Open AccessRegulation of CCN2/Connective tissue growth factor expression in the nucleus pulposus of the intervertebral disc: Role of Smad and activator protein 1 signaling
Author(s) -
Tran Cassie M.,
Markova Dessislava,
Smith Harvey E.,
Susarla Bala,
Ponnappan Ravi Kumar,
Anderson D. Greg,
Symes Aviva,
Shapiro Irving M.,
Risbud Makarand V.
Publication year - 2010
Publication title -
arthritis & rheumatism
Language(s) - English
Resource type - Journals
eISSN - 1529-0131
pISSN - 0004-3591
DOI - 10.1002/art.27445
Subject(s) - ctgf , smad , growth factor , connective tissue , activator (genetics) , western blot , nucleus , transforming growth factor beta , transcription factor , transforming growth factor , biology , chemistry , microbiology and biotechnology , endocrinology , medicine , gene , receptor , genetics
Abstract Objective To investigate transforming growth factor β (TGFβ) regulation of connective tissue growth factor (CTGF) expression in cells of the nucleus pulposus of rats, mice, and humans. Methods Real‐time reverse transcription–polymerase chain reaction and Western blot analyses were used to measure CTGF expression in the nucleus pulposus. Transfections were used to measure the effects of Smads 2, 3, and 7 and activator protein 1 (AP‐1) on TGFβ‐mediated CTGF promoter activity. Results CTGF expression was lower in neonatal rat discs than in skeletally mature rat discs. An increase in CTGF expression and promoter activity was observed in rat nucleus pulposus cells after TGFβ treatment. Deletion analysis indicated that promoter constructs lacking Smad and AP‐1 motifs were unresponsive to treatment. Analysis showed that full‐length Smad3 and the Smad3 MH‐2 domain alone increased CTGF activity. Further evidence of Smad3 and AP‐1 involvement was seen when DN‐Smad3, SiRNA‐Smad3, Smad7, and DN‐AP‐1 suppressed TGFβ‐mediated activation of the CTGF promoter. When either Smad3 or AP‐1 sites were mutated, CTGF promoter induction by TGFβ was suppressed. We also observed a decrease in the expression of CTGF in discs from Smad3‐null mice as compared with those from wild‐type mice. Analysis of human nucleus pulposus samples indicated a trend toward increasing CTGF and TGFβ expression in the degenerated state. Conclusion TGFβ, through Smad3 and AP‐1, serves as a positive regulator of CTGF expression in the nucleus pulposus. We propose that CTGF is a part of the limited reparative response of the degenerated disc.