title: Glial cells drive preconditioning-induced blood-brain barrier protection.
abstract: OBJECTIVE
  The cerebrovascular contribution to ischemic preconditioning (IPC) has been scarcely explored. Using in vivo and in vitro approaches, we investigated the involvement of the blood-brain barrier and the role of its cellular components.
  
  
  METHODS
  Seven-minute occlusion of the right middle cerebral artery, used as in vivo IPC stimulus 4 days before permanent occlusion of the right middle cerebral artery, significantly reduced brain infarct size (8.45±0.7 versus 13.61±0.08 mm3 measured 7 days after injury) and preserved blood-brain barrier function (Evans blue leakage, 0.54±0.1 versus 0.89±0.1 ng/mg). Assessment of neuronal, endothelial, and glial gene expression revealed that IPC specifically increased glial fibrillary acidic protein mRNA, thus showing selective astrocyte activation in IPC-protected mice.
  
  
  RESULTS
  The blood-brain barrier was modeled by coculturing murine primary brain microvessel endothelial and astroglial cells. One-hour oxygen-glucose deprivation (OGD), delivered 24 hours before a 5-hour OGD, acted as an IPC stimulus, significantly attenuating the reduction in transendothelial electric resistance (199.17±11.7 versus 97.72±3.4 Ωcm2) and the increase in permeability coefficients for sodium fluorescein (0.98±0.11×10(-3) versus 1.8±0.36×10(-3) cm/min) and albumin (0.12±0.01×10(-3) versus 0.29±0.07×10(-3) cm/min) induced by severe OGD. IPC also prevented the 5-hour OGD-induced disorganization of the tight junction proteins ZO-1 and claudin-5. IPC on glial (but not endothelial) cells alone preserved transendothelial electric resistance, permeability coefficients, and ZO-1 localization after 5 hours of OGD. Astrocyte metabolic inhibition by fluorocitrate abolished IPC protection, confirming the critical role of astrocytes. IPC significantly increased glial fibrillary acidic protein, interleukin-6, vascular endothelial growth factor-a, and ciliary neurotrophic factor gene expression after OGD in glial cells, indicating that multiple pathways mediate the glial contribution to IPC.
  
  
  CONCLUSIONS
  Our data show that the blood-brain barrier can be directly preconditioned and that astrocytes are major mediators of IPC protection.
authors.affiliation: Laboratory of Inflammation and Nervous System Diseases, Mario Negri Institute, via G. La Masa 19, 20156 Milan, Italy.
authors.forename: Raffaella
authors.initials: R
authors.lastname: Gesuete
authors.forename: Franca
authors.initials: F
authors.lastname: Orsini
authors.forename: Elisa R
authors.initials: ER
authors.lastname: Zanier
authors.forename: Diego
authors.initials: D
authors.lastname: Albani
authors.forename: Maria A
authors.initials: MA
authors.lastname: Deli
authors.forename: Gianfranco
authors.initials: G
authors.lastname: Bazzoni
authors.forename: Maria-Grazia
authors.initials: MG
authors.lastname: De Simoni
mesh_terms: D000818:Animals; D001253:Astrocytes / Q000378:metabolism / Q000473:pathology; D001812:Blood-Brain Barrier / Q000502:physiology*; D002545:Brain Ischemia / Q000378:metabolism / Q000473:pathology* / Q000503:physiopathology*; D062507:Claudin-5; D018920:Coculture Techniques; D004195:Disease Models, Animal; D004730:Endothelium, Vascular / Q000378:metabolism / Q000473:pathology; D005904:Glial Fibrillary Acidic Protein; D020244:Infarction, Middle Cerebral Artery; D019194:Ischemic Preconditioning; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D009419:Nerve Tissue Proteins / Q000378:metabolism; D009457:Neuroglia / Q000473:pathology / Q000502:physiology*; D010750:Phosphoproteins / Q000378:metabolism; D062826:Zonula Occludens-1 Protein
chemical_list: D062507:Claudin-5; C491511:Cldn5 protein, mouse; D005904:Glial Fibrillary Acidic Protein; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D010750:Phosphoproteins; C569043:Tjp1 protein, mouse; D062826:Zonula Occludens-1 Protein; C528900:glial fibrillary astrocytic protein, mouse