Involvement of caspase-1 proteases in hypoxic brain injury. Effects of their inhibitors in developing neurons

To further explore the contribution of caspase-1/interleukin-1 beta-convert ing enzyme in the consequences of hypoxia in developing brain neurons, its temporal expression profile was analysed by immunohistochemistry and wester n blotting in cultured neurons from the embryonic rat forebrain subjected t o a hypoxic stress (95% N-2/5% CO2 for 6 h), and proteolytic activity of ca spase-1 was monitored as a function of time by measuring the degradation of a selective colorimetric substrate (N-acetyl-Tyr-Val-Ala-Asp-p-nitroanilid e). In addition, the influence of pre and posthypoxic treatments by caspase -1 inhibitors (N-acetyl-Tyr-Val-Ala-Asp-aldehyde and N-acetyl-Tyr-Val-Ala-A sp-chloromethylketone) was tested on cell outcome. Hypoxia led to delayed a poptotic neuronal death, with an elevation of the expression of both pro-ca spase-1 and caspase-1 active cleavage product (ICE p20) for up to 96 h afte r cell reoxygenation. As reflected by cleavage of the specific substrate, c aspase-1 activity progressively increased between 24 h and 96 h posthypoxia , and was blocked by inhibitors in a dose-dependent fashion. The inhibitory compounds, including when given 24 h after hypoxia, prevented neuronal dea th, reduced apoptosis hallmarks and also increased the number of mitotic ne urons, suggesting they might promote neurogenesis. Similar observations wer e made when neurons were exposed to a sublethal hypoxia (i.e. 3 h). These data emphasize the participation of caspase-1 in neuronal injury cons ecutive to oxygen deprivation, and provide new insight into the possible ce llular mechanisms by which caspase inhibitors may protect developing brain neurons. (C) 1999 IBRO. Published by Elsevier Science Ltd.