Endothelial cell regulation of salivary gland epithelial patterning

Abstract

Perfusion-independent regulation of epithelial pattern formation by the vasculature during organ development and regeneration is of considerable interest for application in restoring organ function. During murine submandibular salivary gland development, the vasculature co-develops with the epithelium during branching morphogenesis; however, it is not known whether the vasculature has instructive effects on the epithelium. Using pharmacological inhibitors and siRNA knockdown in embryonic organ explants, we determined that VEGFR2-dependent signaling is required for salivary gland epithelial patterning. To test directly for a requirement for endothelial cells in instructive epithelial patterning, we developed a novel ex vivo cell fractionation/reconstitution assay. Immunodepletion of CD31 + endothelial cells in this assay confirmed a requirement for endothelial cells in epithelial patterning of the gland. Depletion of endothelial cells or inhibition of VEGFR2 signaling in organ explants caused an aberrant increase in cells expressing the ductal proteins K19 and K7, with a reduction in Kit + progenitor cells in the endbuds of reconstituted glands. Addition of exogenous endothelial cells to reconstituted glands restored epithelial patterning, as did supplementation with the endothelial cell-regulated mesenchymal factors IGFBP2 and IGFBP3. Our results demonstrate that endothelial cells promote expansion of Kit + progenitor cells and suppress premature ductal differentiation in early developing embryonic submandibular salivary gland buds.

Initial stages of development of the submandibular gland (human embryos at 5.5–8 weeks of development)

“…A recent report indicated the importance of endothelial factors to glandular development in mice (Kwon et al. ).…”

The expression of water channel proteins during human salivary gland development: a topographic study of aquaporins 1, 3 and 5

“…The expression pattern of AQP1 seem to be important for water transport from blood vessels to SG, aiding the formation of the future salivary fluid, corroborating our results (Akamatsu et al 2003 ). In addition, recent studies on the role of vascularisation in mouse salivary glands have reported the requirement of endothelial cells for gland development by promoting expansion of specific progenitor cells during early stages (Kwon et al 2017 ).…”

Salivary gland stem cells: A review of development, regeneration and cancer

“…These interactions are formed at the beginnings of organogenesis: the E11.5 epithelium invaginates into a condensing neural crest-derived mesenchyme containing a discontinuous CD31+ endothelial plexus and recruits SOX10+ neural precursors to form a post-ganglionic parasympathetic ganglion (Figure 2). During the next 36 h the endothelium becomes continuous and axons extend from the ganglion and travel along the developing ductal system to envelope newly forming end buds, thus forming an integrated organ system (Coughlin, 1975; Knosp et al , 2015; Knox et al , 2010; Kwon et al , 2017). As would be expected from such a heterogeneous structure, many pathways are involved in regulating epithelial branching, including those mediated by growth factors such as FGF, EGF, WNT, Hedgehog, and EDA (reviewed in Mattingly et al , 2015) and neurotransmitters acetylcholine (ACh) (Knox et al , 2010) and vasoactive intestinal protein (VIP) (Nedvetsky et al , 2014).…”