Though the occurrence of neuroinflammation after spinal cord injury (SCI) is essential for antigen clearance and tissue repair, excessive inflammation results in cell death and axon dieback. The effect of anti‐inflammatory medicine used in clinical treatment remains debatable owing to the inappropriate therapeutic schedule that does not align with the biological process of immune reaction. A better understanding of the immunity process is critical to promote effective anti‐inflammatory therapeutics. However, cellular heterogeneity, which results in complex cellular functions, is a major challenge. This study performs single‐cell RNA sequencing by profiling the tissue proximity to the injury site at different time points after SCI. Depending on the analysis of single‐cell data and histochemistry observation, an appropriate time window for anti‐inflammatory medicine treatment is proposed. This work also verifies the mechanism of typical anti‐inflammatory medicine methylprednisolone sodium succinate (MPSS), which is found attributable to the activation inhibition of cells with pro‐inflammatory phenotype through the downregulation of pathways such as TNF, IL2, and MIF. These pathways can also be provided as targets for anti‐inflammatory treatment. Collectively, this work provides a therapeutic schedule of 1–3 dpi (days post injury) to argue against classical early pulse therapy and provides some pathways for target therapy in the future.

Neuroinflammation is a necessary response to spinal cord injury (SCI) as it helps to clear antigens and promote tissue repair. However, excessive inflammation can result in the death of cells and axon dieback. The efficacy of anti-inflammatory medication in clinical treatment is still up for debate due to inappropriate therapeutic scheduling that does not align with the biological process of immune response. A better understanding of the immune process is crucial for effective anti-inflammatory therapeutics, but it is challenged by cellular heterogeneity and complex cellular functions. To address this, we conducted a single-cell RNA sequencing study and profiled tissue proximity to the injury site at various time points after SCI. Based on our analysis of single-cell data and histochemistry observations, we recommend an appropriate time window of 1-3 days post-injury for anti-inflammatory medication treatment. We also verified the mechanism of MPSS, a typical anti-inflammatory medication, which was found to inhibit the activation of cells with pro-inflammatory phenotypes by downregulating pathways such as TNF, IL2, and MIF. These pathways could be potential targets for anti-inflammatory treatment. In summary, we recommend a therapeutic schedule of 1-3 days post-injury to argue against classical early pulse therapy and provide potential pathways for target therapy in the future.