ACVR1 drives neuropathic pain by regulating NLRP3-Induced neuronal pyroptosis through the p38 and Smad1/5/8 pathways

Background: Neuropathic pain is characterized by sustained pain hypersensitivity caused by nerve injury. The molecular mechanisms underlying this condition remain poorly understood. This study aims to elucidate the role of ACVR1 and its downstream pathways in mediating neuropathic pain through neuronal Pyroptosis and neuroinflammation.

Methods: A spared nerve injury (SNI) model was established both in male and female mouse to induce neuropathic pain. Behavioral tests, Western blot, PCR, and immunofluorescence were used to assess the expression of ACVR1, p-Smad1/5/8, p-p38, and pyroptosis-related proteins (NLRP3, Caspase-1, and GSDMD-N). ACVR1, p38, and Smad1/5/8 were pharmacologically inhibited to evaluate their roles in neuropathic pain and Pyroptosis.

Results: Behavioral analysis confirmed successful SNI model establishment, marked by reduced paw withdrawal thresholds (PWT). Protein and mRNA expression analysis revealed significant upregulation of ACVR1, p-Smad1/5/8, and p-p38 in the spinal cord, particularly in neurons. Furthermore, SNI enhanced pyroptosis-related protein expression, including NLRP3, Caspase-1, GSDMD-N, IL-1β and IL-18. Inhibition of ACVR1 alleviated mechanical allodynia, reduced neuronal Pyroptosis, and decreased serum IL-1β and IL-18 levels. Similarly, p38 inhibition mitigated NLRP3-induced Pyroptosis without altering ACVR1 expression. In contrast, Smad1/5/8 inhibition by DMH-1 effectively reduced Pyroptosis and inflammation via NLRP3 but had no effect on p38 phosphorylation. Combined p38 and Smad1/5/8 pathway inhibition synergistically decreased pyroptosis-related protein expression, highlighting their interactive roles in ACVR1-mediated neuropathic pain.

Conclusion: These findings suggest that ACVR1 exacerbates neuropathic pain by activating neuronal Pyroptosis and neuroinflammation via the p38 and Smad1/5/8 pathways. Targeting ACVR1 and its downstream signaling pathways may offer novel therapeutic strategies for neuropathic pain.

ACVR1; NLRP3 inflammasome; Neuronal pyroptosis; Neuropathic pain; Smad1/5/8 pathway; p38 MAPK.