Hodges Nicholas A, Barr Ryan W, Murfee Walter L
Tulane University, Department of Biomedical Engineering, New Orleans, LA, 70118, United States; University of Florida, Department of Biomedical Engineering, Gainesville, FL, 32611, United States.
Tulane University, Department of Biomedical Engineering, New Orleans, LA, 70118, United States.
J Neurosci Methods. 2020 Dec 1;346:108923. doi: 10.1016/j.jneumeth.2020.108923. Epub 2020 Sep 1.
Neurovascular patterning is an emerging area of microvascular research. While overlapping molecular signals highlight links between angiogenesis and neurogenesis, advancing our understanding is limited by a lack of in vitro models containing both systems. One potential model is the rat mesentery culture model, which our laboratory has recently introduced as an ex vivo tool to investigate cellular dynamics during angiogenesis in a microvascular network scenario. The objective of this study was to demonstrate the use of the rat mesentery culture model as an ex vivo platform for maintaining the spatiotemporal relationship between blood vessels and peripheral nerves during angiogenesis in adult microvascular networks.
Adult male Wistar rat mesenteric tissue windows were harvested, rinsed in sterile DPBS and medium and then cultured per group: 1) MEM alone and 2) NBM with NGF and 20 % FBS (nerve culture medium). On day 3 post culture tissues were labeled for endothelial (PECAM) and neural (class III β-tubulin, NG2, tyrosine hydroxylase, CGRP) markers.
In MEM alone tissues nerve segment degeneration was supported by discontinuous nerve or absence of nerve marker labeling. Nerve presence at the arteriole level and capillary level was maintained for the nerve culture medium group compared to day 0, non-cultured control group (unstimulated).
The results support the use of specific medium types to maintain nerve presence across cultured microvascular networks and implicates the rat mesentery culture model as a novel ex vivo tool for investigating neurovascular patterning in adult tissues.
神经血管模式形成是微血管研究中一个新兴领域。虽然重叠的分子信号突出了血管生成与神经发生之间的联系，但由于缺乏包含这两个系统的体外模型，我们对其的深入理解受到限制。一种潜在的模型是大鼠肠系膜培养模型，我们实验室最近将其作为一种体外工具引入，用于在微血管网络场景中研究血管生成过程中的细胞动态。本研究的目的是证明大鼠肠系膜培养模型可作为一种体外平台，用于在成年微血管网络血管生成过程中维持血管与周围神经之间的时空关系。
采集成年雄性Wistar大鼠的肠系膜组织窗，用无菌DPBS和培养基冲洗，然后分组培养：1）单独使用MEM和2）添加NGF和20%胎牛血清的NBM（神经培养基）。培养后第3天，对组织进行内皮（PECAM）和神经（III类β-微管蛋白、NG2、酪氨酸羟化酶、降钙素基因相关肽）标记物的标记。
在单独使用MEM的组织中，神经节段退化表现为神经不连续或缺乏神经标记物标记。与第0天未培养的对照组（未刺激）相比，神经培养基组在小动脉水平和毛细血管水平的神经得以保留。
结果支持使用特定类型的培养基来维持培养的微血管网络中的神经存在，并表明大鼠肠系膜培养模型是一种用于研究成年组织中神经血管模式形成的新型体外工具。
