title: TRAF6 upregulation in spinal astrocytes maintains neuropathic pain by integrating TNF-α and IL-1β signaling.
abstract: The proinflammatory cytokines tumor necrosis factor (TNF) α and interleukin (IL) 1β have been strongly implicated in the pathogenesis of neuropathic pain, but the intracellular signaling of these cytokines in glial cells is not fully understood. TNF receptor-associated factor 6 (TRAF6) plays a key role in signal transduction in the TNF receptor superfamily and the IL-1 receptor superfamily. In this study, we investigated the role of TRAF6 in neuropathic pain in mice after spinal nerve ligation (SNL). SNL induced persistent TRAF6 upregulation in the spinal cord. Interestingly, TRAF6 was mainly colocalized with the astrocytic marker glial fibrillary acidic protein on SNL day 10 and partially expressed in microglia on SNL day 3. In cultured astrocytes, TRAF6 was upregulated after exposure to TNF-α or IL-1β. TNF-α or IL-1β also increased CCL2 expression, which was suppressed by both siRNA and shRNA targeting TRAF6. TRAF6 siRNA treatment also inhibited the phosphorylation of c-Jun N-terminal kinase (JNK) in astrocytes induced by TNF-α or IL-1β. JNK inhibitor D-NKI-1 dose-dependently decreased IL-1β-induced CCL2 expression. Moreover, spinal injection of TRAF6 siRNA decreased intrathecal TNF-α- or IL-1β-induced allodynia and hyperalgesia. Spinal TRAF6 inhibition via TRAF6 siRNA, shRNA lentivirus, or antisense oligodeoxynucleotides partially reversed SNL-induced neuropathic pain and spinal CCL2 expression. Finally, intrathecal injection of TNF-α-activated astrocytes induced mechanical allodynia, which was attenuated by pretreatment of astrocytes with TRAF6 siRNA. Taken together, the results suggest that TRAF6, upregulated in spinal cord astrocytes in the late phase after nerve injury, maintains neuropathic pain by integrating TNF-α and IL-1β signaling and activating the JNK/CCL2 pathway in astrocytes.
authors.affiliation: Pain Research Laboratory, Institute of Nautical Medicine, Jiangsu Key Laboratory of Inflammation and Molecular Drug Target, Nantong University, Nantong, Jiangsu, China Department of Nutrition, School of Public Health, Nantong University, Nantong, Jiangsu, China Departments of Anesthesiology and Neurobiology, Duke University Medical Center, Durham, NC 27710, USA.
authors.forename: Ying
authors.initials: Y
authors.lastname: Lu
authors.forename: Bao-Chun
authors.initials: BC
authors.lastname: Jiang
authors.forename: De-Li
authors.initials: DL
authors.lastname: Cao
authors.forename: Zhi-Jun
authors.initials: ZJ
authors.lastname: Zhang
authors.forename: Xin
authors.initials: X
authors.lastname: Zhang
authors.forename: Ru-Rong
authors.initials: RR
authors.lastname: Ji
authors.forename: Yong-Jing
authors.initials: YJ
authors.lastname: Gao
mesh_terms: D000818:Animals; D001253:Astrocytes / Q000187:drug effects / Q000378:metabolism*; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology; D018932:Chemokine CCL2 / Q000378:metabolism; D004195:Disease Models, Animal; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D053583:Interleukin-1beta / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D008297:Male; D051379:Mice; D008813:Mice, Inbred ICR; D009437:Neuralgia / Q000473:pathology*; D010147:Pain Measurement; D012333:RNA, Messenger / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D013116:Spinal Cord / Q000473:pathology; D048029:TNF Receptor-Associated Factor 6 / Q000235:genetics / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D015854:Up-Regulation / Q000187:drug effects / Q000502:physiology*
chemical_list: D018932:Chemokine CCL2; D005904:Glial Fibrillary Acidic Protein; D053583:Interleukin-1beta; D012333:RNA, Messenger; D048029:TNF Receptor-Associated Factor 6; D014409:Tumor Necrosis Factor-alpha
keywords: Astrocytes; CCL2; IL-1beta; JNK; Neuropathic pain; Proinflammatory cytokines; TNF-alpha; TRAF6