Reduction of Brain Barrier Tight Junctional Proteins by Lead Exposure: Role of Activation of Nonreceptor Tyrosine Kinase Src via Chaperon GRP78

Abstract

Lead (Pb) has long been recognized as a neurodevelopmental toxin. Developing blood-brain barrier (BBB) is known to be a target of Pb neurotoxicity; however, the underlying mechanisms are still unclear. Recent evidence suggests that intracellular nonreceptor protein tyrosine kinase Src regulates tight junctional proteins (TJPs). This study was designed to investigate whether Pb acted on the Src-mediated cascade event leading to an altered TJP expression at BBB. Rats aged 20-22 days were exposed to Pb in drinking water (0, 100, 200, and 300 ppm Pb) for eight weeks. Electron microscopic and Western blot analyses revealed a severe leakage of BBB and significantly decreased expressions of TJP occludin and ZO-1. When cultured brain endothelial RBE4 cells were exposed to 10μM Pb for 24 h, expressions of phosphor-Src and an upstream regulator GRP78 were significantly increased by 6.42-fold and 8.29-fold (p < 0.01), respectively. Inactivation of Src pathway by a Src-specific inhibitor reversed Pb-induced downregulation of occludin, but not ZO-1; small interfering RNA knockdown of GRP78 attenuated Pb-induced Src phosphorylation and occludin reduction. Furthermore, Pb exposure caused redistribution of GRP78 from endoplasmic reticulum to cytosol and toward cell member. However, the data from immunoneutralization studies did not show the involvement of cell-surface GRP78 in regulating Src phosphorylation upon Pb exposure, suggesting that the cytosolic GRP78, rather than cell-surface GRP78, was responsible to Pb-induced Src activation and ensuing occludin reduction. Taken together, this study provides the evidence of a novel linkage of GRP78, Src activation to downregulation of occludin, and BBB disruption during Pb exposure.

Spatiotemporal Expression of GRP78 in the Blood Vessels of Rats Treated With 3-Nitropropionic Acid Correlates With Blood–Brain Barrier Disruption

“…Despite the potential relationship between GRP78 expression and BBB integrity, little is known about vessel-associated cells expressing GRP78 in the lesion core. Although a limited number of studies have reported GRP78 expression in vascular endothelial cells ( Yan et al, 2011 ; Birukova et al, 2014 ; Song et al, 2014 ), to our knowledge, this is the first study to show that, in addition to endothelial cells, smooth muscle cells and adventitial cells also express GRP78. In particular, adventitial cells had the following features that were similar to those of perivascular fibroblast-like cells, reported in our previous study, although their exact cellular identity had not been elucidated: they were localized along the outer part of smooth muscle cells, had euchromatic nuclei with prominent nucleoli, and coexpressed GRP78 with nestin ( Shin et al, 2013 ).…”

“…A correlative approach using light and electron microscopy clearly demonstrated that, in these vasculature-associated cells, conspicuous silver-enhanced grains indicative of GRP78 were specifically localized in the rough ER/perinuclear cisternae, indicating that despite the phenotypic heterogeneity of vascular GRP78-positive cells, GRP78 was present as an ER protein in these cells. This ER localization of GRP78 in endothelial cells is unexpected considering that GRP78 is redistributed from the ER to the cytosol or plasma membrane of endothelial cells in vitro , where it acts as a receptor mediating for controlling cell viability and signaling ( Birukova et al, 2014 ; Song et al, 2014 ). In our previous studies, however, GRP78 protein was almost exclusively localized to the rough ER in both constitutive cells (neurons) and cells induced by ER stress (reactive astrocytes and brain macrophages) after focal cerebral ischemia ( Jin et al, 2018a ).…”

“…GRP78, associated with its cofactor HTJ-1, can be translocated and anchored to the cell surface by oxidized phospholipids, suggesting that GRP78 is a novel receptor initiating a protective vascular barrier response to oxidized phospholipids ( Birukova et al, 2014 ). By contrast, lead exposure induces GRP78 expression in brain endothelial cells, which leads to Src activation and subsequent reduction of tight junctional proteins, with ensuing BBB disruption ( Song et al, 2014 ). In addition, a recent study demonstrated that exposure to GRP78-specific recombinant antibodies results in increased BBB leakage, suggesting that GRP78 autoantibodies are associated with BBB disruption in neuromyelitis optica ( Shimizu et al, 2017 ).…”

Post-treatment with Ma-Huang-Tang ameliorates cold-warm-cycles induced rat lung injury

“…These results highly suggest activation of a protein that mediates both para- and trans-cellular pathways. We speculated that this protein is Src, because phosphorylation of Src has been reported not only to increase the expression and phosphorylation of Caveolin-1 4 but also reduce Occluding and ZO-1 from tight junctions 29–31 . This speculation was validated by the finding that Src in rat pulmonary tissue was activated in response to cold-warm-cycles challenge.…”

Blood–Brain Barrier Damage and Dysfunction by Chemical Toxicity

“…Metals, such as manganese (Mn), lead (Pb), and mercury (Hg), are transported on endogenous carriers, which otherwise function in the transport of essential macromolecules (Aschner, 1998;Song et al, 2014). Mn binds readily to transferrin without displacing iron (Fe) in plasma.…”