Department of Medical Pharmacology and Physiology, Dalton Cardiovascular Sciences Center and University of Missouri School of Medicine, Columbia, Missouri, United States of America.
PLoS One. 2013 Dec 31;8(12):e85147. doi: 10.1371/journal.pone.0085147. eCollection 2013.
We describe a novel 3D fibrin matrix model using recombinant hematopoietic stem cell cytokines under serum-free defined conditions which promotes the assembly of human endothelial cell (EC) tubes with co-associated pericytes. Individual ECs and pericytes are randomly mixed together and EC tubes form that is accompanied by pericyte recruitment to the EC tube abluminal surface over a 3-5 day period. These morphogenic processes are stimulated by a combination of the hematopoietic stem cell cytokines, stem cell factor, interleukin-3, stromal derived factor-1α, and Flt-3 ligand which are added in conjunction with fibroblast growth factor (FGF)-2 into the fibrin matrix. In contrast, this tube morphogenic response does not occur under serum-free defined conditions when VEGF and FGF-2 are added together in the fibrin matrices. We recently demonstrated that VEGF and FGF-2 are able to prime EC tube morphogenic responses (i.e. added overnight prior to the morphogenic assay) to hematopoietic stem cell cytokines in collagen matrices and, interestingly, they also prime EC tube morphogenesis in 3D fibrin matrices. EC-pericyte interactions in 3D fibrin matrices leads to marked vascular basement membrane assembly as demonstrated using immunofluorescence and transmission electron microscopy. Furthermore, we show that hematopoietic stem cell cytokines and pericytes stimulate EC sprouting in fibrin matrices in a manner dependent on the α5β1 integrin. This novel co-culture system, under serum-free defined conditions, allows for a molecular analysis of EC tube assembly, pericyte recruitment and maturation events in a critical ECM environment (i.e. fibrin matrices) that regulates angiogenic events in postnatal life.
我们描述了一种新颖的 3D 纤维蛋白基质模型，该模型使用无血清定义条件下的重组造血干细胞细胞因子，促进人内皮细胞（EC）管与共相关周细胞的组装。单个 EC 和周细胞随机混合在一起，EC 管形成，同时在 3-5 天内募集周细胞到 EC 管腔表面。这些形态发生过程受到造血干细胞细胞因子（干细胞因子、白细胞介素 3、基质衍生因子-1α 和 Flt-3 配体）的组合刺激，这些细胞因子与纤维蛋白基质中的成纤维细胞生长因子（FGF）-2 一起添加。相比之下，当无血清定义条件下将 VEGF 和 FGF-2 添加到纤维蛋白基质中时，不会发生这种管形态发生反应。我们最近证明，VEGF 和 FGF-2 能够使内皮细胞管形态发生反应（即在形态发生测定前过夜添加）对纤维蛋白基质中的造血干细胞细胞因子敏感，有趣的是，它们也能使内皮细胞管在 3D 纤维蛋白基质中形态发生。内皮细胞-周细胞相互作用在 3D 纤维蛋白基质中导致明显的血管基底膜组装，如免疫荧光和透射电子显微镜所示。此外，我们表明，造血干细胞细胞因子和周细胞以依赖于α5β1 整合素的方式刺激纤维蛋白基质中的 EC 出芽。在无血清定义条件下，这种新型共培养系统允许在关键细胞外基质环境（即纤维蛋白基质）中对内皮细胞管组装、周细胞募集和成熟事件进行分子分析，该环境调节出生后生命中的血管生成事件。
