Kwon Hae Ryong, Nelson Deirdre A, DeSantis Kara A, Morrissey Jennifer M, Larsen Melinda
Department of Biological Sciences, University at Albany, State University of New York, 1400 Washington Avenue, Albany, NY 12222, USA.
Graduate Program in Molecular, Cellular, Developmental, and Neural Biology, University at Albany, State University of New York, 1400 Washington Avenue, Albany, NY 12222, USA.
Development. 2017 Jan 15;144(2):211-220. doi: 10.1242/dev.142497.
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. Immuno-depletion 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.
在器官发育和再生过程中，脉管系统对上皮细胞模式形成的灌注非依赖性调节对于恢复器官功能的应用具有重要意义。在小鼠下颌下唾液腺发育过程中，脉管系统在分支形态发生过程中与上皮细胞共同发育；然而，尚不清楚脉管系统是否对上皮细胞具有指导作用。我们使用胚胎器官外植体中的药理学抑制剂和小干扰RNA敲低技术，确定唾液腺上皮细胞模式形成需要VEGFR2依赖性信号传导。为了直接测试内皮细胞在指导性上皮细胞模式形成中的必要性，我们开发了一种新型的体外细胞分级分离/重组试验。在该试验中对CD31内皮细胞进行免疫耗竭，证实了内皮细胞在腺体上皮细胞模式形成中的必要性。器官外植体中内皮细胞的耗竭或VEGFR2信号传导的抑制导致表达导管蛋白K19和K7的细胞异常增加，而重组腺体终末芽中的Kit祖细胞减少。向重组腺体中添加外源性内皮细胞可恢复上皮细胞模式形成，补充内皮细胞调节的间充质因子IGFBP2和IGFBP3也可恢复上皮细胞模式形成。我们的结果表明，内皮细胞促进早期发育的胚胎下颌下唾液腺芽中Kit祖细胞的扩增，并抑制导管过早分化。
