Long-term alcohol exposure aggravates ischemic stroke-induced damage by promoting pericyte NLRP3 inflammasome activation via the TLR4/NF-κB pathway in rats

Background: Alcohol consumption is a common lifestyle worldwide and a manageable risk factor for ischemic stroke. Chronic ethanol exposure reportedly causes blood-brain barrier (BBB) dysfunction and neuroinflammation. Pericytes participate in neuroinflammation during cerebral ischemia. However, whether pericytes are involved in alcohol-induced cerebral ischemia/reperfusion injury remains unclear. Therefore, we investigated the activation of Nod-like receptor protein 3 (NLPR3) inflammasome and Toll-like receptor 4 (TLR4)/ nuclear factor kappa-B (NF-κB) pathway in the pericytes in vivo and in vitro. Methods: Rat models of long-term alcohol intake followed by transient middle cerebral artery occlusion stroke (EtOH+tMCAO group) and cell models of oxygen-glucose deprivation/reoxygenation (OGD/R) with alcohol co-treatment were constructed. Results: Worsened infarct volume, neurological scores, and BBB disruption were observed after reperfusion in the EtOH+tMCAO group. Immunofluorescence staining showed increased NLPR3 inflammasome activation in the pericytes at the ischemic penumbra in the alcohol intake group than in the control group. Western blotting showed increased expression of the NLRP3 inflammasome in the pericytes after OGD/R that was significantly amplified by alcohol pretreatment. Alcohol-activated TLR4 in the pericytes in vivo and elevated levels of TLR4/NF-κB pathway-associated protein were observed in the pericytes following alcohol treatment in vitro. Transfecting with TLR4-short hairpin RNA to block TLR4 signaling in OGD/R models with alcohol co-treatment markedly reversed NLRP3 inflammasome activation. In vivo, TAK-242 inhibited TLR4 expression in the EtOH+tMCAO group and alcohol-mediated neurological impairment was significantly alleviated. Conclusion: Long-term alcohol pretreatment aggravated ischemic stroke-induced brain damage by activating NLRP3 inflammasome and TLR4/NF-κB signaling pathway in the pericytes.