MMP-9-Positive neutrophil infiltration is associated to blood-brain barrier breakdown and basal lamina type IV collagen degradation during hemorrhagic transformation after human ischemic stroke

Background and Purpose - An abnormal expression of some matrix metalloproteinases (MMPs) is related with hemorrhagic transformation events after stroke. Our aim was to investigate MMP-2 and MMP-9 in the ischemic brain and its relation with blood-brain barrier breakdown after hemorrhagic transformation in human stroke. Methods - We assessed 5 cases of fatal ischemic strokes with hemorrhagic complications; brain samples were obtained from infarct, hemorrhagic, and contralateral tissue. MMP-9 and MMP-2 content was analyzed by zymography and immunohistochemistry was performed to localize MMP-9 and to assess collagen IV integrity in the basal lamina. Laser capture microdissection was performed to isolate blood-brain barrier vessels to study these MMPs. Results - Overall, MMP-9 levels were higher both in hemorrhagic and nonhemorrhagic infarcted tissue compared to contralateral areas (P < 0.0001 and P < 0.05). Moreover, levels of the cleaved MMP-9 85kDa-form were significantly elevated in the hemorrhagic compared to nonhemorrhagic and contralateral areas (P = 0.033 and P = 0.0001). No changes were found for MMP-2 content. Immunostaining revealed a strong MMP-9 -positive neutrophil infiltration surrounding brain microvessels associated with severe basal lamina type IV collagen degradation and blood extravasation. Microdissection confirmed that content of MMP-9 was similarly high in microvessel endothelium from hemorrhagic and infarcted areas compared to contralateral hemisphere vessels (P < 0.05), pointing to neutrophils surrounding dissected microvessels as the main source of MMP-9 in hemorrhagic areas. Conclusions - Our results show a strong neutrophil infiltration in the infarcted and hemorrhagic areas with local high MMP-9 content closely related to basal lamina collagen IV degradation and blood-brain barrier breakdown. Microvessel and inflammatory MMP-9 response are associated with hemorrhagic complications after stroke.