title: 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.
authors.affiliation: Department of Occupational and Environmental Health and the Ministry-of-Education's Key Laboratory of Hazard Assessment and Control in Special Operational Environment, School of Public Health, Fourth Military Medical University, Xi'an 710032, China.
authors.forename: Han
authors.initials: H
authors.lastname: Song
authors.forename: Gang
authors.initials: G
authors.lastname: Zheng
authors.forename: Xue-Feng
authors.initials: XF
authors.lastname: Shen
authors.forename: Xin-Qin
authors.initials: XQ
authors.lastname: Liu
authors.forename: Wen-Jing
authors.initials: WJ
authors.lastname: Luo
authors.forename: Jing-Yuan
authors.initials: JY
authors.lastname: Chen
mesh_terms: D000818:Animals; D001812:Blood-Brain Barrier / Q000187:drug effects* / Q000378:metabolism; D002199:Capillary Permeability / Q000187:drug effects; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004785:Environmental Pollutants / Q000097:blood / Q000633:toxicity*; D006360:Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D007854:Lead / Q000097:blood / Q000633:toxicity*; D008297:Male; D046529:Microscopy, Electron, Transmission; D055806:Microvessels / Q000648:ultrastructure; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D060888:Real-Time Polymerase Chain Reaction; D062725:Tight Junction Proteins / Q000378:metabolism*; D019061:src-Family Kinases / Q000235:genetics / Q000378:metabolism*; D000091342:Endoplasmic Reticulum Chaperone BiP
chemical_list: D004785:Environmental Pollutants; D006360:Heat-Shock Proteins; D007854:Lead; D034741:RNA, Small Interfering; D062725:Tight Junction Proteins; D019061:src-Family Kinases; C051684:GRP78 protein, rat; D000091342:Endoplasmic Reticulum Chaperone BiP
keywords: GRP78; Lead; Src; occludin; tight junctions