Mechanisms of vasculogenesis in 3D fibrin matrices mediated by the interaction of adipose-derived stem cells and endothelial cells
Mechanisms of vasculogenesis in 3D fibrin matrices mediated by the interaction of adipose-derived stem cells and endothelial cells
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DOI：
10.1007/s10456-014-9439-0
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发表时间：
2014-10-01
期刊：
影响因子：
9.8
通讯作者：
Holnthoner, Wolfgang
中科院分区：
文献类型：
--
作者：
Rohringer, Sabrina;Hofbauer, Pablo;Holnthoner, Wolfgang
组织工程构建物的血管化对于在植入靶部位后提供足够的营养供应和止血至关重要。脂肪源性干细胞（ASC）与生长的内皮细胞（OEC）在纤维蛋白凝胶中的共培养被证明提供了一种有效的体外诱导血管生成的可能性。然而，迄今为止，这两种细胞类型之间的相互作用机制仍不清楚。本研究的目的是评估直接和间接刺激ASC诱导的血管生成的差异，ASC对网络稳定性的影响以及参与血管结构形成的分子机制。内皮细胞（EC）包埋在纤维蛋白凝胶中，含有非包被或ASC包被的微载体珠以及单独的ASC。此外，除了在顶部接种ASC的构建体之外，还使用与ASC条件培养基孵育的EC接种的构建体。通过表达绿色荧光蛋白的细胞或CD 31的免疫染色观察血管网络形成并定量。对来源于纤维蛋白中的共培养物的细胞进行RT-qPCR，以评价培养第一周期间EC标志物基因表达的变化。此外，血管生成相关的蛋白质水平进行测量，通过执行血管生成蛋白质组分析仪阵列。结果表明，内皮细胞和ASC的接近是网络形成所需的，ASC通过发展周细胞特性来稳定EC网络。我们进一步表明ASC通过分泌促血管生成和调节蛋白诱导受控的血管生长。本研究揭示了纤维蛋白基质中EC/ASC相互作用所涉及的血管生成蛋白质谱，并证实了OEC/ASC共培养物用于自体血管组织工程的可用性。
Vascularization of tissue-engineered constructs is essential to provide sufficient nutrient supply and hemostasis after implantation into target sites. Co-cultures of adipose-derived stem cells (ASC) with outgrowth endothelial cells (OEC) in fibrin gels were shown to provide an effective possibility to induce vasculogenesis in vitro. However, the mechanisms of the interaction between these two cell types remain unclear so far. The aim of this study was to evaluate differences of direct and indirect stimulation of ASC-induced vasculogenesis, the influence of ASC on network stabilization and molecular mechanisms involved in vascular structure formation. Endothelial cells (EC) were embedded in fibrin gels either containing non-coated or ASC-coated microcarrier beads as well as ASC alone. Moreover, EC-seeded constructs incubated with ASC-conditioned medium were used in addition to constructs with ASC seeded on top. Vascular network formation was visualized by green fluorescent protein expressing cells or immunostaining for CD31 and quantified. RT-qPCR of cells derived from co-cultures in fibrin was performed to evaluate changes in the expression of EC marker genes during the first week of culture. Moreover, angiogenesis-related protein levels were measured by performing angiogenesis proteome profiler arrays. The results demonstrate that proximity of endothelial cells and ASC is required for network formation and ASC stabilize EC networks by developing pericyte characteristics. We further showed that ASC induce controlled vessel growth by secreting pro-angiogenic and regulatory proteins. This study reveals angiogenic protein profiles involved in EC/ASC interactions in fibrin matrices and confirms the usability of OEC/ASC co-cultures for autologous vascular tissue engineering.