Data_Sheet_5_Knockdown of ATF3 suppresses the progression of ischemic stroke through inhibiting ferroptosis.ZIP

ObjectiveCurrent therapies towards ischemic stroke (IS) are still not satisfied, and alternative strategies targeting ferroptosis may be another choice. The purpose of this study is to screen potential ferroptosis-related genes involving in IS.
MethodsA rat model of IS was established via middle cerebral artery occlusion. Differentially expressed genes (DEGs) were screened from the model rats through transcriptional sequencing. Among the isolated DEGs, the expression of several attractive DEGs relating with ischemic injury was confirmed by qRT-PCR. Then, ATF3 relating with both IS and ferroptosis was selected a candidate gene for functional assays. After knockdown of ATF3 in the model rats, the infarction, histopathology, apoptosis, and ferroptosis in brain tissues were evaluated.
ResultsIS model was successfully established in rats, exhibiting the emergence of infarction area, histopathological injury, and enhanced cell apoptosis. Total 699 up-regulated DEGs and 461 down-regulated DEGs were screened from the model rats. qRT-PCR verified the up-regulation of Hspa1b, Tfpi2, Ptx3, and Atf3, and the down-regulation of Smyd1 and Tacr2 in the Model group compared with those in the Sham group. It is noteworthy that knockdown of ATF3 decreased the infarction area, relieved histopathological injury, weakened apoptosis, and inhibited ferroptosis in the model rats.
ConclusionSeveral candidate genes in relation with IS were revealed. More importantly, knockdown of ATF3 may relieve IS through inhibiting ferroptosis.

研究目的：当前缺血性脑卒中（ischemic stroke, IS）的治疗方案仍不尽人意，靶向铁死亡（ferroptosis）的替代治疗策略或为新的选择。本研究旨在筛选参与缺血性脑卒中的铁死亡相关潜在基因。
研究方法：通过大脑中动脉闭塞（middle cerebral artery occlusion, MCAO）法构建大鼠缺血性脑卒中模型。采用转录组测序技术从模型大鼠脑组织中筛选差异表达基因（Differentially Expressed Genes, DEGs）。从筛选得到的DEGs中，选取数条与缺血性损伤相关的候选DEGs，通过实时荧光定量聚合酶链反应（quantitative real-time polymerase chain reaction, qRT-PCR）验证其表达水平。随后，选取同时与缺血性脑卒中及铁死亡相关的激活转录因子3（Activating Transcription Factor 3, ATF3）作为功能实验的候选基因。在模型大鼠中敲低ATF3后，对脑组织的梗死情况、组织病理学改变、细胞凋亡及铁死亡水平进行评估。
研究结果：大鼠缺血性脑卒中模型构建成功，模型组大鼠出现脑梗死灶、组织病理学损伤及细胞凋亡水平升高。从模型大鼠脑组织中共筛选得到699个上调差异表达基因及461个下调差异表达基因。qRT-PCR验证结果显示，与假手术组相比，模型组大鼠脑组织中Hspa1b、Tfpi2、Ptx3及Atf3的表达显著上调，而Smyd1与Tacr2的表达显著下调。值得注意的是，敲低ATF3可缩小模型大鼠的脑梗死灶面积，减轻组织病理学损伤，抑制细胞凋亡及铁死亡过程。
研究结论：本研究筛选得到多个与缺血性脑卒中相关的候选基因。更为重要的是，敲低ATF3或可通过抑制铁死亡过程缓解缺血性脑卒中的病情。