title: Role of cytotoxic protease granzyme-b in neuronal degeneration during human stroke.
abstract: Infiltration of leukocytes into post-ischemic cerebrum is a well-described phenomenon in stroke injury. Because CD-8(+) T-lymphocytes secrete cytotoxic proteases, including granzyme-b (Gra-b) that exacerbates post-ischemic brain damage, we investigated roles of Gra-b in human stroke. To study the role of Gra-b in stroke, ischemic and non-ischemic tissues (from post-mortem stroke patients) were analyzed using immunoblotting, co-immunoprecipitation, terminal deoxy uridine nick end labeling (TUNEL) and Annexin-V immunostaining, and in vitro neuron survival assays. Activated CG-SH cells and supernatants were used to model leukocyte-dependent injury. Non-ischemic brain tissues were used as non-pathological controls. Non-activated CG-SH cells and supernatants were used as controls for in vitro experiments. Human stroke (ischemic) samples contained significantly higher levels of Gra-b and interferon-gamma inducible protein-10 (IP-10/CXCL10) than non-ischemic controls. In stroke, poly (ADP-ribose) polymerase-1 and heat shock protein-70 were cleaved to canonical proteolytic "signature" fragments by Gra-b. Gra-b was also found to bind to Bid and caspase-3. Gra-b also co-localized with Annexin-V(+) /TUNEL(+) in degenerating neurons. Importantly, Gra-b inhibition protected both normal and ischemia-reperfused neurons against in vitro neurotoxicity mediated by activated CG-SH cells and supernatants. These results suggest that increased leukocyte infiltration and elevated Gra-b levels in the post-stroke brain can induce contact-dependent and independent post-ischemic neuronal death to aggravate stroke injury.
authors.affiliation: Department of Molecular and Cellular Physiology, School of Medicine, Feist-Weiller Cancer Center, Louisiana State University Health Sciences Center, Shreveport, La, USA.
authors.forename: Ganta Vijay
authors.initials: GV
authors.lastname: Chaitanya
authors.forename: Prabhakar
authors.initials: P
authors.lastname: Eeka
authors.forename: Reinhold
authors.initials: R
authors.lastname: Munker
authors.forename: Jonathan Steven
authors.initials: JS
authors.lastname: Alexander
authors.forename: Phanithi Prakash
authors.initials: PP
authors.lastname: Babu
mesh_terms: D000328:Adult; D000368:Aged; D000704:Analysis of Variance; D017209:Apoptosis; D015153:Blotting, Western; D001921:Brain / Q000378:metabolism* / Q000473:pathology; D002545:Brain Ischemia / Q000378:metabolism* / Q000473:pathology; D053148:Caspase 3 / Q000378:metabolism; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D054357:Chemokine CXCL10 / Q000378:metabolism; D005260:Female; D053804:Granzymes / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D047468:Immunoprecipitation; D020287:In Situ Nick-End Labeling; D008297:Male; D008875:Middle Aged; D009336:Necrosis; D009410:Nerve Degeneration / Q000378:metabolism* / Q000473:pathology; D009474:Neurons / Q000378:metabolism / Q000473:pathology; D020521:Stroke / Q000378:metabolism* / Q000473:pathology
chemical_list: C114753:CXCL10 protein, human; D054357:Chemokine CXCL10; D053804:Granzymes; D053148:Caspase 3
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