Open AccessRole of Cytotoxic Protease Granzyme‐b in Neuronal Degeneration During Human Stroke
Author(s) -
Chaitanya Ganta Vijay,
Eeka Prabhakar,
Munker Reinhold,
Alexander Jonathan Steven,
Babu Phanithi Prakash
Publication year - 2011
Publication title -
brain pathology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.986
H-Index - 132
eISSN - 1750-3639
pISSN - 1015-6305
DOI - 10.1111/j.1750-3639.2010.00426.x
Subject(s) - granzyme b , tunel assay , biology , pathology , annexin , proteases , apoptosis , microbiology and biotechnology , cytotoxic t cell , in vitro , medicine , biochemistry , enzyme
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.