Azimi Mohammad S, Motherwell Jessica M, Murfee Walter L
Biomedical Engineering, Tulane University.
Biomedical Engineering, Tulane University;
J Vis Exp. 2017 Feb 9(120):55183. doi: 10.3791/55183.
Angiogenesis, defined as the growth of new blood vessels from pre-existing vessels, involves endothelial cells, pericytes, smooth muscle cells, immune cells, and the coordination with lymphatic vessels and nerves. The multi-cell, multi-system interactions necessitate the investigation of angiogenesis in a physiologically relevant environment. Thus, while the use of in vitro cell-culture models have provided mechanistic insights, a common critique is that they do not recapitulate the complexity associated with a microvascular network. The objective of this protocol is to demonstrate the ability to make time-lapse comparisons of intact microvascular networks before and after angiogenesis stimulation in cultured rat mesentery tissues. Cultured tissues contain microvascular networks that maintain their hierarchy. Immunohistochemical labeling confirms the presence of endothelial cells, smooth muscle cells, pericytes, blood vessels and lymphatic vessels. In addition, labeling tissues with BSI-lectin enables time-lapse comparison of local network regions before and after serum or growth factor stimulation characterized by increased capillary sprouting and vessel density. In comparison to common cell culture models, this method provides a tool for endothelial cell lineage studies and tissue specific angiogenic drug evaluation in physiologically relevant microvascular networks.
血管生成被定义为从已有的血管中生长出新的血管，涉及内皮细胞、周细胞、平滑肌细胞、免疫细胞，以及与淋巴管和神经的协调。多细胞、多系统的相互作用使得在生理相关环境中研究血管生成成为必要。因此，虽然体外细胞培养模型的使用提供了机制上的见解，但一个常见的批评是它们无法重现与微血管网络相关的复杂性。本方案的目的是展示在培养的大鼠肠系膜组织中，对血管生成刺激前后完整微血管网络进行延时比较的能力。培养的组织包含维持其层级结构的微血管网络。免疫组织化学标记证实了内皮细胞、平滑肌细胞、周细胞、血管和淋巴管的存在。此外，用BSI-凝集素标记组织能够对血清或生长因子刺激前后的局部网络区域进行延时比较，其特征是毛细血管芽生和血管密度增加。与常见的细胞培养模型相比，该方法为内皮细胞谱系研究和在生理相关微血管网络中进行组织特异性血管生成药物评估提供了一种工具。
