Inhibition of transforming growth factor-β type II receptor signaling accelerates tooth formation in mouse first branchial arch explants

Abstract

Members of the transforming growth factor-beta (TGF-beta) superfamily signal through their cognate receptors to determine cell phenotypes during embryogenesis. Our previous studies on the regulation of first branchial arch morphogenesis have identified critical components of a hierarchy of different TGF-beta isoforms and their possible functions in regulating tooth and cartilage formation during mandibular morphogenesis. Here we tested the hypothesis that TGF-beta type II receptor (TGF-beta IIR) is a critical component in the TGF-beta signaling pathway regulating tooth formation. To establish the precise location of TGF-beta ligand and its cognate receptor, we first performed detailed analyses of the localization of both TGF-beta2 and TGF-beta IIR during initiation and subsequent morphogenesis of developing embryonic mouse tooth organs. A possible autocrine functional role for TGF-beta and its cognate receptor (TGF-beta IIR) was inferred due to the temporal and spatial localization patterns during the early inductive stages of tooth morphogenesis. Second, loss of function of TGF-beta IIR in a mandibular explant culture model resulted in the acceleration of tooth formation to the cap stage while the mandibular explants in the control group only showed bud stage tooth formation. In addition, there was a significant increase in odontogenic epithelial cell proliferation following TGF-beta IIR abrogation. These results demonstrate, for the first time, that abrogation of the TGF-beta IIR stimulates embryonic tooth morphogenesis in culture and reverses the negative regulation of endogenous TGF-beta signaling upon enamel organ epithelial cell proliferation.

Inhibition of SMAD2 expression prevents murine palatal fusion

“…Immunostaining was completed by following standard procedures (Chai et al, 1999;Hosokawa et al, 2005). Briefly, serial sections (6 m) of cultured palates transfected with control or Smad2 siRNA were cut and mounted onto the same slide to ensure exposure to the same antibody concentration.…”

Receptor‐regulated and inhibitory Smads are critical in regulating transforming growth factorβ–mediated Meckel's cartilage development

“…TGF-␤ and its cognate receptors are important regulators during mandibular morphogenesis (Pelton et al, 1990;Chai et al, 1994Chai et al, , 1999. Targeted disruption of TGF-␤ signaling (such as TGF-␤2 null mutant) results in multiple developmental defects, including a small mandible with its angle missing.…”

“…This AdSmad7 has been used successfully to block the phosphorylation of Smad2 upon TGF-␤ stimulation in various studies, demonstrating its effectiveness as a negative TGF-␤ signaling regulator (Nakao et al, 1999;Zhao et al, 2000). Our previous study has shown that adenoviral vectors are very effective in altering TGF-␤ type II receptor gene expression in cultured mandibular explants (Chai et al, 1999).…”

“…Seventy-five micrograms of total protein from each sample was loaded in each well on a 12% polyacrylamide gel. Western analysis was done as previously described (Chai et al, 1999).…”

“…Competitive PCR has been shown to successfully measure the change in mRNA expression level of mandibular explants after adenovirus treatment (Chai et al, 1999). Primers were designed based on murine Smad7 and Smad6 cDNA sequence and have been demonstrated to detect variation of Smad7 and Smad6 expression level after adenovirus treatment during lung morphogenesis (Zhao et al, 2000).…”