组蛋白甲基转移酶酶增强子的 zeste 同源物 2 通过 H3K27me3 介导的 PI3K/AKT/mTOR 信号通路调节拮抗缺血性脑损伤。

Histone methyltransferase enzyme enhancer of zeste homolog 2 counteracts ischemic brain injury via H3K27me3-mediated regulation of PI3K/AKT/mTOR signaling pathway.

机构信息

Department of Emergency, The First Affiliated Hospital of Hainan Medical University, Haikou, People's Republic of China.

The Institute for Biomedical Engineering and Nano Science, Tongji University School of Medicine, Shanghai, People's Republic of China.

出版信息

Environ Toxicol. 2023 Sep;38(9):2240-2255. doi: 10.1002/tox.23863. Epub 2023 Jun 19.

DOI:10.1002/tox.23863

PMID:37334851

Abstract

BACKGROUND

Epigenetic histone methylation plays a crucial role in cerebral ischemic injury, particularly in the context of ischemic stroke. However, the complete understanding of regulators involved in histone methylation, such as Enhancer of Zeste Homolog 2 (EZH2), along with their functional effects and underlying mechanisms, remains incomplete.

METHODS

Here, we employed a rat model of MCAO (Middle cerebral artery occlusion) and an OGD (Oxygen-Glucose Deprivation) model of primary cortical neurons to study the role of EZH2 and H3K27me3 in cerebral ischemia-reperfusion injury. The infarct volume was measured through TTC staining, while cell apoptosis was detected using TUNEL staining. The mRNA expression levels were quantified through quantitative real-time polymerase chain reaction (qPCR), whereas protein expressions were evaluated via western blotting and immunofluorescence experiments.

RESULTS

The expression levels of EZH2 and H3K27me3 were upregulated in OGD; these expression levels were further enhanced by GSK-J4 but reduced by EPZ-6438 and AKT inhibitor (LY294002) under OGD conditions. Similar trends were observed for mTOR, AKT, and PI3K while contrasting results were noted for UTX and JMJD3. The phosphorylation levels of mTOR, AKT, and PI3K were activated by OGD, further stimulated by GSK-J4, but inhibited by EPZ-6438 and AKT inhibitor. Inhibition of EZH2 or AKT effectively counteracted OGD-/MCAO-induced cell apoptosis. Additionally, inhibition of EZH2 or AKT mitigated MCAO-induced infarct size and neurological deficit in vivo.

CONCLUSIONS

Collectively, our results demonstrate that EZH2 inhibition exerts a protective effect against ischemic brain injury by modulating the H3K27me3/PI3K/AKT/mTOR signaling pathway. The results provide novel insights into potential therapeutic mechanisms for stroke treatment.

摘要

背景

表观遗传组蛋白甲基化在脑缺血损伤中起着至关重要的作用，尤其是在缺血性中风的情况下。然而，对于涉及组蛋白甲基化的调节因子（如 Enhancer of Zeste Homolog 2，EZH2）及其功能影响和潜在机制，我们的理解还不完全。

方法

在这里，我们使用 MCAO（大脑中动脉闭塞）大鼠模型和原代皮质神经元的 OGD（氧葡萄糖剥夺）模型来研究 EZH2 和 H3K27me3 在脑缺血再灌注损伤中的作用。通过 TTC 染色测量梗死体积，通过 TUNEL 染色检测细胞凋亡。通过实时定量聚合酶链反应（qPCR）定量测量 mRNA 表达水平，通过 Western blot 和免疫荧光实验评估蛋白表达。

结果

EZH2 和 H3K27me3 的表达水平在 OGD 中上调；在 OGD 条件下，这些表达水平进一步被 GSK-J4 增强，但被 EPZ-6438 和 AKT 抑制剂（LY294002）降低。mTOR、AKT 和 PI3K 表现出相似的趋势，而 UTX 和 JMJD3 则表现出相反的趋势。mTOR、AKT 和 PI3K 的磷酸化水平被 OGD 激活，进一步被 GSK-J4 刺激，但被 EPZ-6438 和 AKT 抑制剂抑制。EZH2 或 AKT 的抑制有效地抵消了 OGD-/MCAO 诱导的细胞凋亡。此外，EZH2 或 AKT 的抑制减轻了体内 MCAO 诱导的梗死面积和神经功能缺损。