title: Exposure to lipopolysaccharide and/or unconjugated bilirubin impair the integrity and function of brain microvascular endothelial cells.
abstract: BACKGROUND
  Sepsis and jaundice are common conditions in newborns that can lead to brain damage. Though lipopolysaccharide (LPS) is known to alter the integrity of the blood-brain barrier (BBB), little is known on the effects of unconjugated bilirubin (UCB) and even less on the joint effects of UCB and LPS on brain microvascular endothelial cells (BMEC).
  
  
  RESULTS
  Monolayers of primary rat BMEC were treated with 1 µg/ml LPS and/or 50 µM UCB, in the presence of 100 µM human serum albumin, for 4 or 24 h. Co-cultures of BMEC with astroglial cells, a more complex BBB model, were used in selected experiments. LPS led to apoptosis and UCB induced both apoptotic and necrotic-like cell death. LPS and UCB led to inhibition of P-glycoprotein and activation of matrix metalloproteinases-2 and -9 in mono-cultures. Transmission electron microscopy evidenced apoptotic bodies, as well as damaged mitochondria and rough endoplasmic reticulum in BMEC by either insult. Shorter cell contacts and increased caveolae-like invaginations were noticeable in LPS-treated cells and loss of intercellular junctions was observed upon treatment with UCB. Both compounds triggered impairment of endothelial permeability and transendothelial electrical resistance both in mono- and co-cultures. The functional changes were confirmed by alterations in immunostaining for junctional proteins β-catenin, ZO-1 and claudin-5. Enlargement of intercellular spaces, and redistribution of junctional proteins were found in BMEC after exposure to LPS and UCB.
  
  
  CONCLUSIONS
  LPS and/or UCB exert direct toxic effects on BMEC, with distinct temporal profiles and mechanisms of action. Therefore, the impairment of brain endothelial integrity upon exposure to these neurotoxins may favor their access to the brain, thus increasing the risk of injury and requiring adequate clinical management of sepsis and jaundice in the neonatal period.
authors.affiliation: Research Institute for Medicines and Pharmaceutical Sciences, Faculty of Pharmacy, University of Lisbon, Lisbon, Portugal.
authors.forename: Filipa L
authors.initials: FL
authors.lastname: Cardoso
authors.forename: Agnes
authors.initials: A
authors.lastname: Kittel
authors.forename: Szilvia
authors.initials: S
authors.lastname: Veszelka
authors.forename: Inês
authors.initials: I
authors.lastname: Palmela
authors.forename: Andrea
authors.initials: A
authors.lastname: Tóth
authors.forename: Dora
authors.initials: D
authors.lastname: Brites
authors.forename: Mária A
authors.initials: MA
authors.lastname: Deli
authors.forename: Maria A
authors.initials: MA
authors.lastname: Brito
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001253:Astrocytes / Q000187:drug effects / Q000648:ultrastructure; D001663:Bilirubin / Q000494:pharmacology; D001812:Blood-Brain Barrier / Q000187:drug effects / Q000648:ultrastructure; D001921:Brain / Q000098:blood supply / Q000378:metabolism; D002463:Cell Membrane Permeability / Q000187:drug effects; D002478:Cells, Cultured; D062507:Claudin-5; D057167:Claudins / Q000378:metabolism; D018920:Coculture Techniques; D018870:Endoplasmic Reticulum, Rough / Q000187:drug effects; D042783:Endothelial Cells / Q000187:drug effects / Q000648:ultrastructure; D008070:Lipopolysaccharides / Q000494:pharmacology; D008565:Membrane Proteins / Q000378:metabolism; D046529:Microscopy, Electron, Transmission; D055806:Microvessels / Q000187:drug effects / Q000648:ultrastructure; D008928:Mitochondria / Q000187:drug effects; D010750:Phosphoproteins / Q000378:metabolism; D051381:Rats; D062826:Zonula Occludens-1 Protein; D051176:beta Catenin / Q000378:metabolism
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D062507:Claudin-5; D057167:Claudins; C491510:Cldn5 protein, rat; D008070:Lipopolysaccharides; D008565:Membrane Proteins; D010750:Phosphoproteins; C569042:TJP1 protein, human; C569044:Tjp1 protein, rat; D062826:Zonula Occludens-1 Protein; D051176:beta Catenin; D001663:Bilirubin