title: Disruption of blood-brain barrier integrity in postmortem alcoholic brain: preclinical evidence of TLR4 involvement from a binge-like drinking model.
abstract: Inflammatory cytokines and reactive oxygen species are reported to be involved in blood-brain barrier (BBB) disruption. Because there is evidence that ethanol (EtOH) induces release of free radicals, cytokines and inflammatory mediators we examined BBB integrity and matrix metalloproteinase (MMP) activity in postmortem human alcoholic brain and investigated the role of TLR4 signaling in BBB permeability in TLR4-knockout mice under a binge-like EtOH drinking protocol. Immunohistochemical studies showed reduced immunoreactivity of the basal lamina protein, collagen-IV and of the tight junction protein, claudin-5 in dorsolateral prefrontal cortex of alcoholics. There was also increased MMP-9 activity and expression of phosphorylated ERK1/2 and p-38. Greater number of CD45+ IR cells were observed associated with an enhanced neuroinflammatory response reflected by increased GFAP and Iba-1 immunostaining. To further explore effects of high EtOH consumption on BBB integrity we studied TLR4-knockout mice exposed to the drinking in the dark paradigm. Repetitive EtOH exposure in wild-type mice decreased hippocampal expression of laminin and collagen-IV and increased IgG immunoreactivity, indicating IgG extravasation. Western blot analysis also revealed increased MyD88 and p-ERK1/2 levels. None of these changes was observed in TLR4-knockout mice. Collectively, these findings indicate that chronic EtOH increases degradation of tight junctions and extracellular matrix in postmortem human brain and induces a neuroinflammatory response associated with activation of ERK1/2 and p-38 and greater MMP-9 activity. The EtOH-induced effects on BBB impairment are not evident in the hippocampus of TLR4-knockout mice, suggesting the involvement of TLR4 signaling in the underlying mechanism leading to BBB disruption in mice.
authors.affiliation: Departamento de Farmacología, Facultad de Medicina, Universidad Complutense, Instituto de Investigación Sanitaria Hospital 12 de Octubre, Spain.|Red de Trastornos Adictivos del Instituto de Salud Carlos III, Spain.
authors.forename: Ana
authors.initials: A
authors.lastname: Rubio-Araiz
authors.affiliation: Departamento de Farmacología, Facultad de Medicina, Universidad Complutense, Instituto de Investigación Sanitaria Hospital 12 de Octubre, Spain.|Red de Trastornos Adictivos del Instituto de Salud Carlos III, Spain.
authors.forename: Francesca
authors.initials: F
authors.lastname: Porcu
authors.affiliation: Departamento de Farmacología, Facultad de Medicina, Universidad Complutense, Instituto de Investigación Sanitaria Hospital 12 de Octubre, Spain.|Red de Trastornos Adictivos del Instituto de Salud Carlos III, Spain.
authors.forename: Mercedes
authors.initials: M
authors.lastname: Pérez-Hernández
authors.affiliation: Instituto de Neurociencias, Campus de San Juan, Universidad Miguel Hernández-CSIC, Spain.|Red de Trastornos Adictivos del Instituto de Salud Carlos III, Spain.
authors.forename: Mª Salud
authors.initials: MS
authors.lastname: García-Gutiérrez
authors.affiliation: Instituto de Neurociencias, Campus de San Juan, Universidad Miguel Hernández-CSIC, Spain.|Red de Trastornos Adictivos del Instituto de Salud Carlos III, Spain.
authors.forename: María Auxiliadora
authors.initials: MA
authors.lastname: Aracil-Fernández
authors.affiliation: Departamento de Farmacología, Facultad de Medicina, Universidad Complutense, Instituto de Investigación Sanitaria Hospital 12 de Octubre, Spain.|Red de Trastornos Adictivos del Instituto de Salud Carlos III, Spain.
authors.forename: María Dolores
authors.initials: MD
authors.lastname: Gutierrez-López
authors.affiliation: Departamento de Patología Celular, Centro de Investigación Príncipe Felipe, Spain.|Red de Trastornos Adictivos del Instituto de Salud Carlos III, Spain.
authors.forename: Consuelo
authors.initials: C
authors.lastname: Guerri
authors.affiliation: Instituto de Neurociencias, Campus de San Juan, Universidad Miguel Hernández-CSIC, Spain.|Red de Trastornos Adictivos del Instituto de Salud Carlos III, Spain.
authors.forename: Jorge
authors.initials: J
authors.lastname: Manzanares
authors.affiliation: Departamento de Farmacología, Facultad de Medicina, Universidad Complutense, Instituto de Investigación Sanitaria Hospital 12 de Octubre, Spain.|Red de Trastornos Adictivos del Instituto de Salud Carlos III, Spain.
authors.forename: Esther
authors.initials: E
authors.lastname: O'Shea
authors.affiliation: Departamento de Farmacología, Facultad de Medicina, Universidad Complutense, Instituto de Investigación Sanitaria Hospital 12 de Octubre, Spain.|Red de Trastornos Adictivos del Instituto de Salud Carlos III, Spain.
authors.forename: María Isabel
authors.initials: MI
authors.lastname: Colado
mesh_terms: D000328:Adult; D000368:Aged; D000437:Alcoholism / Q000150:complications* / Q000235:genetics / Q000378:metabolism; D000818:Animals; D001344:Autopsy; D063425:Binge Drinking / Q000150:complications* / Q000235:genetics / Q000378:metabolism; D001812:Blood-Brain Barrier / Q000187:drug effects* / Q000378:metabolism / Q000503:physiopathology; D015153:Blotting, Western; D001921:Brain / Q000187:drug effects* / Q000378:metabolism / Q000503:physiopathology; D002492:Central Nervous System Depressants / Q000378:metabolism / Q000494:pharmacology; D004195:Disease Models, Animal; D000431:Ethanol / Q000378:metabolism / Q000494:pharmacology*; D005260:Female; D006801:Humans; D008297:Male; D020780:Matrix Metalloproteinase 9 / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008875:Middle Aged; D019108:Tight Junctions / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D051197:Toll-Like Receptor 4 / Q000235:genetics / Q000378:metabolism*
chemical_list: D002492:Central Nervous System Depressants; C493487:Tlr4 protein, mouse; D051197:Toll-Like Receptor 4; D000431:Ethanol; D020780:Matrix Metalloproteinase 9
keywords: MAPK; TLR4; blood-brain barrier; ethanol; human brain; metalloproteinase