Mechanical stimulation simulating osteopathic pressing manipulation inhibits nociceptive hypersensitivity and synovial inflammation in knee osteoarthritis rats by modulating trafficking dynamics of transient receptor potentials via complexin2

Nociceptive hypersensitivity and synovial inflammation promote the progression of knee osteoarthritis (KOA) and joint pain, yet effective therapeutic strategies remain limited. This study investigated the mechanism of osteopathic pressing manipulation (OPM), a traditional Chinese spinal orthopedic technique, in alleviating KOA pain and inflammation. Using a rat KOA model, we demonstrated that moderate-intensity OPM (MOPM) significantly improved mechanical withdrawal threshold and reduced synovial inflammation markers IL-6, TNF-α and nociceptors TRPV1, TRPA1. RNA sequencing of dorsal root ganglion (DRG) tissues revealed that MOPM reversed KOA-induced upregulation of complexin 2 (CPLX2), a key modulator of SNARE complex-mediated cellular membrane trafficking. Mechanistically, MOPM suppressed CPLX2-dependent vesicle transport, thereby reducing membrane localization of TRP channels and secretion of pain mediators NGF, CGRP. In vitro co-culture experiments using DRG neurons and fibroblast-like synoviocytes confirmed that mechanical stress mimicking OPM attenuated trafficking dynamics-related protein expression VAMP2, SNAP25, and syntaxin1 via CPLX2 downregulation. Critically, CPLX2 siRNA synergized with MOPM to enhance anti-nociceptive and anti-inflammatory effects. These findings identify OPM as a non-pharmacological intervention capable of modulating CPLX2-mediated neuronal trafficking dynamics of TRPV1 and TRPA1, thereby alleviating pain and inflammation associated with KOA.