Matrix metalloproteinase-mediated disruption of tight junction proteins in cerebral vessels is reversed by synthetic matrix metalloproteinase inhibitor in focal ischemia in rat.
Matrix metalloproteinases (MMPs) disrupt the blood-brain barrier (BBB) during reperfusion. Occludin and claudins are recently described tight junction proteins (TJPs) that form the BBB. We hypothesized that the opening of the BBB was because of the degradation of TJPs by the MMPs. Spontaneously hypertensive rats had a 90 mins middle cerebral artery occlusion with reperfusion for 2, 3, or 24 h. Matrix metalloproteinases were measured by immunohistochemistry and in situ and gel zymography. Real-time polymerase chain reaction (PCR) measured mRNAs of MMP-2 and -9, furin, membrane-type MMP (MT1-MMP), occludin, and claudin-5. There was opening of the BBB in the piriform cortex after 3 h of reperfusion, and an MMP inhibitor, BB-1101 (30 mg/kg), prevented the opening. At 3 h, in situ zymograms showed gelatinase activity. Zymography and PCR showed greater increases in MMP-2 than in MMP-9. There were increased mRNA and immunohistochemistry for MT1-MMP and furin, which activate MMP-2. Claudin-5 and occludin mRNA expression decreased at 2 h in both hemispheres with fragments of both proteins seen on Western blot by 3 h on the ischemic side; treatment with BB-1101 reversed the degradation of the TJPs. Immunohistochemistry at 3 h showed fragmented TJPs within the endothelial cell clefts. By 24 h, in situ zymography showed gelatinase activity and gel zymography showed elevated levels of MMP-9. Disrupted TJPs previously seen in endothelial cells appeared in the surrounding astrocytes. Our results provide direct evidence that MMPs open the BBB by degrading TJPs and that an MMP inhibitor prevents degradation of the TJPs by MMPs.
Yang
Yi
Y
Department of Neurology, University of New Mexico Health Sciences Center, Albuquerque, New Mexico 87131-0001, USA.
Estrada
Eduardo Y
EY
Thompson
Jeffrey F
JF
Liu
Wenlan
W
Rosenberg
Gary A
GA
D000818:Animals; D001253:Astrocytes / Q000187:drug effects / Q000378:metabolism; D001812:Blood-Brain Barrier / Q000187:drug effects; D015153:Blotting, Western; D002545:Brain Ischemia / Q000188:drug therapy* / Q000378:metabolism; D002536:Cerebral Arteries / Q000187:drug effects / Q000378:metabolism*; D002550:Cerebral Veins / Q000187:drug effects / Q000378:metabolism*; D062507:Claudin-5; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism; D020536:Enzyme Activators / Q000494:pharmacology; D005070:Evans Blue; D045683:Furin / Q000096:biosynthesis; D018093:Gelatinases / Q000378:metabolism; D007150:Immunohistochemistry; D020244:Infarction, Middle Cerebral Artery / Q000473:pathology; D008297:Male; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D061965:Matrix Metalloproteinase Inhibitors; D020782:Matrix Metalloproteinases / Q000096:biosynthesis / Q000502:physiology*; D008565:Membrane Proteins / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D062793:Occludin; D011480:Protease Inhibitors / Q000494:pharmacology*; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D011918:Rats, Inbred SHR; D015427:Reperfusion Injury / Q000473:pathology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013395:Sucrose; D019108:Tight Junctions / Q000187:drug effects / Q000378:metabolism*
D062507:Claudin-5; C491510:Cldn5 protein, rat; D020536:Enzyme Activators; D061965:Matrix Metalloproteinase Inhibitors; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D062793:Occludin; C568987:Ocln protein, rat; D011480:Protease Inhibitors; D012333:RNA, Messenger; D005070:Evans Blue; D013395:Sucrose; D045683:Furin; D018093:Gelatinases; D020782:Matrix Metalloproteinases; D020778:Matrix Metalloproteinase 2