HMGB1/TLR4 axis promotes pyroptosis after ICH by activating the NLRP3 inflammasome

We previously reported that the HMGB1/TLR4 axis promoted inflammation during the acute phase of intracerebral hemorrhage. Given that this phase is known to involve neuronal pyroptosis and neuroinflammation, here we explore whether HMGB1/TLR signaling activate inflammasome and pyroptosis after intracerebral hemorrhage.

Autologous blood was injected into Sprague-Dawley rats to induce intracerebral hemorrhage. Neurological deficits were assessed using a modified neurological severity score. These expression and localization of NLRP1 and NLRP3 inflammasomes, as well as the levels of pyroptosis and pyroptosis-associated proteins were assessed using Western blot or immunocytochemistry. These experiments were repeated in animals that received treatment with short interfering RNAs against NLRP1 or NLRP3, with HMGB1 inhibitor ethyl pyruvate or TLR4 inhibitor TAK-242.

Intracerebral hemorrhage upregulated NLRP1 and NLRP3 in the ipsilateral striatum and increased the proportions of these cells that were pyroptosis-positive. Additionally, the levels of caspase protein family (e.g., pro-caspase-1 and caspase-1), apoptosis-associated speck-like protein (ASC), pro-interleukin-1β (IL-1β), and IL-1β were also elevated. These effects on pyroptosis and associated neurological deficit, were partially reversed by knockdown of NLRP1 or NLRP3, or by inhibition of HMGB1 or TLR4. Inhibition of HMGB1 or TLR4 resulted in the downregulation NLRP3 but not NLRP1.

The HMGB1/TLR4 signaling may activate the NLRP3 inflammasome during the acute phase of intracerebral hemorrhage, resulting in the inflammatory process known as pyroptosis. These insights suggest potential therapeutic targets for the mitigation tissue injury and associated neurological deficits following hemorrhagic stroke.