Hypersensitivity of myelinated A-fibers via toll-like receptor 5 promotes mechanical allodynia in tenascin-X-deficient mice associated with Ehlers-Danlos syndrome

Deficiency of an extracellular matrix glycoprotein tenascin-X (TNX) leads to a human heritable disorder Ehlers-Danlos syndrome, and TNX-deficient patients complain of chronic joint pain, myalgia, paresthesia, and axonal polyneuropathy. We previously reported that TNX-deficient (Tnxb^-/-) mice exhibit mechanical allodynia and hypersensitivity to myelinated A-fibers. Here, we investigated the pain response of Tnxb^-/- mice using pharmacological silencing of A-fibers with co-injection of N-(2,6-Dimethylphenylcarbamoylmethyl) triethylammonium bromide (QX-314), a membrane-impermeable lidocaine analog, plus flagellin, a Toll-like Receptor 5 (TLR5) ligand. Intraplantar co-injection of QX-314 and flagellin significantly increased the paw withdrawal threshold to transcutaneous sine wave stimuli at frequencies of 250 Hz (Aδ fiber responses) and 2000 Hz (Aβ fiber responses), but not 5 Hz (C fiber responses) in wild-type mice. The QX-314 plus flagellin-induced silencing of Aδ- and Aβ-fibers was also observed in Tnxb^-/- mice. Co-injection of QX-314 and flagellin significantly inhibited the mechanical allodynia and neuronal activation of the spinal dorsal horn in Tnxb^-/- mice. Interestingly, QX-314 alone inhibited the mechanical allodynia in Tnxb^-/- mice, and it increased the paw withdrawal threshold to stimuli at frequencies of 250 Hz and 2000 Hz in Tnxb^-/- mice, but not in wild-type mice. The inhibition of mechanical allodynia induced by QX-314 alone was blocked by intraplantar injection of a TLR5 antagonist TH1020 in Tnxb^-/- mice. These results suggest that mechanical allodynia due to TNX deficiency is caused by the hypersensitivity of Aδ- and Aβ-fibers, and it is induced by constitutive activation of TLR5.