Stroke serves as a prevalent cerebrovascular disorder with severe cerebral ischemia/reperfusion (CIR) injury, in which neural stem cells (NSCs) play critical roles in the recovery of cerebral function. Circular RNAs (circRNAs) have been widely found to participate in stroke and NSC modulation. However, the role of circRNA TTC3 (circTTC3) in the regulation of CIR injury and NSCs remains elusive. Here, we aimed to explore the impact of circTTC3 on CIR injury and NSCs. The middle cerebral artery occlusion/repression (MCAO/R) model was established in C57BL/6J mice. The primary astrocytes were isolated from the cerebellum from C57BL/6J mice. The primary NSCs were obtained from rat embryos. The effect of circTTC3 on CIR injury and NSCs was analyzed by TTC staining, qPCR, Western blot, LDH colorimetric kits, MTT assays, Annexin V-FITC Apoptosis Detection Kit, luciferase reporter gene assays, and others in the system. Significantly, the expression of circTTC3 was elevated in the MCAO/R mice and oxygen and glucose deprivation (OGD)-treated astrocytes. The depletion of circTTC3 attenuated cerebral infarction, neurological score, and brain water content. The OGD treatment induced apoptosis and the levels of lactate dehydrogenase (LDH) in the astrocytes, in which circTTC3 depletion reduced this phenotype in the system. Moreover, the depletion of circTTC3 promoted the proliferation and upregulated the nestin and β-tubulin III expression in NSCs. Mechanically, circTTC3 was able to sponge miR-372-3p, and miR-372-3p can target Toll-like receptor 4 (TLR4) in NSCs. The miR-372-3p inhibitor or TLR4 overexpression could reverse circTTC3 depletion-mediated astrocyte OGD injury and NSC regulation. Thus, we conclude that circTTC3 regulates CIR injury and NSCs by the miR-372-3p/TLR4 axis in cerebral infarction. Our finding presents new insight into the mechanism by which circTTC3 modulates CIR injury and NSC dysfunction. CircTTC3, miR-372-3p, and TLR4 may serve as potential targets for the treatment of CIR injury during stroke.

中文翻译：

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环形RNA TTC3通过miR-372-3p / TLR4轴调节脑梗死中的脑缺血-再灌注损伤和神经干细胞

中风是一种严重的脑缺血/再灌注（CIR）损伤的普遍性脑血管疾病，其中神经干细胞（NSC）在脑功能恢复中起关键作用。广泛发现环状RNA（circRNA）参与中风和NSC调节。但是，circRNA TTC3（circTTC3）在CIR损伤和NSC调控中的作用仍然难以捉摸。在这里，我们旨在探讨circTTC3对CIR损伤和NSC的影响。在C57BL / 6J小鼠中建立了大脑中动脉闭塞/抑制（MCAO / R）模型。从C57BL / 6J小鼠小脑中分离出原代星形胶质细胞。主要的NSC来自大鼠胚胎。通过TTC染色，qPCR，Western印迹，LDH比色试剂盒，MTT分析，分析了circTTC3对CIR损伤和NSC的作用。系统中的膜联蛋白V-FITC细胞凋亡检测试剂盒，荧光素酶报告基因检测及其他。值得注意的是，在MCAO / R小鼠和氧和葡萄糖剥夺（OGD）处理的星形胶质细胞中，circTTC3的表达升高。circTTC3的耗竭减弱了脑梗塞，神经学评分和脑含水量。OGD处理诱导星形胶质细胞凋亡和乳酸脱氢酶（LDH）的水平，其中circTTC3耗竭减少了该表型。此外，circTTC3的消耗促进了NSCs的增殖并上调了Nestin和β-tubulinIII的表达。从机械上讲，circTTC3能够海绵化miR-372-3p，而miR-372-3p可以靶向NSC中的Toll样受体4（TLR4）。miR-372-3p抑制剂或TLR4过表达可以逆转circTTC3耗竭介导的星形胶质细胞OGD损伤和NSC调节。因此，我们得出结论，circTTC3通过miR-372-3p / TLR4轴在脑梗死中调节CIR损伤和NSC。我们的发现为circTTC3调节CIR损伤和NSC功能障碍提供了新的见解。CircTTC3，miR-372-3p和TLR4可能作为中风期间CIR损伤治疗的潜在靶标。