EFFECT OF OSTEOPONTIN ON EXPRESSION OF MATRIX METALLOPROTEINASE 13 IN HUMAN KNEE OSTEOARTHRITIC CHONDROCYTES_Chinese Journal of Reparative and Reconstructive Surgery

Authors：

 JIANGWei ¹ , LEIGuanghua ² , LINBowen ¹ , WANGHua ¹ , LüMeng ¹ , GAOShuguang ² , ZHANGFangjie ² , ZENGChao ²

1. Department of Bone and Joint, Shenzhen People's Hospital, Second Clinical Medical College of Jinan University, Shenzhen Guangdong, 518020, P. R. China;
2. Department of Orthopaedics, Xiangya Hospital, Central South University;

Corresponding author：

JIANGWei, Email: jw8509@163.com

Keywords：

Osteopontin; Matrix metalloproteinase 13; Osteoarthritis; Chondrocyte

DOI：

10.7507/1002-1892.20140291

Video：

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Objective To investigate the role of osteopontin (OPN) on the expressions of matrix metalloproteinase 13 (MMP-13) mRNA and protein in human knee osteoarthritic chondrocytes and to find the optimal time and concentration for OPN treatment. Methods Chondrocytes were isolated from articular cartilage tissues of patients with primary osteoarthritis (OA) and cultured using one step digestive treatment with collagenase typeⅡ. The chondrocytes were then identified using immunohistochemistry of collagen typeⅡ. The first generation of chondrocytes were stimulated with OPN at a concentration of 1μg/mL for 0, 24, 48, and 72 hours, and with OPN at the concentrations of 0, 0.5, 1, 2, and 4μg/mL for 48 hours. The levels of MMP-13 mRNA and protein expressions were measured with real-time fluorescent quantitative PCR and Western blot. Results The immunohistochemical staining showed that first generation of chondrocytes expressed collagen typeⅡ. Both MMP-13 mRNA and protein expression levels in OA chondrocytes increased significantly in the presence of OPN (1μg/ mL) and peaked at 48 hours after incubation, showing significant difference between different time points (P < 0.05). The MMP-13 mRNA expression level in OA chondrocytes at the OPN concentration of 1μg/mL was significantly higher than those at the other concentrations (P < 0.05), and the MMP-13 protein expression level at the OPN concentration of 1μg/mL was significantly higher than that at 0μg/mL (P < 0.05). MMP-13 protein expression level at the OPN concentrations of 0.5, 2, and 4μg/mL were significantly higher than that at 0μg/mL (P < 0.05). Conclusion OPN induces up-regulation of MMP-13 mRNA and protein expressions in human knee osteoarthritic chondrocytes in time-and dose-dependent manners. The optimal time and concentration for OPN treatment are 48 hours and 1μg/mL, respectively.

Citation： JIANGWei, LEIGuanghua, LINBowen, WANGHua, LüMeng, GAOShuguang, ZHANGFangjie, ZENGChao. EFFECT OF OSTEOPONTIN ON EXPRESSION OF MATRIX METALLOPROTEINASE 13 IN HUMAN KNEE OSTEOARTHRITIC CHONDROCYTES. Chinese Journal of Reparative and Reconstructive Surgery, 2014, 28(11): 1342-1345. doi: 10.7507/1002-1892.20140291 Copy

1.	Wieland HA, Michaelis M, Kirschbaum BJ, et al. Osteoarthritis-an untreatable disease? Nat Rev Drug Discov, 2005, 4(4):331-344.
2.	Lawrence RC, Felson DT, Helmick CG, et al. Estimates of the prevalence of arthritis and other rheumatic conditions in the United States. PartⅡ. Arthritis Rheum, 2008, 58(1):26-35.
3.	Bijlsma JW, Berenbaum F, Lafeber FP. Osteoarthritis:an update with relevance for cl inical practice. Lancet, 2011, 377(9783):2115-2126.
4.	Heinegård D, Saxne T. The role of the cartilage matrix in osteoarthritis. Nat Rev Rheumatol, 2011, 7(1):50-56.
5.	Troeberg L, Nagase H. Proteases involved in cartilage matrix degradation in osteoarthritis. Biochim Biophys Acta, 2012, 1824(1):133-145.
6.	Gao SG, Li KH, Zeng KB, et al. Elevated osteopontin level of synovial fluid and articular cartilage is associated with disease severity in knee osteoarthritis patients. Osteoarthritis Cartilage, 2010, 18(1):82-87.
7.	Honsawek S, Tanavalee A, Sakdinakiattikoon M, et al. Correlation of plasma and synovial fluid osteopontin with disease severity in knee osteoarthritis. Clin Biochem, 2009, 42(9):808-812.
8.	Hasegawa M, Segawa T, Maeda M, et al. Thrombin-cleaved osteopontin levels in synovial fluid correlate with disease severity of knee osteoarthritis. J Rheumatol, 2011, 38(1):129-134.
9.	Neuhold LA, Killar L, Zhao W, et al. Postnatal expression in hyaline cartilage of constitutively active human collagenase-3(MMP-13) induces osteoarthritis in mice. J Clin Invest, 2001, 107(1):35-44.
10.	Kevorkian L, Young DA, Darrah C, et al. Expression profil ing of metalloproteinases and their inhibitors in cartilage. Arthritis Rheum, 2004, 50(1):131-141.
11.	Little CB, Barai A, Burkhardt D, et al. Matrixmetalloproteinase 13-deficient mice are resistant to osteoarthritic cartilage erosion but not chondrocyte hypertrophy or osteophyte development. Arthritis Rheum, 2009, 60(12):3723-3733.
12.	Roach HI, Yamada N, Cheung KS, et al. Association between the abnormal expression of matrix-degrading enzymes by human osteoarthritic chondrocytes and demethylation of specific CpG sites in the promoter regions. Arthritis Rheum, 2005, 52(10):3110-3124.
13.	Muddasani P, Norman JC, Ellman M, et al. Basic fibroblast growth factor activates the MAPK and NFkappaB pathways that converge on Elk-1 to control production of matrix metalloproteinase-13 by human adult articular chondrocytes. J Biol Chem, 2007, 282(43):31409-31421.
14.	Burrage PS, Mix KS, Brinckerhoff CE. Matrix metalloproteinases:role in arthritis. Front Biosci, 2006, 11:529-543.
15.	Miwa HE, Gerken TA, Huynh TD, et al. Mammalian expression of full-length bovine aggrecan and link protein:formation of recombinant proteoglycan aggregates and analysis of proteolytic cleavage by ADAMTS-4 and MMP-13. Biochim Biophys Acta, 2006, 1760(3):472-486.
16.	Mori N, Majima T, Iwasaki N, et al. The role of osteopontin in tendon tissue remodeling after denervation-induced mechanical stress deprivation. Matrix Biol, 2007, 26(1):42-53.
17.	Berge G, Pettersen S, Grotterød I, et al. Osteopontin-an important downstream effector of S100A4-mediated invasion and metastasis. Int J Cancer, 2011, 129(4):780-790.
18.	Matsui Y, Iwasaki N, Kon S, et al. Accelerated development of agingassociated and instabil ity-induced osteoarthritis in osteopontindeficient mice. Arthritis Rheum, 2009, 60(8):2362-2371.

1. Wieland HA, Michaelis M, Kirschbaum BJ, et al. Osteoarthritis-an untreatable disease? Nat Rev Drug Discov, 2005, 4(4):331-344.
2. Lawrence RC, Felson DT, Helmick CG, et al. Estimates of the prevalence of arthritis and other rheumatic conditions in the United States. PartⅡ. Arthritis Rheum, 2008, 58(1):26-35.
3. Bijlsma JW, Berenbaum F, Lafeber FP. Osteoarthritis:an update with relevance for cl inical practice. Lancet, 2011, 377(9783):2115-2126.
4. Heinegård D, Saxne T. The role of the cartilage matrix in osteoarthritis. Nat Rev Rheumatol, 2011, 7(1):50-56.
5. Troeberg L, Nagase H. Proteases involved in cartilage matrix degradation in osteoarthritis. Biochim Biophys Acta, 2012, 1824(1):133-145.
6. Gao SG, Li KH, Zeng KB, et al. Elevated osteopontin level of synovial fluid and articular cartilage is associated with disease severity in knee osteoarthritis patients. Osteoarthritis Cartilage, 2010, 18(1):82-87.
7. Honsawek S, Tanavalee A, Sakdinakiattikoon M, et al. Correlation of plasma and synovial fluid osteopontin with disease severity in knee osteoarthritis. Clin Biochem, 2009, 42(9):808-812.
8. Hasegawa M, Segawa T, Maeda M, et al. Thrombin-cleaved osteopontin levels in synovial fluid correlate with disease severity of knee osteoarthritis. J Rheumatol, 2011, 38(1):129-134.
9. Neuhold LA, Killar L, Zhao W, et al. Postnatal expression in hyaline cartilage of constitutively active human collagenase-3(MMP-13) induces osteoarthritis in mice. J Clin Invest, 2001, 107(1):35-44.
10. Kevorkian L, Young DA, Darrah C, et al. Expression profil ing of metalloproteinases and their inhibitors in cartilage. Arthritis Rheum, 2004, 50(1):131-141.
11. Little CB, Barai A, Burkhardt D, et al. Matrixmetalloproteinase 13-deficient mice are resistant to osteoarthritic cartilage erosion but not chondrocyte hypertrophy or osteophyte development. Arthritis Rheum, 2009, 60(12):3723-3733.
12. Roach HI, Yamada N, Cheung KS, et al. Association between the abnormal expression of matrix-degrading enzymes by human osteoarthritic chondrocytes and demethylation of specific CpG sites in the promoter regions. Arthritis Rheum, 2005, 52(10):3110-3124.
13. Muddasani P, Norman JC, Ellman M, et al. Basic fibroblast growth factor activates the MAPK and NFkappaB pathways that converge on Elk-1 to control production of matrix metalloproteinase-13 by human adult articular chondrocytes. J Biol Chem, 2007, 282(43):31409-31421.
14. Burrage PS, Mix KS, Brinckerhoff CE. Matrix metalloproteinases:role in arthritis. Front Biosci, 2006, 11:529-543.
15. Miwa HE, Gerken TA, Huynh TD, et al. Mammalian expression of full-length bovine aggrecan and link protein:formation of recombinant proteoglycan aggregates and analysis of proteolytic cleavage by ADAMTS-4 and MMP-13. Biochim Biophys Acta, 2006, 1760(3):472-486.
16. Mori N, Majima T, Iwasaki N, et al. The role of osteopontin in tendon tissue remodeling after denervation-induced mechanical stress deprivation. Matrix Biol, 2007, 26(1):42-53.
17. Berge G, Pettersen S, Grotterød I, et al. Osteopontin-an important downstream effector of S100A4-mediated invasion and metastasis. Int J Cancer, 2011, 129(4):780-790.
18. Matsui Y, Iwasaki N, Kon S, et al. Accelerated development of agingassociated and instabil ity-induced osteoarthritis in osteopontindeficient mice. Arthritis Rheum, 2009, 60(8):2362-2371.

Chinese Journal of Reparative and Reconstructive Surgery

EFFECT OF OSTEOPONTIN ON EXPRESSION OF MATRIX METALLOPROTEINASE 13 IN HUMAN KNEE OSTEOARTHRITIC CHONDROCYTES

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