Abstract
Purpose :
Pericytes, cells crucially important to maintain a healthy microvasculature, make direct cell connections with vascular endothelial cells yet the functional significance of these contacts remains largely unexplored. In mice lacking the cell signaling protein Notch 3 or in diabetic retinopathy it is well known that pericytes drop-out from the vasculature, thus, we wanted to test the hypothesis that pericytes lose direct connection to endothelial cells by analyzing the ultrastructure.
Methods :
Serial section transmission electron microscopy (TEM) was used to image mouse retinal ganglion cell layer capillaries in wild type, Notch 3 knock out (KO), and STZ induced diabetic mice (n=3 mice for each condition and mice were 6-8 months old). The dataset of TEM images that was segmented consisted of 19 WT, 21 diabetic, and 17 Notch 3 vessels. For each vessel a series of z-sections were acquired: ~14 images per vessel in WT (271 total images), ~16 images per vessel in Notch 3 KO (281 total images), and ~13 images per vessel in diabetic (283 total images). Images were segmented manually using Image J v4 to trace the boundaries of the basement membrane, endothelial cells, mural cells, and peg-and-socket connections (Fig 1A). The segmented images were analyzed using Matlab 2021a to measure contact lengths between mural cells and endothelial cells and peg-and-socket properties.
Results :
This study revealed significant differences in the structure of endothelial cell-pericyte connections in diabetic and Notch 3 KO animals when compared to WT. While the vessels analyzed in each group were of similar diameter and amount of pericyte, there was an increase in the depth of endothelial cell-pericyte contacts in diabetic and Notch 3 KO (Fig 1B)
Conclusions :
As the Notch 3 receptor plays an important role in cell signaling between pericytes and endothelial cells and diabetes is also known to disrupt Notch 3 signaling, our hypothesis for the enlarged peg phenotype is that the pericytes and endothelial cells actively increase their contact surface to compensate for loss of Notch 3 signaling.
This abstract was presented at the 2024 ARVO Imaging in the Eye Conference, held in Seattle, WA, May 4, 2024.