Virginia Research Day 2022

Medical Student Research Biomedical

15 ARCHITECTURE OF THE JEJUNAL AND COLONIC MICROVASCULATURE

Glynn B. Reno; Natalie Jeong Min Kim; Tatiana M. Midkiff; Christina B. Bagnati; Swapnil K. Sonkusare; Anthony J.M. Bauer Corresponding author: gbhorkott@liberty.edu

Liberty University College of Osteopathic Medicine University of Virginia

Background: The intestinal microvasculature is essential in maintaining intestinal oxygenation, digestion, absorption, and mucosal barrier function. Many diseases result in gut ischemia, inflammation, and abnormal vascular plasticity. Surprisingly, the architecture of the intestinal microvasculature has not been clearly delineated and its relationship to other cell types within the gut wall has not been elucidated. Our objective was to construct the 3D architecture of the murine intestinal microvasculature and show its relationship to the enteric nervous system and constitutive innate immune cells. Methods: A laser-scanning confocal system was used to image live intact segments or fixed whole-mounts of the C57Bl/6 intestine. The microvasculature was labeled with isolectin-B4 (IB4) and dextran conjugated to fluorophores. Neutrophils were labeled with Hanker-Yates reagent, macrophages with a F4/80 + antibody, mast cells with avidin and enteric nervous system with IB4. An Andor high speed confocal system was used to visualize the endothelial

present. Neutrophils were absent within the muscularis but arranged in a perivascular manner within the submucosa and distributed within the mucosa. Dragonfly confocal imaging of the endothelial specific Cdh5-GCaMP8 expressing microvasculature elucidated the 3D architecture of the jejunal vascular endothelial cell network of arterioles, venules, and lymph vessels. Conclusions: The results have elucidated the detailed 3D architecture of the intestinal microvasculature and has demonstrated its relationship to the ENS, macrophages, mast cells and neutrophils. Of particular interest is the tight intertwining of the muscularis capillary network with the myenteric plexus. And, also, the potential role of the perivascular macrophages in regulating intestinal blood flow.

architecture of the arterioles, venules and lymphatics of Cdh5-GCaMP8 expressing mice. Results: 3D imaging demonstrated that the mesenteric arteries entered the bowel and coursed through the muscularis externa, arborized within the submucosal layer sending arteriole projections outwards into the muscularis and inwards to the mucosa. The jejunal muscularis capillaries formed a singular planar myenteric network between the circular and longitudinal muscle layers entwining the myenteric plexus. Whereas, in the thicker colonic muscularis capillaries were observed running parallel deep along the circular muscle bundles. Venous drainage proceeded in reverse order. Interestingly, the jejunal muscularis capillary network was closely associated with the myenteric plexus, but not the submucosal plexus. Muscularis macrophages formed a dense network within the myenteric capillary system and a sparse network between the circular muscle bundles. Submucosal macrophages encased arterioles and venules. Few submucosal perivascular mast cells were

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