retroviral transduction with SOX9 enhances re-expression of the chondrocyte phenotype in passaged osteoarthritic human articular chondrocytes. Osteoarthritis Cartilage 2005; 13: 80-89.
35. Lefebvre V, Huang W, Harley VR, Goodfellow PN, de Crombrugghe B. SOX9 is a potent activator of the chondrocyte-specific enhancer of the pro alpha1 (II) collagen gene. Mol Cell Biol 1997; 17: 2336-2346.
36. Catterall JB, Carrere S, Koshy PJ. Synergistic induction of matrix metalloproteinase 1 by interleukin-1alpha and oncostatin M in human chondrocytes involves signal transducer and activator of transcription and activator protein 1 transcription factors via a novel mechanism. Arthritis Rheum 2001; 44: 2296-2310.
37. Stremme S, Duerr S, Bau B, Schmid E, Aigner T. MMP-8 is only a minor gene product of human adult articular chondrocytes of the knee. Clin Exp Rheumatol 2003; 21: 205-209.
38. Neuhold LA, Killar L, Zhao W. Postnatal expression in hyaline cartilage of constitutively active human collagenase-3 (MMP-13) induces osteoarthritis in mice. J Clin Invest 2001; 107: 35-44.
39. Maiotti M, Monteleone G, Tarantino U, Fasciglion EG, Marini S, Coletta M. Correlation between osteoarthritic cartilage damage and levels of proteinases and proteinase inhibitors in synovial fluid from the knee joint. Arthroscopy 2000; 16: 522-526.
40. MarteL-Pelletier J, McCollum R, Fujimoto N, Obata K, Cloutier JM, Pelletier JP. Excess of metalloproteases over tissue inhibitor of metalloprotease may contribute to cartilage degradation in osteoarthritis and rheumatoid arthritis. Lab Invest 1994; 70: 807-815.
41. Franchimont P, Bassler C. Effects of hormones and local growth factors on articular chondrocyte metabolism. J Rheumatol 1991; 18 (Suppl 27): 68-70.
42. Sandell LJ, Dudek EJ. Insulin-like growth factor I stimulates type II collagen gene expression in cultured chondrocytes. Trans Orthop Res Soc 1988; 35: 300.
43. Tesch GH, Handley CJ, Cornell HJ, Herington AC. Effects of free and bound insulin-like growth factors on proteoglycan metabolism in articular cartilage explants. J Orthop Res 1992; 10: 14-22.
44. Kim HJ, Im GL. Chondrogenic Differentiation of Adipose Tissue-Derived MesenchymalStem Cells: Greater Doses of Growth Factor are Necessary. J Orthop Res 2009; 27:612-619.
45. Dean PW, Nelson JK, Schumacher J. Effects of age and prosthesis material on in vitro cartilage retention of laryngoplasty prostheses in horses. American Journal of Veterinary Research 1990; 51: 114-117.
46. Su S, Grover J, Roughley PJ, DiBattista JA, Martel-Pelletier J, Pelletier JP, et al. Expression of the tissue inhibitor of metalloproteinases (TIMP) gene family in normal and osteoarthritic joints. Rheumatology International 1999; 18: 183-191.
47. Clark IM, Powell LK, Ramsey S, Hazleman BL, Cawston TE. The measurement of collagenase, tissue inhibitor of metalloproteinases (timp), and collagenase—timp complex in synovial fluids from patients with osteoarthritis and rheumatoid arthritis. Arthritis and Rheumatology 1993; 36: 372-379.
48. Cawston TE. Proteinases and inhibitors. British medical bulletin 1995; 51: 385-401.
49. Freemont AJ, Hampson V, Tilman R, Goupille P, Taiwo Y, Hoyland JA. Gene expression of matrix metalloproteinases 1, 3, and 9 by chondrocytes in osteoarthritic human knee articular cartilage is zone and grade specific. Annals of The Rheumatic Diseases 1997; 56: 542-548.
50. Shlopov BV, Gumanovskaya ML, Hasty KA. Autocrine regulation of collagenase 3 (matrix metalloproteinase 13) during osteoarthritis. Arthritis and Rheumatism 2000; 43: 195-205.
51. Freemont AJ, Byers RJ, Taiwo YO, Hoyland JA. In situ zymographic localisation of type II collagen degrading activity in osteoarthritic human articular cartilage. Annals of the rheumatic diseases 1999; 58: 357-365.
52. Billinghurst RC, Dahlberg L, Lonescu M, Reiner A, Bourne R, Rorabeck C. Enhanced cleavage of type II collagen by collagenases in osteoarthritic articular cartilage. J Clin Invest 1997; 99: 1534-1545.
53. Rodríguez-Manzaneque JC, Westling J, Thai SN, Luque A, Knauper V, Murphy G. ADAMTS1 cleaves aggrecan at multiple sites and is differentially inhibited by metalloproteinase inhibitors. Biochem Biophys Res Commun 2002; 293: 501-508.
54. Geyer M, Grassel S, Straub RH, Schett G, Dinser R, Grifka J. Differential transcriptome analysis of intraarticular lesional vs intact cartilage reveals new candidate genes in osteoarthritis pathophysiology. Osteoarthritis and Cartilage, 2009; 17: 328-335.
55. Geyer M, Grassel S, Straub RH, Schett G, Dinser R, Grifka J. Differential transcriptome analysis of intraarticular lesional vs intact cartilage reveals new candidate genes in osteoarthritis pathophysiology. Osteoarthritis and Cartilage, 2009; 17: 328-335.
56. Karlsson C, Dehne T, Lindahl A, Brittberg M, Pruss A, Sittinger M. Genome-wide expression profiling reveals new candidate genes associated with osteoarthritis. Osteoarthritis and Cartilage, 2010; 18: 581-592.
57. Ramos YF, den Hollander W, Bovee JV, Bomer N, Van der Breggen R, Lakenberg N. Genes involved in the osteoarthritis process identified through genome wide expression analysis in articular cartilage; the RAAK study. PLoS One, 2014; 9: e103056.
58. Wachsmuth L, Bau B, Fan Z, Pecht A, Gerwin N, Aigner T. ADAMTS-1, a gene product of articular chondrocytes in vivo and in vitro, is downregulated by interleukin 1beta. J Rheumatol 2004; 31: 315-320.
59. Davidson RK, Waters JG, Kevorkian L, Darrah C, Cooper A, Donell ST. Expression profiling of metalloproteinases and their inhibitors in synovium and cartilage. Arthritis Res Ther 2006; 8: R124-R133.
60. Kevorkian L, Young DA, Darrah C, Donell ST, Shepstone L, Porter S. Expression profiling of metalloproteinases and their inhibitors in cartilage. Arthritis Rheum 2004; 50: 131-141.
61. Swingler TE, Waters JG, Davidson RK, Pennington CJ, Puente XS, Darrah C. Degradome expression profiling in human articular cartilage. Arthritis Res Ther 2009; 11: R96.
62. Lacey DL, Timms E, Tan HL, Kelley MJ, Dunstan CR, Burgess T. Osteoprotegerin ligand is a cytokine that regulates osteoclast differentiation and activation. Cell 1998; 93: 165-176.
63. Kong YY, Feige U, Sarosi I, Bolon B, Tafuri A, Morony S. Activated T cells regulate bone loss and joint destruction in adjuvant arthritis through osteoprotegerin ligand. Nature 1999; 402: 304-309.
64. Simonet WS, Lacey DL, Dunstan CR, Kelley M, Chang MS, Luthy R. Osteoprotegerin: a novel secreted protein involved in the regulation of bone density. Cell 1997; 89: 309-319.
65. Yasuda H, Shima N, Nakagawa N, Yamaguchi K, Kinosaki M, Mochizuki S. Osteoclast differentiation factor is a ligand for osteoprotegerin/osteoclastogenesis-inhibitory factor and is identical to TRANCE/RANKL. Proc Natl Acad Sci USA 1998; 95: 3597-3602.
66. Wong BR, Rho J, Arron J, Robinson E, Orlinick J, Chao M. TRANCE is a novel ligand of the tumor necrosis factor receptor family that activates c-Jun N-terminal kinase in T cells. J Biol Chem 1997; 272: 25190-25194.
67. Kartsogiannis V, Zhou H, Horwood NJ, Thomas RJ, Hards DK, Quinn JMLoalization of RANKL (receptor activator of NFkB ligand) mRNA and protein in skeletal and extraskeletal tissues. Bone 1999; 25: 525-534.
68. Roodman GD. Cell biology of the osteoclast. Exp Hematol. 1999; 27: p. 1229-1241.
69. Ahmed N, Dreier R, Göpferich A, Grifka J, Grässel S. Soluble signalling factors derived from differentiated cartilage tissue affect chondrogenic differentiation of rat adult marrow stromal cells. Cell Physiol Biochem 2007; 20: 665-678.
70. Bian L, Zhai DY, Mauck RL, Burdick JA. Coculture of human mesenchymal stem cells and articular chondrocytes reduces hypertrophy and enhances functional properties of engineered cartilage. Tissue Eng A 2011; 17: 1137-1145.
71. Babur BK, Kabiri M, Klein TJ, Lott WB, Doran MR. The rapid manufacture of uniform composite multicellular-biomaterial micropellets, their assembly into macroscopic organized tissues, and potential applications in cartilage tissue engineering. PLoS One 2015; 10: e0122250.