FGF10/FGFR2b signaling plays essential roles during in vivo embryonic submandibular salivary gland morphogenesis

Abstract

BackgroundAnalyses of Fgf10 and Fgfr2b mutant mice, as well as human studies, suggest that FGF10/FGFR2b signaling may play an essential, nonredundant role during embryonic SMG development. To address this question, we have analyzed the SMG phenotype in Fgf10 and Fgfr2b heterozygous and null mutant mice. In addition, although previous studies suggest that the FGF10/FGFR2b and FGF8/FGFR2c signaling pathways are functionally interrelated, little is known about the functional relationship between these two pathways during SMG development. We have designed in vivo and in vitro experiments to address this question.ResultsWe analyzed Fgf10 and Fgfr2b heterozygous mutant and null mice and demonstrate dose-dependent SMG phenotypic differences. Hypoplastic SMGs are seen in Fgf10 and Fgfr2b heterozygotes whereas SMG aplasia is seen in Fgf10 and Fgfr2b null embryos. Complementary in vitro studies further indicate that FGF10/FGFR2b signaling regulates SMG epithelial branching and cell proliferation. To delineate the functional relationship between the FGF10/FGFR2b and FGF8/FGFR2c pathways, we compared the SMG phenotype in Fgfr2c+/Δ/Fgf10+/- double heterozygous mice to that seen in wildtype, Fgf10+/- (Fgfr2c+/+/Fgf10+/-) and Fgfr2c+/Δ (Fgfr2c+/Δ/Fgf10+/+) single heterozygous mutant littermates and demonstrate genotype-specific SMG phenotypes. In addition, exogenous FGF8 was able to rescue the abnormal SMG phenotype associated with abrogated FGFR2b signaling in vitro and restore branching to normal levels.ConclusionOur data indicates that FGF10/FGFR2b signaling is essential for the SMG epithelial branching and histodifferentiation, but not earliest initial bud formation. The functional presence of other endogenous signaling pathways could not prevent complete death of embryonic SMG cells in Fgf10 and Fgfr2b null mice. Though we were able to rescue the abnormal phenotype associated with reduced in vitro FGF10/FGFR2b signaling with exogenous FGF8 supplementation, our results indicate that the FGF10/FGFR2b and FGF8/FGFR2c are nonredundant signaling pathways essential for in vivo embryonic SMG development. What remains to be determined is the in vivo functional relationship between the FGF10/FGFR2b signal transduction pathway and other key signaling pathways, and how these pathways are integrated during embryonic SMG development to compose the functional epigenome.

Immunolocalization of FGF-2, -7, -8, -10 and FGFR-1–4 during regeneration of the rat submandibular gland

“…In the present study, moderate immunoreactions for FGFR-1 were observed at the ICD of normal and Day 14, and slight immunoreactions were found in NFAC and MAC during regeneration. Previous studies have shown that FGFR-2 as the receptor of FGF-10 is very important in submandibular gland development (Celli et al 1998;Ohuchi et al 2000;Jaskoll et al 2005). However, our results showed that FGFR-2 was not expressed in the duct ligation model of submandibular gland regeneration.…”

“…Out of these, fibroblast growth factors (FGFs) consisting 22 family members (Itoh and Ornitz 2004) play crucial roles. For example, FGF-8/FGF receptor (FGFR)-2(IIIc) (Jaskoll et al 2002) and FGF-10/FGFR-2(IIIb) (Ohuchi et al 2000;Jaskoll et al 2005) signals contribute to bud formation and branching of the embryonic submandibular gland. FGF-2 (Madan and Kramer 2005) and -7 (Guo et al 1993) play possibly roles in differentiation of the salivary gland.…”

Parasympathetic Innervation Regulates Tubulogenesis in the Developing Salivary Gland

“…However, expression of Fgf10 and FGFR2b targets, Etv4 and 5 , were unchanged (Figure 7C). In addition, reduction of FGF10/FGFR2b signaling ex vivo (Figure S6A (Steinberg et al, 2005)) or in vivo (Figure S6B; (Jaskoll et al, 2005)) had no effect on ductal development. Together, these data indicate that deficient VIP-dependent innervation results in impaired SMG tubulogenesis in vivo .…”

“…Nrg1-III deficient salivary glands were devoid of functional nerves and exhibited aberrant ductal morphogenesis: epithelial branching was reduced (Figure 7A and B) with disrupted lumen formation (Figure 7A, ZO1 immunostaining, lower panel) and expansion (Figure 7D and F, arrows). We next asked whether this phenotype was due to loss of FGF10/FGFR2b signaling, an essential regulator of epithelial branching in the SMG (Jaskoll et al, 2005). However, expression of Fgf10 and FGFR2b targets, Etv4 and 5 , were unchanged (Figure 7C).…”

Anatomy, Biogenesis and Regeneration of Salivary Glands

“…This syndrome presents as aplasia and/or hypoplasia of the salivary and lacrimal glands, and also abnormalities of the ears, eyes, face, mouth, teeth, digits and genitourinary system. Again, mice with similar genetic mutations have similar defects in salivary gland biogenesis and have been used to study how FGF signaling affects gland biogenesis [24,25]. …”