Scleraxis upregulated by transforming growth factor-ß1 signaling inhibits tension-induced osteoblast differentiation of priodontal ligament cells via ephrin A2.

During tooth movement in orthodontic treatment, bone formation and resorption occur on the tension and compression sides of the alveolar bone, respectively. Although the bone formation activity increases in the periodontal ligament (PDL) on the tension side, the PDL itself is not ossified and maintains its homeostasis, indicating that there are negative regulators of bone formation in the PDL. Our previous report suggested that scleraxis (Scx) has an inhibitory effect on ossification of the PDL on the tension side through the suppression of calcified extracellular matrix formation. However, the molecular biological mechanisms of Scx-modulated inhibition of ossification in the tensioned PDL are not fully understood. The aim of the present study is to clarify the inhibitory role of Scx in osteoblast differentiation of PDL cells and its underlying mechanism. Our in vivo experiment using a mouse experimental tooth movement model showed that Scx expression was increased during early response of the PDL to tensile force. Scx knockdown upregulated expression of alkaline phosphatase, an early osteoblast differentiation marker, in the tensile force-loaded PDL cells in vitro. Transforming growth factor (TGF)-ß1-Smad3 signaling in the PDL was activated by tensile force and inhibitors of TGF-ß receptor and Smad3 suppressed the tensile force-induced Scx expression in PDL cells. Tensile force induced ephrin A2 (Efna2) expression in the PDL and Efna2 knockdown upregulated alkaline phosphatase expression in PDL cells under tensile force loading. Scx knockdown eliminated the tensile force-induced Efna2 expression in PDL cells. These findings suggest that the TGF-ß1-Scx-Efna2 axis is a novel molecular mechanism that negatively regulates the tensile force-induced osteoblast differentiation of PDL cells.

Ten-m/Odz3 regulates migration and differentiation of chondrogenic ATDC5 cells via RhoA-mediated actin reorganization.

Tenascin-like molecule major (Ten-m)/odd Oz (Odz), a type II transmembrane molecule, is well known to modulate neural development. We have reported that Ten-m/Odz3 is expressed in cartilaginous tissues and cells. Actin cytoskeleton and its regulator ras homolog gene family member A (RhoA) are closely associated with chondrogenesis. The present study aimed to evaluate the function and molecular mechanism of Ten-m/Odz3 during chondrogenesis, focusing on RhoA and the actin cytoskeleton. Ten-m/Odz3 was expressed in precartilaginous condensing mesenchyme in mouse limb buds. Ten-m/Odz3 knockdown in ATDC5 induced actin cytoskeleton reorganization and change of cell shape through modulation of RhoA activity and FGF2 expression. Ten-m/Odz3 knockdown suppressed ATDC5 migration and expression of genes associated with chondrogenesis, such as Sox9 and type II collagen, via RhoA. On the other hand, Ten-m/Odz3 knockdown inhibited proliferation of ATDC5 in a RhoA-independent manner. These findings suggest that Ten-m/Odz3 plays an important role in early chondrogenesis regulating RhoA-mediated actin reorganization.

Scleraxis and osterix antagonistically regulate tensile force-responsive remodeling of the periodontal ligament and alveolar bone.

The periodontal ligament (PDL) is a mechanosensitive noncalcified fibrous tissue connecting the cementum of the tooth and the alveolar bone. Here, we report that scleraxis (Scx) and osterix (Osx) antagonistically regulate tensile force-responsive PDL fibrogenesis and osteogenesis. In the developing PDL, Scx was induced during tooth eruption and co-expressed with Osx. Scx was highly expressed in elongated fibroblastic cells aligned along collagen fibers, whereas Osx was highly expressed in the perialveolar/apical osteogenic cells. In an experimental model of tooth movement, Scx and Osx expression was significantly upregulated in parallel with the activation of bone morphogenetic protein (BMP) signaling on the tension side, in which bone formation compensates for the widened PDL space away from the bone under tensile force by tooth movement. Scx was strongly expressed in Scx(+)/Osx(+) and Scx(+)/Osx(-) fibroblastic cells of the PDL that does not calcify; however, Scx(-)/Osx(+) osteogenic cells were dominant in the perialveolar osteogenic region. Upon BMP6-driven osteoinduction, osteocalcin, a marker for bone formation was downregulated and upregulated by Scx overexpression and knockdown of endogenous Scx in PDL cells, respectively. In addition, mineralization by osteoinduction was significantly inhibited by Scx overexpression in PDL cells without affecting Osx upregulation, suggesting that Scx counteracts the osteogenic activity regulated by Osx in the PDL. Thus, Scx(+)/Osx(-), Scx(+)/Osx(+) and Scx(-)/Osx(+) cell populations participate in the regulation of tensile force-induced remodeling of periodontal tissues in a position-specific manner.