Inhibiting ceramide synthase 5 expression in microglia decreases neuroinflammation after spinal cord injury

JOURNAL/nrgr/04.03/01300535-202510000-00026/figure1/v/2024-11-26T163120Z/r/image-tiff Microglia, the resident monocyte of the central nervous system, play a crucial role in the response to spinal cord injury. However, the precise mechanism remains unclear. To investigate the molecular mechanisms by which microglia regulate the neuroinflammatory response to spinal cord injury, we performed single-cell RNA sequencing dataset analysis, focusing on changes in microglial subpopulations. We found that the MG1 subpopulation emerged in the acute/subacute phase of spinal cord injury and expressed genes related to cell pyroptosis, sphingomyelin metabolism, and neuroinflammation at high levels. Subsequently, we established a mouse model of contusive injury and performed intrathecal injection of siRNA and molecular inhibitors to validate the role of ceramide synthase 5 in the neuroinflammatory responses and pyroptosis after spinal cord injury. Finally, we established a PC12-BV2 cell co-culture system and found that ceramide synthase 5 and pyroptosis-associated proteins were highly expressed to induce the apoptosis of neuron cells. Inhibiting ceramide synthase 5 expression in a mouse model of spinal cord injury effectively reduced pyroptosis. Furthermore, ceramide synthase 5-induced pyroptosis was dependent on activation of the NLRP3 signaling pathway. Inhibiting ceramide synthase 5 expression in microglia in vivo reduced neuronal apoptosis and promoted recovery of neurological function. Pla2g7 formed a “bridge” between sphingolipid metabolism and ceramide synthase 5-mediated cell death by inhibiting the NLRP3 signaling pathway. Collectively, these findings suggest that inhibiting ceramide synthase 5 expression in microglia after spinal cord injury effectively suppressed microglial pyroptosis mediated by NLRP3, thereby exerting neuroprotective effects.