Development of Quantitative Tools for the Characterization and Analysis of the Blood Brain Barrier in Normal and MPS IIIB Mice

Abstract

The Blood Brain Barrier (BBB) functions to control homeostasis within the central nervous system (CNS) via strict control over the passage of molecules into and out of the brain. The goal of this barrier is to protect the CNS from harmful external factors while allowing entry of essential nutrients and removal of metabolic byproducts. This restrictive nature of the BBB is due to the protein network at the borders of adjoining cerebral endothelial cells known as the tight junctions (TJs) and the relationship of other supporting cells and proteins such as astrocytes, pericytes, microglia, and the basement membrane and extracellular matrix proteins associated with the endothelial cells. While beneficial in health, the restrictive nature of the BBB has proved to be a hindrance towards drug administration in many brain diseases, blocking pharmaceutical compounds from diffusing out of the blood vessels and into the brain parenchyma. However, certain diseases such as the metabolic disease Mucopolysaccharidosis IIIB, have been shown to cause breaches in the BBB's integrity, thus suggesting a possible mechanism to administer treatment around this restriction could be to utilize a specific disease's own pathology. In order to understand the extent of BBB dysfunction a systematic and quantitative analysis of the BBB's cellular and protein components and their relationships in each disease of interest during the disease's progression compared to normal conditions is required. This project used advancing technologies in confocal microscopy and 3D image analysis to develop and utilize methodologies to analyze each of the components of the BBB in wild type mice with the goal to quantify their relationship with the cerebral microvascular endothelial cells. In doing so valuable image processing protocols were developed revealing the inadequacies of traditional 2D methods and has helped begin to shed light on the effects of MPS IIIB on the BBB including disrupted endothelial cells along the tight junctions, increased astrocyte contact area and points of contact with endothelial cells, and no changes in the contact area of desmin positive pericytes with endothelial cells.