Hyperhomocysteinemia increases permeability of the blood-brain barrier by NMDA receptor-dependent regulation of adherens and tight junctions

Abstract

IntroductionMicrovessels are responsible for facilitating the exchange of nutrients between blood and tissue while maintaining a barrier against the undesired movement of other molecules. This barrier is most finely tuned and structurally competent at the interface between the blood and the brain, the blood-brain barrier (BBB). The BBB is formed by intercellular junctions that limit paracellular diffusion combined with a low-flux transcellular system that selectively transports molecules either into or out of the brain parenchyma. Increasing evidence implicates chronic BBB permeability in the onset of neurologic and neurovascular diseases (eg, Alzheimer disease, dementia, small-vessel disease, stroke). 1 Although the BBB prevents the flux of harmful substances into the brain, it can also exclude the delivery of therapeutic drugs. Understanding the molecular regulation of the BBB is therefore important for the understanding of disease etiology and treatment.Paracellular diffusion of molecules at the BBB is minimized by endothelial cell-cell (EC-EC) junctions. Chief among these are the adherens junctions and tight junctions, of which vascularendothelial cadherin (VE-cadherin; VEC) and claudin-5 are wellknown members, respectively. Although VEC is endothelial specific, the claudin family of proteins is found in many epithelial tissues. However, claudin-5 is specific to tight junctions of ECs and is required to block paracellular diffusion of small molecules (Ͻ ϳ 800 Da) in the microvasculature of the brain. 2 In particular, claudin-5 is now recognized as the predominant claudin isoform in tight junctions of the BBB. Although claudins 1, 10, 11, and 12 (at least) are also expressed in brain endothelium, claudin-5 mRNA is nearly 600-fold higher than the other claudins expressed in microvascular ECs freshly isolated from the brain. 3 On electron microscopy, plasma membranes of adjacent cells associate close enough to merge without apparent intercellular space at tight junctions. 4 Homocysteine (Hcy) is an intermediate aminothiol derived from methionine catabolism, and its elevation in plasma, known as hyperhomocysteinemia (HHcy), is a chronic condition associated with cerebral small-vessel disease, stroke, and dementia (Alzheimer and vascular). 5,6 The list of Hcy-mediated vasculopathies is still growing but includes endothelial dysfunction, vessel wall malformations, loss of extracellular matrix collagen, and loosening of the BBB in rodents and humans. 5,6 Mounting evidence suggests that an increase in BBB permeability is an important factor in initiating cerebral small-vessel disease and lacunar stroke. 7,8 Hence, identifying the mechanisms by which HHcy opens the BBB is of great clinical interest. We reported that treating human umbilical vein ECs with Hcy increases EC monolayer permeability and decreases expression of claudin-5. 9 However, the mechanism by which Hcy triggers these events has not been elucidated, and the intracellular mechanisms for decreased expression of claudin-5 in HHcy are unknown.Recently, a ...

Expression of the ALS-Causing Variant hSOD1G93A Leads to an Impaired Integrity and Altered Regulation of Claudin-5 Expression in an in Vitro Blood–Spinal Cord Barrier Model

“…In the nucleus, pβ-cat forms a complex with FoxO1 and T-cell factor and the subsequent interaction of this complex with the claudin-5 promoter was shown to result in reduced levels of claudin-5. 20,29,30 In our study, we observed increased levels of p[S675]β-cat in the hSOD1 G93A cells compared with the wild-type cells indicating a mechanistic explanation for the reduced TJ protein levels. Furthermore, it was recently shown that the nuclear levels of β-cat are increased in whole lysates of ALS spinal cords of hSOD1 G93A mice, 31 pointing again to a direct involvement of mutant SOD expression in impaired barrier integrity due to an altered pathway responsible for TJ protein expression.…”

“…Thus, unphosphorylated FoxO1 is translocated to the nucleus, forming a complex with pβ-cat and T-cell factor and thereby repressing the claudin-5 expression. [18][19][20] To assess whether the expression of mutant SOD1 affects this pathway, we analyzed the phosphorylation states of β-cat, AKT, and FoxO1 by western blot (Figure 4). We observed increased levels of pβ-cat and decreased levels of pAKT in mutant bEnd.3 cells compared with SOD1 WT overexpressing cells ( Figures 4A and 4D).…”

Addition of NMDA-receptor antagonist MK801 during oxygen/glucose deprivation moderately attenuates the upregulation of glucose uptake after subsequent reoxygenation in brain endothelial cells

“…Interestingly, in the present work we have observed effects by MK801 in our BBB in vitro model without using astrocytes indicating the presence of alternative pathways. Recently, deleterious effects of homocysteine onto the BBB were reported to be mediated by NMDAR [2]. Furthermore, we and others were able to show that MK801 alone can affect the proteome and the intracellular Ca-level of BBB models [16,17].…”

Role of MicroRNA29b in Blood–Brain Barrier Dysfunction during Hyperhomocysteinemia: An Epigenetic Mechanism

“…29,30 Previous studies from our and other labs have also shown that high Hcy levels impart significant influence in BBB opening and thus raise permeability. 3,[31][32][33] Therefore, we created BBB disruption model by using Hcy in bEnd.3 cells and to validate in vitro findings, we used genetically hyperhomocysteinemic CBS þ / À mice models for in vivo studies. Previous studies also described bEnd.3 cells as a convenient and useful model to study BBB functions.…”