Department of Anesthesiology, The Second Hospital of Tianjin Medical University, Tianjin 300211, China; Department of Anesthesiology, Tianjin Medical University General Hospital, Tianjin 300052, China.
Department of Anesthesiology, Tianjin Medical University General Hospital, Tianjin 300052, China; Tianjin Research Institute of Anesthesiology, Tianjin 300052, China.
Int Immunopharmacol. 2021 Jun;95:107583. doi: 10.1016/j.intimp.2021.107583. Epub 2021 Mar 24.
Sepsis-associated encephalopathy (SAE) can cause acute and long-term cognitive impairment and increase the mortality rate in sepsis patients, and we previously reported that 2% hydrogen gas (H) inhalation has a therapeutic effect on SAE, but the underlying mechanism remains unclear. Dynamic DNA methylation, which catalyzed by DNA methyltransferases (DNMTs), is involved in the formation of synaptic plasticity and cognitive memory in the central nervous system. And brain-derived neurotrophic factor (BDNF), to be a key signaling component in activity-dependent synaptic plasticity, can be induced by neuronal activity accompanied by hypomethylation of its promoter IV. This study was designed to illustrate whether H can mediate SAE by alter the BDNF promoter IV methylation mediated by DNMTs. We established an SAE model by cecal ligation and perforation (CLP) in C57BL/6 mice. The Morris water maze test from the 4th to the 10th day after sham or CLP operations were used to evaluate mouse cognitive function. Hippocampal tissues were isolated at the 24 after sham or CLP surgery. Pro-inflammatory cytokines including tumor necrosis factor-α (TNF-α), Interleukin-6 (IL-6) and High Mobility Group Box 1 (HMGB1) were measured by enzyme-linked immunosorbent assay (ELISA). mRNA or protein levels of DNMTs (DNMT1, DNMT3a and DNMT3b), BDNF promoter IV and total BDNF were detected by RT-PCR and Western blot tests. Immunofluorescence staining were used to determine the expressions of DNMT1 and DNMT3a. The quantitative methylation analysis of the 11 CpG island of the promoter region of BDNF exon IV was determined using theAgena's MassARRAY EpiTYPER system. We found that 2% H inhalation can reduce pro-inflammatory factors, alleviate DNMT1, DNMT3a but not DNMT3b expression, make hypomethylation of BDNF promoter IV at 5 CpG sites, enhance the BDNF levels and then decrease escape latency but increase platform crossing times in septic mice. Our results suggest that 2% H inhalation may alleviate SAE through altering the regulation of BDNF promoter IV methylation which mediated by DNMT1 and DNMT3a in the hippocampus of septic mice.
脓毒症相关性脑病（SAE）可导致脓毒症患者急性和长期认知障碍，并增加死亡率，我们之前报道过 2%氢气（H）吸入对 SAE 具有治疗作用，但潜在机制尚不清楚。动态 DNA 甲基化由 DNA 甲基转移酶（DNMTs）催化，参与中枢神经系统中突触可塑性和认知记忆的形成。脑源性神经营养因子（BDNF）作为活性依赖性突触可塑性的关键信号成分，可以在其启动子 IV 低甲基化的情况下被神经元活动诱导。本研究旨在阐明 H 是否可以通过改变 DNMTs 介导的 BDNF 启动子 IV 甲基化来介导 SAE。我们通过盲肠结扎和穿孔术（CLP）在 C57BL/6 小鼠中建立 SAE 模型。假手术或 CLP 手术后第 4 至 10 天进行 Morris 水迷宫测试，以评估小鼠的认知功能。假手术或 CLP 手术后第 24 天分离海马组织。通过酶联免疫吸附试验（ELISA）测量促炎细胞因子，包括肿瘤坏死因子-α（TNF-α）、白细胞介素-6（IL-6）和高迁移率族蛋白 B1（HMGB1）。通过 RT-PCR 和 Western blot 试验检测 DNMTs（DNMT1、DNMT3a 和 DNMT3b）、BDNF 启动子 IV 和总 BDNF 的 mRNA 或蛋白水平。通过免疫荧光染色确定 DNMT1 和 DNMT3a 的表达。使用 Agena 的 MassARRAY EpiTYPER 系统对 BDNF 外显子 IV 启动子区域的 11 个 CpG 岛进行定量甲基化分析。我们发现 2%H 吸入可以降低促炎因子，减轻 DNMT1、DNMT3a 但不减轻 DNMT3b 的表达，使 BDNF 启动子 IV 在 5 个 CpG 位点发生低甲基化，增强 BDNF 水平，从而降低脓毒症小鼠的逃避潜伏期，但增加平台穿越次数。我们的结果表明，2%H 吸入可能通过改变脓毒症小鼠海马中 DNMT1 和 DNMT3a 介导的 BDNF 启动子 IV 甲基化的调节来缓解 SAE。
