Abstract：
BACKGROUND: Rab3a regulates vesicle secretion and transport. Emerging evidences have shown that extracellular vesicles (EVs) can reach target lesions of injured spinal cords and exert a positive effect on these lesions. However, the molecular mechanism by which Rab3a regulates vesicle secretion to ameliorate spinal cord injury (SCI) is not fully understood.
METHODS: An SCI rat model was established which was used to examine the pathological changes and Rab3a expression in spinal cord tissue. Rab3a was overexpressed in the model rats to demonstrate its effect on SCI repair. Rab3a was also knocked down in neuronal cells to verify its role in vesicle secretion and neuronal cells. The binding protein of Rab3a was identified by Co-IP and mass spectrometry.
RESULTS: Rab3a was significantly downregulated in SCI rats and Rab3a overexpression promoted SCI repair. Rab3a knockdown inhibited the secretion of neuronal cell-derived EVs. Compared to the EVs from the equal number of control neuronal cells, EVs from Rab3a-knockdown neuronal cells promoted M1 macrophage polarization, which in turn, promoted neuronal cell apoptosis. Mechanistically, STXBP1 was identified as a binding protein of Rab3a, and their interaction promoted the secretion of neuronal cell-derived EVs. Furthermore, METTL2b was significantly downregulated in SCI rats, and METTL2b knockdown significantly reduced Rab3a protein expression.
CONCLUSIONS: These results suggest that Rab3a promotes the secretion of neuronal cell-derived EVs by interacting with its binding protein STXBP1. Neuronal cells-derived EVs inhibited the polarization of M1 macrophages in the spinal cord microenvironment, thereby promoting SCI repair. Our findings provide a theoretical basis for the clinical treatment of SCI.
METHODS: An SCI rat model was established which was used to examine the pathological changes and Rab3a expression in spinal cord tissue. Rab3a was overexpressed in the model rats to demonstrate its effect on SCI repair. Rab3a was also knocked down in neuronal cells to verify its role in vesicle secretion and neuronal cells. The binding protein of Rab3a was identified by Co-IP and mass spectrometry.
RESULTS: Rab3a was significantly downregulated in SCI rats and Rab3a overexpression promoted SCI repair. Rab3a knockdown inhibited the secretion of neuronal cell-derived EVs. Compared to the EVs from the equal number of control neuronal cells, EVs from Rab3a-knockdown neuronal cells promoted M1 macrophage polarization, which in turn, promoted neuronal cell apoptosis. Mechanistically, STXBP1 was identified as a binding protein of Rab3a, and their interaction promoted the secretion of neuronal cell-derived EVs. Furthermore, METTL2b was significantly downregulated in SCI rats, and METTL2b knockdown significantly reduced Rab3a protein expression.
CONCLUSIONS: These results suggest that Rab3a promotes the secretion of neuronal cell-derived EVs by interacting with its binding protein STXBP1. Neuronal cells-derived EVs inhibited the polarization of M1 macrophages in the spinal cord microenvironment, thereby promoting SCI repair. Our findings provide a theoretical basis for the clinical treatment of SCI.
摘要：
背景：Rab3a调节囊泡分泌和转运。研究表明，细胞外囊泡（EV）可以到达受损脊髓的目标病变，并对这些病变发挥积极作用。然而,Rab3a调节囊泡分泌以改善脊髓损伤（SCI）的分子机制尚不完全清楚。
方法：建立SCI大鼠模型，观察脊髓组织中Rab3a的表达及病理改变。Rab3a在模型大鼠中过表达以证明其对SCI修复的作用。Rab3a也在神经元细胞中被敲低，以验证其在囊泡分泌和神经元细胞中的作用。通过Co-IP和质谱鉴定Rab3a的结合蛋白。
结果：Rab3a在SCI大鼠中显著下调，Rab3a过表达促进SCI修复。Rab3a敲低抑制神经元细胞衍生的EV的分泌。来自Rab3a敲低神经元细胞的EV促进M1巨噬细胞极化，反过来，促进神经元细胞凋亡。机械上,STXBP1被鉴定为Rab3a的结合蛋白，它们的相互作用促进了神经元细胞衍生的电动汽车的分泌。此外，在SCI大鼠中METTL2显著下调，METTL2b敲除显著降低Rab3a蛋白表达。
结论：这些结果表明Rab3a通过与其结合蛋白STXBP1相互作用促进神经元细胞衍生的EV的分泌。EV抑制脊髓微环境中M1巨噬细胞的极化，从而促进SCI修复。本研究结果为SCI的临床治疗提供了理论依据。
方法：建立SCI大鼠模型，观察脊髓组织中Rab3a的表达及病理改变。Rab3a在模型大鼠中过表达以证明其对SCI修复的作用。Rab3a也在神经元细胞中被敲低，以验证其在囊泡分泌和神经元细胞中的作用。通过Co-IP和质谱鉴定Rab3a的结合蛋白。
结果：Rab3a在SCI大鼠中显著下调，Rab3a过表达促进SCI修复。Rab3a敲低抑制神经元细胞衍生的EV的分泌。来自Rab3a敲低神经元细胞的EV促进M1巨噬细胞极化，反过来，促进神经元细胞凋亡。机械上,STXBP1被鉴定为Rab3a的结合蛋白，它们的相互作用促进了神经元细胞衍生的电动汽车的分泌。此外，在SCI大鼠中METTL2显著下调，METTL2b敲除显著降低Rab3a蛋白表达。
结论：这些结果表明Rab3a通过与其结合蛋白STXBP1相互作用促进神经元细胞衍生的EV的分泌。EV抑制脊髓微环境中M1巨噬细胞的极化，从而促进SCI修复。本研究结果为SCI的临床治疗提供了理论依据。
