SOX9 is a major negative regulator of cartilage vascularization, bone marrow formation and endochondral ossification

Abstract

SUMMARYSOX9 is a transcription factor of the SRY family that regulates sex determination, cartilage development and numerous other developmental events. In the foetal growth plate, Sox9 is highly expressed in chondrocytes of the proliferating and prehypertrophic zone but declines abruptly in the hypertrophic zone, suggesting that Sox9 downregulation in hypertrophic chondrocytes might be a necessary step to initiate cartilage-bone transition in the growth plate. In order to test this hypothesis, we generated transgenic mice misexpressing Sox9 in hypertrophic chondrocytes under the control of a BAC-Col10a1 promoter. The transgenic offspring showed an almost complete lack of bone marrow in newborns, owing to strongly retarded vascular invasion into hypertrophic cartilage and impaired cartilage resorption, resulting in delayed endochondral bone formation associated with reduced bone growth. In situ hybridization analysis revealed high levels of Sox9 misexpression in hypertrophic chondrocytes but deficiencies of Vegfa, Mmp13, RANKL and osteopontin expression in the non-resorbed hypertrophic cartilage, indicating that Sox9 misexpression in hypertrophic chondrocytes inhibits their terminal differentiation. Searching for the molecular mechanism of SOX9-induced inhibition of cartilage vascularization, we discovered that SOX9 is able to directly suppress Vegfa expression by binding to SRY sites in the Vegfa gene. Postnatally, bone marrow formation and cartilage resorption in transgenic offspring are resumed by massive invasion of capillaries through the cortical bone shaft, similar to secondary ossification. These findings imply that downregulation of Sox9 in the hypertrophic zone of the normal growth plate is essential for allowing vascular invasion, bone marrow formation and endochondral ossification.

Signaling Cascades Governing Cdc42-Mediated Chondrogenic Differentiation and Mensenchymal Condensation

“…The discrepancy in our findings could be explained by the different cell types and treatments among these studies. Consistent with our findings indicating that Sox9 is involved in TGF-b/Cdc42-mediated chondrogenesis, transgenic mice misexpressing Sox9 in hypertrophic chondrocytes reveal the features of growth plates that are similar to those of Prx1Cre; Cdc42 fl/fl mice (Hattori et al 2010). In addition, a study of TGF-b type II receptor (Tgfbr2) knockout mice (Prx1Cre; Tgfbr2 fl/f ) also reveals that Col2-expressing cells exhibited a less-organized columnar distribution than the controls, whereas hypertrophic chondrocytes are larger but decreased in expression of Col10 characterized in both Prx1Cre; Cdc42 fl/fl and Col2Cre;Cdc42 fl/fl long bones (Spagnoli et al 2007;Aizawa et al 2012;Suzuki et al 2015).…”

Endochondral Ossification for Enhancing Bone Regeneration: Converging Native Extracellular Matrix Biomaterials and Developmental Engineering In Vivo

“…27,43,57,58,[61][62][63][64][65][66][67][68][69][70][71][72][73][74][75][76] While cartilage tissue engineering strategies have focused on inducing and maintaining chondrogenic phenotypes, the induction and modulation of cartilage hypertrophy is critical to the progression of EC ossification in bone regeneration designs. 9,33,34,[37][38][39][40][41][42][43]72,[77][78][79][80][81][82][83][84][85] The key transcription factor involved in initial chondrogenesis is SOX9, whereas the primary hypertrophy associated factors are runtrelated transcription factor 2 (Runx2) and myocyte enhancer factor 2C (MEF2C). 33,34,41,78 In this critical maturation stage of EC ossification, chondrocytes experience a large increase in volume (*5-to 10-fold), and downstream proteins activated by Runx2 and MEF2C transcription factors begin to remodel the surrounding ECM.…”

CCAAT/Enhancer-binding Protein β Regulates the Repression of Type II Collagen Expression during the Differentiation from Proliferative to Hypertrophic Chondrocytes

“…Furthermore, SOX9 was reported to directly bind and activate cartilage-specific regulatory elements of Col2a1, resulting in its cartilage-specific gene expression (7,8). Although SOX9 is indispensable for chondrogenesis, it was also reported to work as a negative regulator of hypertrophic differentiation (9). Several studies have reported that Winglesstype murine mammary tumor virus integration site (10), bone morphogenetic protein 2 (BMP2) (11), parathyroid hormonerelated protein (PTHrP) (12), IL-1␤ (13), and Runt-related transcription factor 2 (RUNX2) (14) can regulate SOX9 expression or activity in arthritic chondrocytes or during chondrocyte differentiation.…”