Embryonic salivary gland dysmorphogenesis in Twisted gastrulation deficient mice.

OBJECTIVE: Mouse Twisted gastrulation gene (Twsg1) expression is found throughout embryonic development, including substantial levels in the first branchial arch that gives rise to the submandibular salivary gland (SMG). We addressed the proposition that normal Twsg1 expression is critical to normal SMG ontogenesis. DESIGN: Utilizing C57BL/6 embryos that were Twsg1-/- homozygotes, as well as wild type and heterozygote littermates, we investigated SMG development from gestational day 13 to newborn. RESULTS: Twsg1 protein is immunodetected in epithelia throughout SMG development. Twsg1-/- embryos display widely variable craniofacial phenotypes that range from normal to severe holoprosencephaly/agnathia with no mandibular arch or stomodeum. The SMG phenotypes are correlated with the external craniofacial phenotype, ranging from normal to agenesis/aplasia. CONCLUSIONS: It is evident that normal Twsg1 expression is critical for normal mouse SMG ontogenesis. Twsg1 loss of function is ultimately epistatic to the epigenome under normal physiologic conditions, but not always so. The reduced penetrance and variable expressivity seen in the SMGs of Twsg1-/- embryos is a challenging enigma.

FGF8 dose-dependent regulation of embryonic submandibular salivary gland morphogenesis.

FGF8 has been shown to play important morphoregulatory roles during embryonic development. The observation that craniofacial, cardiovascular, pharyngeal, and neural phenotypes vary with Fgf8 gene dosage suggests that FGF8 signaling induces differences in downstream responses in a dose-dependent manner. In this study, we investigated if FGF8 plays a dose-dependent regulatory role during embryonic submandibular salivary gland (SMG) morphogenesis. We evaluated SMG phenotypes of Fgf8 hypomorphic mice, which have decreased Fgf8 gene function throughout embryogenesis. We also evaluated SMG phenotypes of Fgf8 conditional mutants in which Fgf8 function has been completely ablated in its expression domain in the first pharyngeal arch ectoderm from the time of arch formation. Fgf8 hypomorphs have hypoplastic SMGs, whereas conditional mutant SMGs exhibit ontogenic arrest followed by involution and are absent by E18.5. SMG aplasia in Fgf8 ectoderm conditional mutants indicates that FGF8 signaling is essential for the morphogenesis and survival of Pseudoglandular Stage and older SMGs. Equally important, the presence of an initial SMG bud in Fgf8 conditional mutants indicates that initial bud formation is FGF8 independent. Mice heterozygous for either the Fgf8 null allele (Fgf8(+/N)) or the hypomorphic allele (Fgf8(+/H)) have SMGs that are indistinguishable from wild-type (Fgf8(+/+)) mice which suggest that there is not only an FGF8 dose-dependent phenotypic response, but a nonlinear, threshold-like, epistatic response as well. We also found that enhanced FGF8 signaling induced, and abrogated FGF8 signaling decreased, SMG branching morphogenesis in vitro. Furthermore, since FGF10 and Shh expression is modulated by Fgf8 levels, we postulated that exogenous FGF10, Shh, or FGF10 + Shh peptide supplementation in vitro would largely "rescue" the abnormal SMG phenotype associated with decreased FGF8 signaling. This is as expected, though there is no synergistic effect with FGF10 + Shh peptide supplementation. These in vitro experiments model the principle that mutations have different effects in the context of different epigenotypes.