Anti-arthritic Effect of the Spirocyclopiperazinium Salt Compound LXM-15 in Rats and Its Underlying Mechanism

In this study, we aimed to evaluate the effects of the spirocyclopiperazinium salt compound LXM-15 on rheumatoid arthritis induced by complete Freund’s adjuvant (CFA) in rats and investigate the underlying mechanism. The results showed that LXM-15 significantly inhibited the paw edema and ankle swelling, and alleviated the mechanical allodynia and thermal hyperalgesia responses in the CFA rats. The histopathological results revealed that LXM-15 ameliorated the infiltration of inflammatory cells and joint destruction. The micro-CT scan showed that LXM-15 alleviated bone erosion and increased BMD in the ankle joints of the CFA rats. Western blot analyses showed that LXM-15 significantly reduced the upregulation of phospho-JAK2, phospho-STAT3, phospho-IκBα, and phospho-NF-κBp65, and the overexpression of BDNF in the dorsal root ganglions. ELISA result showed that the protein level of TNF-α in the paw tissue was decreased upon LXM-15 treatment. RT-PCR analysis showed that the mRNA expression levels of c-fos and BDNF were reduced in the dorsal root ganglions by LXM-15 treatment. The LXM-15-mediated anti-arthritic effects were abolished by treatment with hexamethonium (a peripheral nicotinic receptor antagonist), atropine methylnitrate (a peripheral muscarinic receptor antagonist), methyllycaconitine citrate (a selective α7 nicotinic receptor antagonist), and tropicamide (a selective M4 muscarinic receptor antagonist). Collectively, our results demonstrate that LXM-15 exerts anti-arthritic effects in CFA rats. The underlying mechanism may be related to the activation of the peripheral α7 nicotinic receptor and M4 muscarinic receptor by LXM-15, further suppressing the activation of the JAK2/STAT3 and IκBα/NF-κBp65 signaling pathways and, eventually, inhibiting the expression levels of TNF-α, BDNF, and c-fos.