Contents lists available at ScienceDirect Brain, Behavior, and Immunity journal homepage: www.elsevier.com/locate/ybrbi Full-length Article Spinal interleukin-10 produces antinociception in neuropathy through microglial β-endorphin expression, separated from antineuroinflammation Hai-Yun Wu, Xiao-Fang Mao, Xue-Qi Tang, Usman Ali, Evhy Apryani, Hao Liu, Xin-Yan Li, Yong-Xiang Wang ⁎ King’s Lab, Shanghai Jiao Tong University School of Pharmacy, Shanghai 200240, China ARTICLE INFO Keywords: Interleukin 10 (IL-10) Microglia β-Endorphin Antinociception Antineuroinflammation Neuroinflammatory cytokines ABSTRACT Interleukin 10 (IL-10) is antinociceptive in various animal models of pain without induction of tolerance, and its mechanism of action was generally believed to be mediated by inhibition of neuroinflammation. Here we re- ported that intrathecal IL-10 injection dose dependently attenuated mechanical allodynia and thermal hyper- algesiain male and female neuropathic rats, with ED 50 values of 40.8 ng and 24 ng, and E max values of 61.5% MPE and 100% MPE in male rats. Treatment with IL-10 specifically increased expression of the β-endorphin (but not prodynorphin) gene and protein in primary cultures of spinal microglia but not in astrocytes or neurons. Intrathecal injection of IL-10 stimulated β-endorphin expression from microglia but not neurons or astrocytes in both contralateral and ipsilateral spinal cords of neuropathic rats. However, intrathecal injection of the β-en- dorphin neutralizing antibody, opioid receptor antagonist naloxone, or μ-opioid receptor antagonist CTAP completely blocked spinal IL-10-induced mechanical antiallodynia, while the microglial inhibitor minocycline and specific microglia depletor reversed spinal IL-10-induced β-endorphin overexpression and mechanical an- tiallodynia. IL-10 treatment increased spinal microglial STAT3 phosphorylation, and the STAT3 inhibitor NSC74859 completely reversed IL-10-increased spinal expression of β-endorphin and neuroinflammatory cy- tokines and mechanical antiallodynia. Silence of the Bcl3 and Socs3 genes nearly fully reversed IL-10-induced suppression of neuroinflammatory cytokines (but not expression of β-endorphin), although it had no effect on mechanical allodynia. In contrast, disruption of the POMC gene completely blocked IL-10-stimulated β-en- dorphin expression and mechanical antiallodynia, but had no effect on IL-10 inhibited expression of neuroin- flammatory cytokines. Thus this study revealed that IL-10 produced antinociception through spinal microglial β- endorphin expression, but not inhibition of neuroinflammation. 1. Introduction IL-10, one of the most important antiinflammatory and im- munosuppressive cytokines, is involved in infectious and autoimmune diseases, like ulcerative colitis, Crohn’s disease, and multiple sclerosis (Fedorak et al., 2000; Fillatreau et al., 2002). In the central nervous system, IL-10 exhibits neuroprotection and antinociception in various rodent models (Ooboshi et al., 2005; Scholz and Woolf, 2007; Johnston et al., 2008). For example, intrathecal delivery of recombinant IL-10 blocked neuropathic pain induced by chronic constriction injury (Wagner et al., 1998), spared nerve injury (Wang et al., 2012b), or cancer chemotherapy with paclitaxel (Ledeboer et al., 2007). IL-10 was also effective in attenuating inflammatory pain induced by snake venom phospholipase A2 (Laughlin et al., 2000), chronic orofacial in- flammation (Shimizu et al., 2009), acetic acid and zymosan (Vale et al., 2003). Interestingly, sustained IL-10 treatment did not induce anti- nociceptive tolerance in chronic pain states (Wagner et al., 1998; Milligan et al., 2006b; Dengler et al., 2014). Thus, to produce durable antinociception in chronic pain, IL-10 gene therapy using both in- trathecal viral and non-viral delivery systems has been successfully employed in treating chronic pain in rodent models (Milligan et al., 2005a,b, 2006a,b; Soderquist et al., 2010). IL-10 could be a potential strategy to treat clinical pain, particularly chronic neuropathic pain, an unmet medical need. Accumulated evidence indicated that spinal microglia played a crucial role in neuropathic pain, particularly at the initial stage through https://doi.org/10.1016/j.bbi.2018.06.015 Received 2 March 2018; Received in revised form 30 May 2018; Accepted 15 June 2018 ⁎ Corresponding author at: King’s Lab, Shanghai Jiao Tong University School of Pharmacy, 800 Dongchuan Road, Shanghai 200240, China. E-mail address: yxwang@sjtu.edu.cn (Y.-X. Wang). Abbreviations: GLP-1, glucagon-like peptide-1; IL-10, interleukin 10; TNF-α, tumor necrosis factor-α; IL-1β, interleukin-1β; IL-6, interleukin 6; p38 MAPK, p38 mitogen-activated protein kinase; NF-κB, nuclear factor κB; STAT3, signal transducer and activator of transcription 3; Bcl3, B-cell lymphoma 3; Bcl2, B-cell lymphoma 2; Socs3, suppressor of cytokine signaling 3; DUSP1, dual-specificity phosphatase-1; LPS, lipopolysaccrade; NSAIDs, nonsteroidal antiinflammatory drugs; IL-1ra, IL-1 receptor antagonist Brain, Behavior, and Immunity xxx (xxxx) xxx–xxx 0889-1591/ © 2018 Elsevier Inc. All rights reserved. Please cite this article as: Wu, H.-Y., Brain, Behavior, and Immunity (2018), https://doi.org/10.1016/j.bbi.2018.06.015