Abstract：
Pyroptosis has been proven to be responsible for secondary brain injury after intracerebral hemorrhage (ICH). A recent study reported that Raf kinase inhibitor protein (RKIP) inhibited assembly and activation of inflammasome in macrophages. Our present study aimed to investigate the effects of RKIP on inflammasome-mediated neuronal pyroptosis and underlying neuroprotective mechanisms in experimental ICH. Here, we showed that RKIP expression was decreased both in cerebrospinal fluid (CSF) samples from patients with ICH and in the peri-hematoma tissues after experimental ICH. In mouse ICH model, activation of RKIP remarkably improved neurological deficits, reduced brain water content and BBB disruption, and promoted hematoma absorption at 24 h after ICH, as well as alleviated neuronal degeneration, reduced membrane pore formation, and downregulated pyroptotic molecules NLRP3, caspase-1 P20, GSDMD-N, and mature IL-1β. Besides, RKIP activation decreased the number of caspase-1 P20-positive neurons after ICH. However, RKIP inhibitor reserved the neuroprotective effects of RKIP at 24 h following ICH. Moreover, RKIP could bind with ASC, then interrupt the assembly of NLRP3 inflammasome. Mechanistically, inhibiting the caspase-1 by VX-765 attenuated brain injury and suppressed neuronal pyroptosis after RKIP inhibitor-pretreated ICH. In conclusion, our findings indicated that activation of RKIP could attenuate neuronal pyroptosis and brain injury after ICH, to some extent, through ASC/Caspase-1/GSDMD pathway. Thus, RKIP may be a potential target to attenuate brain injury via its anti-pyroptosis effect after ICH.
摘要：
已证明，发热是脑出血（ICH）后继发性脑损伤的原因。最近的一项研究报道，Raf激酶抑制剂蛋白（RKIP）抑制巨噬细胞中炎性小体的组装和活化。我们本研究旨在研究RKIP对实验性ICH中炎性小体介导的神经元焦亡和潜在的神经保护机制的影响。这里,我们显示,RKIP在ICH患者的脑脊液(CSF)和实验性ICH后的血肿周围组织中的表达均降低.在小鼠ICH模型中，RKIP的激活显著改善了神经功能缺损，减少脑含水量和血脑屏障破坏，并在ICH后24h促进血肿吸收，以及减轻神经元变性，减少膜孔形成，和下调的热解分子NLRP3，caspase-1P20，GSDMD-N，和成熟的IL-1β。此外,RKIP激活减少了ICH后caspase-1P20阳性神经元的数量。然而,RKIP抑制剂在ICH后24小时保留了RKIP的神经保护作用。此外，RKIP可以与ASC绑定，然后中断NLRP3炎性体的组装。机械上,通过VX-765抑制caspase-1减轻了RKIP抑制剂预处理的ICH后的脑损伤并抑制了神经元的焦亡。总之,我们的发现表明RKIP的激活可以减轻ICH后神经元的焦亡和脑损伤，在某种程度上,通过ASC/Caspase-1/GSDMD途径。因此，RKIP可能是减轻脑出血后脑损伤的潜在靶点。
