miR-494/Nrdp1轴调控C/EBP-β泛素化在脑出血后巨噬细胞极化中的机制研究

Our previous studies showed that: (1) The metabolites of cerebral hemorrhage decreased the expression of ubiquitin E3 ligase Nrdp1 and promoted the M1 polarization of macrophages; (2) The miR-494 expression of macrophages induced by hematoma metabolites was upregulated. (3) Nrdp1 may be the target protein of miR-494; and Nrdp1 promoted the ubiquitination of C/EBP-β. Therefore, we speculate that the metabolites of cerebral hemorrhage may inhibit the ubiquitin E3 ligase Nrdp1 by blocking up the expression of miR-494, blocking the ubiquitination and activation of C/EBP-β, thereby promoting the M1 polarization of macrophages and aggravating inflammatory injury of nerve function. In this experiment, we used double luciferase, WB, FACS and laser confocal technology to control the expression of exogenous miR-494 and detect the level of C/EBP-β, macrophage M1/M2 polarization and neuroinflammation after cerebral hemorrhage. This study provide a new perspective to clarify ubiquitination in key regulatory mechanism of inflammatory injury of cerebral hemorrhage and explore the mode of action of the "miR-494-C/EBP-β ubiquitin-cell polarization ", and provide new theory and strategy for miRNA regulation of macrophage polarization in the treatment of cerebral hemorrhage .

我们前期研究表明：(1) 脑出血的代谢产物降低泛素E3连接酶Nrdp1的表达，并促进巨噬细胞M1极化；(2) 血肿代谢产物诱导巨噬细胞的miR-494表达显著上调。(3) Nrdp1可能是miR-494的靶蛋白；并且Nrdp1促进C/EBP-β的泛素化。因此，我们推测脑出血代谢产物可能通过上调miR-494的表达，抑制泛素E3连接酶Nrdp1，阻断C/EBP-β的泛素化活化，从而促进巨噬细胞的M1型极化，加重神经功能炎症损伤。本实验拟运用双荧光素酶、WB、FACS、激光共聚焦等技术，通过调控外源性miR-494的表达，检测脑出血后巨噬细胞C/EBP-β的水平，巨噬细胞M1/M2极化和神经炎症情况。本研究将从全新的角度阐明泛素化在脑出血炎症损伤的关键调控机制，深入探讨“miR-494-C/EBP-β泛素化-细胞极化”的作用模式，并为miRNA调控巨噬细胞极化在脑出血的治疗提供新理论和策略。

前期研究表明：(1) 脑出血的代谢产物降低泛素E3连接酶Nrdp1的表达，并促进巨噬细胞M1极化；(2) 血肿代谢产物诱导巨噬细胞的miR-494表达显著上调。(3) Nrdp1可能是miR-494的靶蛋白；并且Nrdp1促进C/EBP-β的泛素化。因此，我们推测脑出血代谢产物可能通过上调miR-494的表达，抑制泛素E3连接酶Nrdp1，阻断C/EBP-β的泛素化活化，从而促进巨噬细胞的M1型极化，加重神经功能炎症损伤。本实验运用双荧光素酶、WB、FACS、激光共聚焦等技术，通过调控外源性miR-494的表达，检测脑出血后巨噬细胞C/EBP-β的水平，巨噬细胞M1/M2极化和神经炎症情况。本研究将从全新的角度阐明泛素化在脑出血炎症损伤的关键调控机制，深入探讨“miR-494-C/EBP-β泛素化-细胞极化”的作用模式，并为miRNA调控巨噬细胞极化在脑出血的治疗提供新理论和策略。