The dual inhibitors of receptor interacting protein kinase-1 and -3 (RIP1 and RIP3) play an important role in cell death processes and inflammatory responses. White matter injury (WMI), a leading cause of neurodevelopmental disabilities in preterm infants, which is characterized by extensive myelination disturbances and demyelination. Neuroinflammation, leads to the loss and differentiation-inhibition of oligodendrocyte precursor cells (OPCs), represents a major barrier to myelin repair. Whether the novel RIP1/RIP3 dual inhibitor ZJU-37 can promote transplanted OPCs derived from human neural stem cells (hOPCs) survival, differentiation and myelination remains unclear. In this study, we investigated the effect of ZJU-37 on myelination and neurobehavioral function in a neonatal rat WMI model induced by hypoxia and ischemia. In vivo, P3 rat pups were subjected to right common carotid artery ligation and hypoxia, and then treated with ZJU-37 or/and hOPCs, then OPCs apoptosis, myelination, glial cell and NLRP3 inflammasome activation together with cognitive outcome were evaluated at 12 weeks after transplantation. In vitro, the effect of ZJU-37 on NLRP3 inflammasome activation in astrocytes induced by oxygen–glucose deprivation (OGD) were examined by western blot and immunofluorescence. The effect of ZJU-37 on OPCs apoptosis induced by the conditioned medium from OGD-injured astrocytes (OGD-astrocyte-CM) was analyzed by flow cytometry and immunofluorescence. ZJU-37 combined with hOPCs more effectively decreased OPC apoptosis, promoted myelination in the corpus callosum and improved behavioral function compared to ZJU-37 or hOPCs treatment. In addition, the activation of glial cells and NLRP3 inflammasome was reduced by ZJU-37 or/and hOPCs treatment in the neonatal rat WMI model. In vitro, it was also confirmed that ZJU-37 can suppress NLRP3 inflammasome activation in astrocytes induced by OGD. Not only that, the OGD-astrocyte-CM treated with ZJU-37 obviously attenuated OPC apoptosis and dysdifferentiation caused by the OGD-astrocyte-CM. The novel RIP1/RIP3 dual inhibitor ZJU-37 may promote OPC survival, differentiation and myelination by inhibiting NLRP3 inflammasome activation in a neonatal rat model of WMI with hOPC graft.

中文翻译：

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新型 RIP1/RIP3 双抑制剂促进 hOPC 移植大鼠新生儿白质损伤模型的 OPC 存活和髓鞘形成

受体相互作用蛋白激酶-1 和 -3（RIP1 和 RIP3）的双重抑制剂在细胞死亡过程和炎症反应中发挥重要作用。白质损伤 (WMI)，早产儿神经发育障碍的主要原因，其特征是广泛的髓鞘形成障碍和脱髓鞘。神经炎症导致少突胶质细胞前体细胞 (OPC) 的丧失和分化抑制，是髓鞘修复的主要障碍。新型 RIP1/RIP3 双重抑制剂 ZJU-37 是否可以促进来自人类神经干细胞 (hOPCs) 的移植 OPCs 的存活、分化和髓鞘形成尚不清楚。在这项研究中，我们研究了 ZJU-37 对缺氧和缺血诱导的新生大鼠 WMI 模型中髓鞘形成和神经行为功能的影响。体内，P3幼鼠右颈总动脉结扎缺氧，然后用ZJU-37或/和hOPCs处理，然后在移植后12周评估OPCs凋亡、髓鞘形成、神经胶质细胞和NLRP3炎性体激活以及认知结果。在体外，通过蛋白质印迹和免疫荧光检查 ZJU-37 对氧-葡萄糖剥夺（OGD）诱导的星形胶质细胞中 NLRP3 炎性体激活的影响。通过流式细胞术和免疫荧光分析ZJU-37对OGD损伤星形胶质细胞（OGD-星形胶质细胞-CM）条件培养基诱导的OPCs凋亡的影响。与ZJU-37或hOPCs治疗相比，ZJU-37联合hOPCs更有效地减少OPC细胞凋亡，促进胼胝体髓鞘形成并改善行为功能。此外，在新生大鼠 WMI 模型中，ZJU-37 或/和 hOPCs 治疗降低了胶质细胞和 NLRP3 炎性体的活化。在体外，还证实ZJU-37可以抑制OGD诱导的星形胶质细胞中NLRP3炎性体的激活。不仅如此，经ZJU-37处理的OGD-星形胶质细胞-CM明显减弱了OGD-星形胶质细胞-CM引起的OPC细胞凋亡和分化异常。新型 RIP1/RIP3 双重抑制剂 ZJU-37 可能通过抑制 hOPC 移植 WMI 新生大鼠模型中 NLRP3 炎性体的激活来促进 OPC 存活、分化和髓鞘形成。ZJU-37处理的OGD-星形胶质细胞-CM明显减弱了OGD-星形胶质细胞-CM引起的OPC细胞凋亡和分化异常。新型 RIP1/RIP3 双重抑制剂 ZJU-37 可能通过抑制 hOPC 移植 WMI 新生大鼠模型中 NLRP3 炎性体的激活来促进 OPC 存活、分化和髓鞘形成。ZJU-37处理的OGD-星形胶质细胞-CM明显减弱了OGD-星形胶质细胞-CM引起的OPC细胞凋亡和分化异常。新型 RIP1/RIP3 双重抑制剂 ZJU-37 可能通过抑制 hOPC 移植 WMI 新生大鼠模型中 NLRP3 炎性体的激活来促进 OPC 存活、分化和髓鞘形成。