FGF8 dose-dependent regulation of embryonic submandibular salivary gland morphogenesis

Abstract

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.

Ectodysplasin and Wnt pathways are required for salivary gland branching morphogenesis

“…Although the function of Shh in SMG morphogenesis is not entirely clear, Fgf8, a potent mitogen that is essential for the growth of SMG (Jaskoll et al, 2004b), has been proposed to act downstream of Shh, as exogenous Fgf8 was shown to rescue the growth defect of cyclopamine-treated SMGs (Jaskoll et al, 2004a). In line with this, qRT-PCR analysis of Eda-null SMGs at later stages (E16 to newborn) indicated that both Shh and Fgf8 were downregulated (Melnick et al, 2009).…”

FGFR2b signaling regulates ex vivo submandibular gland epithelial cell proliferation and branching morphogenesis

“…Targeted overexpression of FGF7 with the K14 promoter caused smaller salivary glands, excessive salivation, and a delay in gland differentiation (Guo et al, 1993). FGF8-conditional mutant and FGF8-hypomorphic mice also have defects in SMG development and differentiation (Jaskoll et al, 2004). Taken together, these studies suggest a primary role for FGF10/FGFR2b signaling in the initiation of the gland, an essential role for FGF8 at later stages, and possibly a role for FGF7 during differentiation.…”

Involvement of Wnt, Eda and Shh at defined stages of sweat gland development

“…The specification of appendage type may then be effected at least in part by variations in the Shh pathway (see below). In contrast to hair follicles and sweat glands, salivary gland early stage development depends instead on Fgf and Pitx2 pathways (Jaskoll et al, 2004b;Tucker, 2007). Eda and Shh pathways are then required for latestage branching morphogenesis (Häärä et al, 2011;Patel et al, 2011).…”