title: Dynamic platelet accumulation at the site of the occluded middle cerebral artery and in downstream microvessels is associated with loss of microvascular integrity after embolic middle cerebral artery occlusion.
abstract: Information is lacking regarding dynamic platelet accumulation at the site of the occluded middle cerebral artery (MCA) and the relationship between platelet aggregation in downstream cerebral microvessels and loss of perfusion and vascular integrity of these microvessels. In the present study, we employed a model of embolic MCA occlusion in the rat to simultaneously measure temporal and spatial profiles of platelet accumulation at the site of the embolus occluding the MCA and within downstream cerebral microvessels. We also measured the integrity of microvessels and matrix metalloproteinase (MMP) activity in ischemic brain. Rats (n=36) were subjected to embolic MCA occlusion. Immunohistochemistry was used to detect microvascular integrity, plasminogen activator inhibitor 1 (PAI-1) and the deposition of fibrin. SDS-PAGE zymography was used to measure MMP2 and MMP9 activities. Accumulation of platelets and increases in PAI-1 immunoreactivity at the site of the embolus occluding the MCA were detected 1 h (n=7) and 4 h (n=7) after ischemia, respectively, and numbers of GPIIb/IIIa immunoreactive downstream cerebral microvessels increased significantly (209+/-59; n=7; P<0.05) 4 h after ischemia, suggesting dynamic platelet aggregation. A significant (n=7; P<0.01) diffuse loss of type IV collagen immunoreactivity in microvessels was temporally associated with platelet GPIIb/IIIa immunoreactivity within the vessels. Triple immunostaining revealed that microvessels containing platelet aggregates exhibited loss of type IV collagen immunoreactivity and both intra- and extra-vascular fibrin deposition, suggesting that intravascular platelet aggregation is associated with decreases in the integrity of the microvascular basal lamina and blood-brain barrier leakage. A significant increase (P<0.05) in MMP9 was detected at 4 h (n=3) and 24 h (n=3) after ischemia but levels of MMP2 were not significantly changed in ischemic brain. Our data suggest that dynamic platelet aggregation in ischemic brain may contribute to time-dependent resistance to fibrinolysis. In addition, platelet deposition and increased MMP9 coincided with degradation of type IV collagen and loss of vascular integrity. These data suggest an important role for post-occlusive distal platelet deposition in the pathophysiology of stroke.
authors.affiliation: Department of Neurology, Henry Ford Health Sciences Center, 2799 West Grand Boulevard, Detroit, MI 48202, USA.
authors.forename: Z G
authors.initials: ZG
authors.lastname: Zhang
authors.forename: L
authors.initials: L
authors.lastname: Zhang
authors.forename: W
authors.initials: W
authors.lastname: Tsang
authors.forename: A
authors.initials: A
authors.lastname: Goussev
authors.forename: C
authors.initials: C
authors.lastname: Powers
authors.forename: K L
authors.initials: KL
authors.lastname: Ho
authors.forename: D
authors.initials: D
authors.lastname: Morris
authors.forename: S S
authors.initials: SS
authors.lastname: Smyth
authors.forename: B S
authors.initials: BS
authors.lastname: Coller
authors.forename: M
authors.initials: M
authors.lastname: Chopp
mesh_terms: D000818:Animals; D001777:Blood Coagulation / Q000502:physiology*; D001792:Blood Platelets / Q000166:cytology / Q000502:physiology* / Q000648:ultrastructure; D001812:Blood-Brain Barrier / Q000502:physiology; D001921:Brain / Q000201:enzymology* / Q000473:pathology / Q000648:ultrastructure; D002545:Brain Ischemia / Q000201:enzymology* / Q000473:pathology / Q000503:physiopathology; D002560:Cerebrovascular Circulation / Q000502:physiology; D024141:Collagen Type IV / Q000378:metabolism; D004195:Disease Models, Animal; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D020244:Infarction, Middle Cerebral Artery / Q000201:enzymology / Q000473:pathology / Q000503:physiopathology*; D007797:Laminin / Q000378:metabolism; D008297:Male; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism; D020782:Matrix Metalloproteinases / Q000378:metabolism*; D008833:Microcirculation / Q000473:pathology / Q000503:physiopathology* / Q000648:ultrastructure; D008854:Microscopy, Electron; D017711:Nonlinear Dynamics; D017395:Plasminogen Activator Inhibitor 1 / Q000378:metabolism; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D013997:Time Factors
chemical_list: D024141:Collagen Type IV; D007797:Laminin; D017395:Plasminogen Activator Inhibitor 1; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D020782:Matrix Metalloproteinases; D020778:Matrix Metalloproteinase 2; D020780:Matrix Metalloproteinase 9