# Caspase‐1 inhibition prevents neuronal death by targeting the canonical inflammasome pathway of pyroptosis in a murine model of cerebral ischemia

## TL;DR

This study shows that inhibiting caspase-1 with Vx765 reduces neuronal death and improves recovery in a mouse model of stroke by blocking pyroptosis.

## Contribution

The study demonstrates that Vx765, a clinically tested drug, can target pyroptosis to reduce brain injury in cerebral ischemia.

## Key findings

- Vx765 treatment significantly reduced infarct volumes in mice at multiple time points after stroke.
- Vx765 improved motor recovery and preserved neuronal structures by inhibiting pyroptosis-related proteins.
- Elevated levels of GSDMD, inflammasomes, and IL-1β were observed in ischemic regions and reduced by Vx765.

## Abstract

The involvement of pyroptosis in ischemic stroke remains to be established. Therefore, we used the specific pyroptosis inhibitor Vx765 as an experimental intervention target in a murine model of stroke.

A total of 564 C57BL/6 mice were subjected to photothrombotic procedures and treated via gavage with Vx765 at 1‐hour post‐ischemia. We subsequently assessed the expression of Gasdermin D (GSDMD), inflammasomes, caspase‐1, and interleukin‐1β (IL‐1β) using immunofluorescence (IF) and Western blot (WB) analyses. We also examined ultrastructural changes of cortical neurons with transmission electron microscopy (TEM) and measured infarct volumes dynamically by magnetic resonance imaging (MRI). Moreover, we evaluated the neurologic deficits by modified neurological severity scores, the rotarod test, and Treadscan.

Elevated expression of GSDMD and GSDMD p30, the pore‐forming subunit, was evident in the peri‐ischemic region on days one and three post‐ischemia. The neuronal plasma, nuclear, and mitochondrial membranes showed ultrastructural damage at day three post‐stroke. Elevated expression of inflammasomes, caspase‐1, and IL‐1β was also present on days one and three post‐injury. There were significant differences between Vx765‐treated and vehicle groups in mean infarct volumes (14.36 vs 21.52 mm3; 12.34 vs 18.56 mm3; 4.13 vs 10.06 mm3; P < .05 at day one, three, and seven post‐surgery, respectively). Mice treated with Vx765 showed better motor recovery as assessed by serial behavior tests and had better neuronal survival, which was attributable to pyroptosis inhibition, as illustrated by downregulated expression of the effector protein GSDMD, inflammasomes, caspase‐1, and IL‐1β. Besides, treatment with Vx765 preserved neuronal membrane structures after the ischemic injury.

Pyroptosis emerges as an important pathway for neuronal death in an acute ischemic stroke. Vx765, a low molecular weight drug that has proven safe in clinical epilepsy trials, has potential therapeutic value for cerebral ischemia by targeting the canonical inflammasome pathway of pyroptosis.