RT Journal Article
SR Electronic
T1 Sensitization of knee-innervating sensory neurons by tumor necrosis factor-α activated fibroblast-like synoviocytes: an in vitro, co-culture model of inflammatory pain
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 791251
DO 10.1101/791251
A1 Sampurna Chakrabarti
A1 Luke A. Pattison
A1 Charity N. Bhebhe
A1 Gerard Callejo
A1 David C. Bulmer
A1 Ewan St. John Smith
YR 2019
UL http://biorxiv.org/content/early/2019/10/07/791251.abstract
AB Background Pain is a principle contributor to the global burden of arthritis with peripheral sensitization being a major cause of arthritis-related pain. Within the knee joint, distal endings of dorsal root ganglion neurons (knee neurons) interact with fibroblast-like synoviocytes (FLS) and the inflammatory mediators they secrete, which are thought to promote peripheral sensitization. Therefore, we investigated the communication between knee neurons and FLS in a co-culture system.Methods Inflammation in FLS (isolated from mouse patella) was induced by stimulation with tumor necrosis factor-α (TNF-FLS). Expression of relevant genes, secretion of cytokines and functional acid response was assessed using polymerase chain reactions, inflammatory antibody array blots and Ca2+ imaging respectively. Electrophysiology was performed on dissociated knee neurons in mono-culture, co-culture with control FLS or TNF-FLS, or supernatant derived from TNF-FLS cultures to determine electrical excitability and sensitivity to transient receptor potential (TRP) agonists. Two group comparisons were conducted using Student’s t-tests (distributed variable) or chi-sq tests (categorical variable). ANOVA followed by Tukey’s post-hoc test was performed for comparing more than two groups.Results Compared to control, TNF-FLS showed increased expression of interleukin-6 (Il-6, p < 0.01, unpaired t-test) and Il-1r1 (p < 0.05, unpaired t-test), and enhanced cytokine secretion. FLS also responded to acidic stimuli and the percentage of TNF-FLS acid responders increased in the pH range 6.0-5.0 (p < 0.001, chi-sq test). Co-culture of DRG neurons with TNF-FLS or supernatant derived from TNF-FLS depolarized the resting membrane potential (p < 0.01, ANOVA), increased spontaneous action potential firing (p < 0.05, ANOVA) and enhanced TRPV1 function (p < 0.05, ANOVA) of knee neurons consistent with a role for FLS in the sensitization of pain sensing nerves in arthritis.Conclusions TNF-FLS have a pro-nociceptive phenotype, displaying enhanced acid sensitivity and increased secretion of pro-inflammatory cytokines. Co-culture with TNF-FLS or supernatant from TNF-FLS, induces hyperexcitability of knee neurons and enhances TRPV1 function. Data from this study demonstrate the ability of FLS activated by TNF-α to promote neuronal sensitization, results that highlight the importance of both non-neuronal and neuronal cells to the development of pain in arthritis.