Estrogen-induced neuroprotection is mediated by an increase in caspase-12-associated apoptosis following MCAO

Estrogen has received considerable attention over the past decade as a potential therapeutic agent against various forms of neurodegenerative diseases including stroke. Experimental data in animal models of stroke have provided exhaustive evidence of the neuroprotective properties of this steroid hormone. Our laboratory in particular has demonstrated that estrogen significantly reduces (∼50%) ischemic cell death resulting from permanent occlusion of the middle cerebral artery (MCAO). However, the cellular and molecular mechanisms implicated in the protective actions of estrogen in this experimental model have yet to be elucidated. Accumulating evidence suggests that endoplasmic reticulum stress-induced apoptosis is linked to ischemic cell death and that estrogen interacts with various components of the endoplasmic reticulum stress response. We therefore hypothesized that estrogen enhances endoplasmic reticulum stress-associated apoptosis via activation of caspase-12 to decrease necrotic cell death, thus limiting infarct volume. Estrogen significantly decreased procaspase-12 expression in both the infarct and peri-infarct regions at 1 hour post-MCAO and this corresponded with an increase in active caspase-12 at 2 and 3 hours post-MCAO. An increase in m-calpain expression was observed in the infarct region at 1 hour post-MCAO with estrogen treatment, suggesting m-calpain may play a role in activating caspase-12 following MCAO. TUNEL staining of brain sections showed a significantly higher number of TUNEL positive cells in estrogen treated animals at 4 hours past-MCAO compared to those treated with saline. The amount of necrotic cell death following MCAO was assessed by the presence of lactate dehydrogenase (LDH) in the cerebrospinal fluid (CSF) surrounding the frontal cortex and the results indicated a significant elevation in LDH levels following MCAO compared to sham occlusion. However, estrogen treatment did not decrease the concentration of LDH in the CSF following MCAO when compared to saline treatment. Overall, these data are consistent with the hypothesis that the protective actions of estrogen observed following permanent MCAO in male rats involve an increase in caspase-12 mediated apoptotic cell death.