Abstract
Blood-brain barrier (BBB) impairment clearly accelerates brain disease progression. As ways to prevent injury-induced barrier dysfunction remain elusive, better understanding of how BBB cells interact and modulate barrier integrity is needed. Our metabolomic profiling study showed that cell-specific adaptation to injury correlates well with metabolic reprogramming at the BBB. In particular we noted that primary astrocytes (AC) contain comparatively high levels of glutathione (GSH)-related metabolites compared to primary endothelial cells (EC). Injury significantly disturbed redox balance in EC but not AC motivating us to assess 1) whether an AC-EC GSH shuttle supports barrier stability and 2) the impact of GSH on EC function. Using an isotopic labeling/tracking approach combined with Time-of-Flight Mass Spectrometry (TOF-MS) we prove that AC constantly shuttle GSH to EC even under resting conditions - a flux accelerated by injury conditions in vitro. In correlation, co-culture studies revealed that blocking AC GSH generation and secretion via siRNA-mediated γ-glutamyl cysteine ligase (GCL) knockdown significantly compromises EC barrier integrity. Using different GSH donors, we further show that exogenous GSH supplementation improves barrier function by maintaining organization of tight junction proteins and preventing injury-induced tight junction phosphorylation. Thus the AC GSH shuttle is key for maintaining EC redox homeostasis and BBB stability suggesting GSH supplementation could improve recovery after brain injury.
Highlights
Multiple studies suggest that stabilizing brain vascular function in patients with neurological disease could arrest or even reverse the course of brain disorders [2], but ways to attain this goal remain elusive
Using an isotopic labeling/ tracking approach combined with Time-of-Flight Mass Spectrometry (TOF-MS) we prove that AC secreted GSH is constantly shuttled to endothelial cells (EC), and this flux is accelerated by injury conditions
We are convinced that better understanding of the mechanisms by which perivascular cells support barrier function could provide new insight for future strategies aimed at modulating barrier function
Summary
Blood-brain barrier (BBB) breakdown and/or dysfunction occurs in many neurological diseases, and significantly contributes to disease progression [1,2,3]. Multiple studies suggest that stabilizing brain vascular function in patients with neurological disease could arrest or even reverse the course of brain disorders [2], but ways to attain this goal remain elusive. Specialized endothelial cells (EC) that are in close contact with perivascular astrocytes and pericytes, form the inner wall of brain microvessels. We are convinced that better understanding of how perivascular cells regulate both physiological and pathological EC responses will provide valuable insight into how to modulate BBB functionality
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