Fan Xuehui, Chen Hongping, Xu Chen, Wang Yingju, Yin Pengqi, Li Meng, Tang Zhanbin, Jiang Fangchao, Wei Wan, Song Jihe, Li Guozhong, Zhong Di
Department of Neurology, The First Affiliated Hospital of Harbin Medical University, Harbin, China.
Front Pharmacol. 2022 May 24;13:834948. doi: 10.3389/fphar.2022.834948. eCollection 2022.
Ischemic stroke is the most common stroke incident. Sphingosine-1-phosphate (S1P) receptor 3 (S1PR3) is a member of the downstream G protein-coupled receptor family of S1P. The effect of S1PR3 on ischemic stroke remains elusive. We downloaded two middle cerebral artery occlusion (MCAO) microarray datasets from the Gene Expression Omnibus (GEO) database and screened differentially expressed genes (DEGs). Then, we performed a weighted gene coexpression network analysis (WGCNA) and identified the core module genes related to ischemic stroke. We constructed a protein-protein interaction (PPI) network for the core genes in which DEGs and WGCNA intersected. Finally, we discovered that S1PR3 was involved as the main member of the red proteome. Then, we explored the mechanism of S1PR3 in the mouse tMCAO model. The S1PR3-specific inhibitor CAY10444 was injected into the abdominal cavity of mice after cerebral ischemia/reperfusion (I/R) injury, and changes in the expression of blood-brain barrier-related molecules were measured using PCR, western blotting, and immunofluorescence staining. Both GEO datasets showed that S1PR3 was upregulated during cerebral I/R in mice. WGCNA revealed that the light yellow module had the strongest correlation with the occurrence of IS. We determined the overlap with DEGs, identified 146 core genes that are potentially related to IS, and constructed a PPI network. Finally, S1PR3 was found to be the main member of the red proteome. In the mouse cerebral I/R model, S1PR3 expression increased 24 h after ischemia. After the administration of CAY10444, brain edema and neurological deficits in mice were ameliorated. CAY10444 rescued the decreased expression of the tight junction (TJ) proteins zonula occludens 1 (ZO1) and occludin after ischemia induced by transient MCAO (tMCAO) and reduced the increase in aquaporin 4 (AQP4) levels after tMCAO, preserving the integrity of the BBB. Finally, we found that S1PR3 is involved in regulating the mitogen-activated protein kinase (MAPK) and (phosphatidylinositol-3 kinase/serine-threonine kinase) PI3K-Akt signaling pathways. S1PR3 participates in the regulation of blood-brain barrier damage after cerebral I/R. S1PR3 is expected to be an indicator and predictor of cerebral ischemia, and drugs targeting S1PR3 may also provide new ideas for clinical medications.
缺血性中风是最常见的中风事件。1-磷酸鞘氨醇（S1P）受体3（S1PR3）是S1P下游G蛋白偶联受体家族的成员。S1PR3对缺血性中风的影响尚不清楚。我们从基因表达综合数据库（GEO）下载了两个大脑中动脉闭塞（MCAO）微阵列数据集，并筛选了差异表达基因（DEG）。然后，我们进行了加权基因共表达网络分析（WGCNA），并确定了与缺血性中风相关的核心模块基因。我们为DEG和WGCNA相交的核心基因构建了蛋白质-蛋白质相互作用（PPI）网络。最后，我们发现S1PR3作为红色蛋白质组的主要成员参与其中。然后，我们在小鼠大脑中动脉闭塞（tMCAO）模型中探索了S1PR3的作用机制。在脑缺血/再灌注（I/R）损伤后，将S1PR3特异性抑制剂CAY10444注入小鼠腹腔，并使用聚合酶链反应（PCR）、蛋白质免疫印迹法和免疫荧光染色法检测血脑屏障相关分子表达的变化。两个GEO数据集均显示，小鼠脑I/R期间S1PR3表达上调。WGCNA显示，浅黄色模块与缺血性中风的发生相关性最强。我们确定了与DEG的重叠部分，鉴定了146个可能与缺血性中风相关的核心基因，并构建了PPI网络。最后，发现S1PR3是红色蛋白质组的主要成员。在小鼠脑I/R模型中，缺血24小时后S1PR3表达增加。给予CAY10444后，小鼠脑水肿和神经功能缺损得到改善。CAY10444挽救了短暂性大脑中动脉闭塞（tMCAO）诱导的缺血后紧密连接（TJ）蛋白闭合蛋白1（ZO1）和闭锁蛋白表达的降低，并降低了tMCAO后水通道蛋白4（AQP4）水平的升高，维持了血脑屏障的完整性。最后，我们发现S1PR3参与调节丝裂原活化蛋白激酶（MAPK）和磷脂酰肌醇-3激酶/丝氨酸-苏氨酸激酶（PI3K-Akt）信号通路。S1PR3参与脑I/R后血脑屏障损伤的调节。S1PR3有望成为脑缺血的一个指标和预测因子，靶向S1PR3的药物也可能为临床用药提供新思路。
