Background Oxidative stress (OS) and inflammatory reactions play pivotal roles in secondary brain injury after traumatic brain injury (TBI). Histone deacetylase 3 (HDAC3) controls the acetylation of histones and non-histones, which has a significant impact on the central nervous system’s reaction to damage. This research determined the implications of RGFP966, a new and specific inhibitor of HDAC3, for the antioxidant (AO) systems mediated by nuclear factor erythroid2-related factor 2 (Nrf2) and the Nod-like receptor protein 3 (NLRP3) inflammasome in TBI. The study also studied the underlying mechanisms of RGFP966’s actions. Our objective was to examine the impacts and underlying RGFP966 mechanisms in TBI. Methods In vitro, a rat cortical neuron OS model was induced by H2O2, followed by the addition of RGFP966 to the culture medium. Neurons were collected after 24 h for western blot (WB), terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and 2′-7′-dichlorodihydrofluorescein diacetate staining. In vivo, RGFP966 (10 mg/kg) was administered post-TBI. Brain tissue water content and modified neurological severity scores were assessed 72 h post-injury. Cortical tissues surrounding the focal injury were subjected to western blot, TUNEL staining, Nissl staining and immunofluorescence/immunohistochemistry staining, and malondialdehyde level, hindered glutathione content and superoxide dismutase activity were measured. Serum was collected for the enzyme-linked immunosorbent assay. Nrf2-specific shRNA lentivirus was injected into the lateral ventricle of rats for 7 days, and cerebral cortex tissue was analyzed by WB and real-time polymerase chain reaction. Results During in vitro and in vivo experiments, RGFP966 suppressed HDAC3 expression, promoted Nrf2 nuclear translocation, activated downstream AO enzymes, mitigated excessive reactive oxygen species production and alleviated nerve cell apoptosis. RGFP966 effectively reduced brain edema and histological damage and enhanced neurological and cognitive function in rats with TBI. RGFP966 markedly inhibited NLRP3 inflammasome activation mediated by high-mobility group box 1 (HMGB1)/toll-like receptor 4 (TLR4). Nrf2 knockdown in TBI rats attenuated the AO and anti-inflammatory, neuroprotective impacts of RGFP966. Conclusions Overall, our findings demonstrate that RGFP966 can mitigate the first brain damage and neurological impairments in TBI. The underlying mechanism involves triggering the Nrf2-mediated AO system and negatively regulating the HMGB1/TLR4-mediated NLRP3 inflammasome pathway.

中文翻译：

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组蛋白脱乙酰酶 3 特异性抑制剂 RGFP966 通过激活 Nrf2 通路减轻创伤性脑损伤后的氧化应激和炎症


背景氧化应激（OS）和炎症反应在创伤性脑损伤（TBI）后继发性脑损伤中发挥着关键作用。组蛋白脱乙酰酶 3 (HDAC3) 控制组蛋白和非组蛋白的乙酰化，这对中枢神经系统对损伤的反应具有重大影响。这项研究确定了 RGFP966（一种新的 HDAC3 特异性抑制剂）对 TBI 中由核因子红细胞 2 相关因子 2 (Nrf2) 和 Nod 样受体蛋白 3 (NLRP3) 炎性体介导的抗氧化 (AO) 系统的影响。该研究还研究了 RGFP966 作用的潜在机制。我们的目标是研究 TBI 中的影响和潜在的 RGFP966 机制。方法在体外，采用H2O2诱导大鼠皮质神经元操作系统模型，然后在培养基中添加RGFP966。 24小时后收集神经元进行蛋白质印迹（WB）、末端脱氧核苷酸转移酶dUTP缺口末端标记（TUNEL）和2'-7'-二氯二氢荧光素二乙酸酯染色。在体内，TBI 后施用 RGFP966 (10 mg/kg)。损伤后 72 小时评估脑组织含水量和修正的神经严重程度评分。对病灶周围皮质组织进行蛋白质印迹、TUNEL染色、尼氏染色和免疫荧光/免疫组化染色，并测定丙二醛水平、受阻性谷胱甘肽含量和超氧化物歧化酶活性。收集血清用于酶联免疫吸附测定。将Nrf2特异性shRNA慢病毒注射到大鼠侧脑室7天，通过WB和实时聚合酶链反应对大脑皮质组织进行分析。 结果在体外和体内实验中，RGFP966 抑制 HDAC3 表达，促进 Nrf2 核转位，激活下游 AO 酶，减轻过多的活性氧产生并减轻神经细胞凋亡。 RGFP966有效减少TBI大鼠的脑水肿和组织学损伤，增强神经和认知功能。 RGFP966 显着抑制由高迁移率族盒 1 (HMGB1)/toll 样受体 4 (TLR4) 介导的 NLRP3 炎性体激活。 TBI 大鼠中的 Nrf2 敲低减弱了 RGFP966 的 AO 和抗炎、神经保护作用。结论 总的来说，我们的研究结果表明 RGFP966 可以减轻 TBI 中的首次脑损伤和神经损伤。其潜在机制涉及触发 Nrf2 介导的 AO 系统并负向调节 HMGB1/TLR4 介导的 NLRP3 炎性体途径。