Aberrant hypertrophy in Smad3‐deficient murine chondrocytes is rescued by restoring transforming growth factor β–activated kinase 1/activating transcription factor 2 signaling: A potential clinical implication for osteoarthritis

Abstract

AbstractObjectiveTo investigate the biologic significance of Smad3 in the progression of osteoarthritis (OA), the crosstalk between Smad3 and activating transcription factor 2 (ATF‐2) in the transforming growth factor β (TGFβ) signaling pathway, and the effects of ATF‐2 overexpression and p38 activation in chondrocyte differentiation.MethodsJoint disease in Smad3‐knockout (Smad3−/−) mice was examined by microfocal computed tomography and histologic analysis. Numerous in vitro methods including immunostaining, real‐time polymerase chain reaction, Western blotting, an ATF‐2 DNA‐binding assay, and a p38 kinase activity assay were used to study the various signaling responses and protein interactions underlying the altered chondrocyte phenotype in Smad3−/− mice.ResultsIn Smad3−/− mice, an end‐stage OA phenotype gradually developed. TGFβ‐activated kinase 1 (TAK1)/ATF‐2 signaling was disrupted in Smad3−/− mouse chondrocytes at the level of p38 MAP kinase (MAPK) activation, resulting in reduced ATF‐2 phosphorylation and transcriptional activity. Reintroduction of Smad3 into Smad3−/− cells restored the normal p38 response to TGFβ. Phosphorylated p38 formed a complex with Smad3 by binding to a portion of Smad3 containing both the MAD homology 1 and linker domains. Additionally, Smad3 inhibited the dephosphorylation of p38 by MAPK phosphatase 1 (MKP‐1). Both ATF‐2 overexpression and p38 activation repressed type X collagen expression in wild‐type and Smad3−/− chondrocytes. P38 was detected in articular cartilage and perichondrium; articular and sternal chondrocytes expressed p38 isoforms α, β, and γ, but not δ.ConclusionSmad3 is involved in both the onset and progression of OA. Loss of Smad3 abrogates TAK1/ATF‐2 signaling, most likely by disrupting the Smad3–phosphorylated p38 complex, thereby promoting p38 dephosphorylation and inactivation by MKP‐1. ATF‐2 and p38 activation inhibit chondrocyte hypertrophy. Modulation of p38 isoform activity may provide a new therapeutic approach for OA.