title: Occludin oligomeric assembly at tight junctions of the blood-brain barrier is disrupted by peripheral inflammatory hyperalgesia.
abstract: Tight junctions (TJs) at the blood-brain barrier (BBB) dynamically alter paracellular diffusion of blood-borne substances from the peripheral circulation to the CNS in response to external stressors, such as pain, inflammation, and hypoxia. In this study, we investigated the effect of lambda-carrageenan-induced peripheral inflammatory pain (i.e., hyperalgesia) on the oligomeric assembly of the key TJ transmembrane protein, occludin. Oligomerization of integral membrane proteins is a critical step in TJ complex assembly that enables the generation of tightly packed, large multiprotein complexes capable of physically obliterating the interendothelial space to inhibit paracellular diffusion. Intact microvessels isolated from rat brains were fractionated by detergent-free density gradient centrifugation, and gradient fractions were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis/ Western blot. Injection of lambda-carrageenan into the rat hind paw produced after 3 h a marked change in the relative amounts of oligomeric, dimeric, and monomeric occludin isoforms associated with different plasma membrane lipid raft domains and intracellular compartments in endothelial cells at the BBB. Our findings suggest that increased BBB permeability (i.e., leak) associated with lambda-carrageenan-induced peripheral inflammatory pain is promoted by the disruption of disulfide-bonded occludin oligomeric assemblies, which renders them incapable of forming an impermeant physical barrier to paracellular transport.
authors.affiliation: Department of Medical Pharmacology, University of Arizona College of Medicine, Tucson, Arizona 85745, USA. gwenm@email.arizona.edu
authors.forename: Gwen
authors.initials: G
authors.lastname: McCaffrey
authors.forename: Melissa J
authors.initials: MJ
authors.lastname: Seelbach
authors.forename: William D
authors.initials: WD
authors.lastname: Staatz
authors.forename: Nicole
authors.initials: N
authors.lastname: Nametz
authors.forename: Carolyn
authors.initials: C
authors.lastname: Quigley
authors.forename: Chris R
authors.initials: CR
authors.lastname: Campos
authors.forename: Tracy A
authors.initials: TA
authors.lastname: Brooks
authors.forename: Thomas P
authors.initials: TP
authors.lastname: Davis
mesh_terms: D000818:Animals; D001693:Biological Transport, Active / Q000187:drug effects; D001812:Blood-Brain Barrier / Q000378:metabolism* / Q000503:physiopathology; D002351:Carrageenan / Q000494:pharmacology; D002451:Cell Compartmentation / Q000187:drug effects; D004058:Diffusion / Q000187:drug effects; D042783:Endothelial Cells / Q000378:metabolism / Q000648:ultrastructure; D005260:Female; D006930:Hyperalgesia / Q000139:chemically induced / Q000150:complications / Q000503:physiopathology*; D007249:Inflammation / Q000139:chemically induced / Q000150:complications / Q000503:physiopathology*; D046911:Macromolecular Substances / Q000378:metabolism; D021962:Membrane Microdomains / Q000187:drug effects / Q000378:metabolism; D008565:Membrane Proteins / Q000096:biosynthesis / Q000378:metabolism*; D008833:Microcirculation / Q000378:metabolism / Q000648:ultrastructure; D062793:Occludin; D011485:Protein Binding / Q000187:drug effects; D021122:Protein Subunits / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D019108:Tight Junctions / Q000378:metabolism* / Q000648:ultrastructure
chemical_list: D046911:Macromolecular Substances; D008565:Membrane Proteins; D062793:Occludin; C568987:Ocln protein, rat; D021122:Protein Subunits; D002351:Carrageenan