Ptbp1 Knockdown in Glial Cells Promotes Motor and Sensory Function Recovery After Peripheral Nerve Injury

ABSTRACT Background: Peripheral nerve injury (PNI) frequently causes persistent sensory and motor deficits with limited therapeutic options. While Ptbp1‐mediated astrocyte reprogramming shows promise in central nervous system repair, its role in PNI—particularly regarding spinal cord astrocytes and dorsal root ganglia (DRG) satellite glial cells (SGCs)—remains unexplored. Aims: This study aimed to determine whether Ptbp1 knockdown in glial cells enhances functional recovery after sciatic nerve injury (SNI) by dual mechanisms: (1) converting spinal cord astrocytes to motor neurons and polarizing them toward neuroprotective A2 phenotype, and (2) activating regenerative signaling pathways in DRG SGCs. Materials & Methods: C57BL/6J mice underwent SNI followed by intrathecal injection of AAV‐GFAP‐CasRx‐Ptbp1 (targeting Ptbp1 in astrocytes/SGCs) or control virus. Primary astrocytes and SGCs were transfected with Ptbp1 siRNA in vitro. Assessments included functional recovery (Basso Mouse Scale, Louisville Swim Score, Hargreaves test, von Frey assay), axonal regeneration (HE/β3‐tubulin/SCG‐10 staining), transcriptome/ATAC sequencing, and molecular analyses (immunofluorescence for DCX/Islet1/ntng2‐NGL‐2; Western blot for Ptbp1/GDNF/C3). Results: Ptbp1 was upregulated in spinal cord astrocytes and DRG SGCs post‐SNI. Its knockdown accelerated motor/sensory functional recovery and axonal regeneration. Mechanistically, in the spinal cord, Ptbp1 depletion induced astrocyte‐to‐motor neuron conversion (upregulation of DCX/Islet1/Map2) and polarized astrocytes toward A2 phenotype (upregulation of S100a10/GDNF; downregulation of C3). In DRG, it activated the ntng2/NGL‐2 pathway in SGCs, enhancing sensory axon regeneration (upregulation of ATF3/GAP43). Ntng2 blockade abolished sensory regeneration, confirming pathway dependence. Discussion: Ptbp1 knockdown promotes PNI repair through spatially distinct mechanisms: spinal cord astrocyte reprogramming/A2 polarization synergizes with DRG SGC‐mediated ntng2/NGL‐2 activation. While astrocyte‐to‐neuron conversion was limited, dominant A2 polarization provided neuroprotection. The absence of SGC transdifferentiation highlights cell‐type‐specific responses. Limitations include low conversion efficiency and interspecies regenerative differences. Conclusion: Targeting Ptbp1 in glial cells accelerates PNI recovery by dual regenerative mechanisms: motor function restoration via astrocyte‐derived neuron replenishment and A2 polarization, coupled with sensory repair through ntng2/NGL‐2 pathway activation. This establishes Ptbp1 as a promising therapeutic target for nerve injuries.