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: Ganta Vijay Chaitanya, Prabhakar Eeka, Reinhold Munker, Jonathan Steven Alexander, Phanithi Prakash Babu
• Journal: Brain pathology (Zurich, Switzerland) (Brain Pathol) Vol. 21 Issue 1 Pg. 16-30 (Jan 2011) ISSN: 1750-3639 [Electronic] Switzerland
• PMID: 20825413 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: © 2010 The Authors; Brain Pathology © 2010 International Society of Neuropathology.

Chemical References

• CXCL10 protein, human
• Chemokine CXCL10
• Granzymes
• Caspase 3

Topics

• Adult
• Aged
• Analysis of Variance
• Apoptosis
• Blotting, Western
• Brain (metabolism, pathology)
• Brain Ischemia (metabolism, pathology)
• Caspase 3 (metabolism)
• Cell Line, Tumor
• Cells, Cultured
• Chemokine CXCL10 (metabolism)
• Female
• Granzymes (metabolism)
• Humans
• Immunohistochemistry
• Immunoprecipitation
• In Situ Nick-End Labeling
• Male
• Middle Aged
• Necrosis
• Nerve Degeneration (metabolism, pathology)
• Neurons (metabolism, pathology)
• Stroke (metabolism, pathology)