PMID- 32032782 OWN - NLM STAT- MEDLINE DCOM- 20210427 LR - 20220531 IS - 1090-2139 (Electronic) IS - 0889-1591 (Print) IS - 0889-1591 (Linking) VI - 87 DP - 2020 Jul TI - N-methyl-d-aspartate receptor subunit 2B on keratinocyte mediates peripheral and central sensitization in chronic post-ischemic pain in male rats. PG - 579-590 LID - S0889-1591(19)31215-2 [pii] LID - 10.1016/j.bbi.2020.02.003 [doi] AB - The spinal N-methyl-d-aspartate (NMDA) receptor, and particularly its NR2B subunit, plays a pivotal role in neuropathic pain. However, the role of peripheral NMDA receptor in neuropathic pain is less well understood. We first treated cultured human keratinocytes, HaCaT cells with NMDA or NR2B-specific antagonist, ifenprodil and evaluated the level of total and phosphorylated NR2B at 24 h using Western blot. Next, using the chronic post-ischemia pain (CPIP) model, we administered NMDA or ifenprodil subcutaneously into the hind paws of male rats. Nociceptive behaviors were assessed by measuring mechanical and thermal withdrawal thresholds. Expression and phosphorylation of NR2B on keratinocyte were analyzed at 6, 12, 18, and 24 h on day 1 (initiation of pain) as well as day 2, 6, 10 and 14 (development and maintenance of pain) after the ischemia. The level of peripheral sensitization-related proteins (nuclear factor-kappaB (NF-kappaB), extracellular regulated protein kinases (ERK), and interleukin-1beta (IL-1beta)) in epidermis and dorsal root ganglion (DRG) were evaluated by immunofluorescence and western blot. Central sensitization-related C-fos induction, as well as astrocytes and microglia activation in the spinal cord dorsal horn (SDH) were studied using immunofluorescence. Administration of NMDA upregulated NR2B phosphorylation on HaCaT cells. CPIP-induced mechanical allodynia and thermal hyperalgesia were intensified by NMDA and alleviated by ifenprodil. CPIP resulted in an early upregulation of NR2B (peaked at 24 h) and late phosphorylation of NR2B (peaked at 14d) in hindpaw keratinocytes. CPIP led to an upregulation and phosphorylation of NF-kappaB and ERK, as well as an increased IL-1beta production in the ipsilateral skin and DRG. CPIP-associated c-fos induction in SDH persisted from acute to chronic stages after ischemia, while microglia and astrocyte activation were only observed in chronic phase. These CPIP-induced changes were also suppressed by ifenprodil administered subcutaneously in the hind paw. Our findings reveal a previously unrecognized role of keratinocyte NMDA receptor subunit 2B in peripheral and central nociceptive sensitization induced by CPIP. CI - Copyright (c) 2020 The Authors. Published by Elsevier Inc. All rights reserved. FAU - Xu, Xiaohan AU - Xu X AD - Department of Anesthesiology, Chinese Academy of Medical Sciences & Peking Union Medical College Hospital, Beijing 100730, China. FAU - Tao, Xin AU - Tao X AD - Department of Anesthesiology, Chinese Academy of Medical Sciences & Peking Union Medical College Hospital, Beijing 100730, China; Department of Infectious Disease, Southern Medical University Nanfang Hospital, Guangzhou, Guangdong 510515, China. FAU - Huang, Ping AU - Huang P AD - Department of Pain Management, Anesthesiology Institute, Cleveland Clinic, Cleveland, OH 44195, United States; Department of Inflammation and Immunity, Lerner Research Institute, Cleveland Clinic, Cleveland, OH 44195, United States. FAU - Lin, Feng AU - Lin F AD - Department of Inflammation and Immunity, Lerner Research Institute, Cleveland Clinic, Cleveland, OH 44195, United States. FAU - Liu, Qing AU - Liu Q AD - Department of Anesthesiology, Chinese Academy of Medical Sciences & Peking Union Medical College Hospital, Beijing 100730, China. FAU - Xu, Li AU - Xu L AD - Department of Anesthesiology, Chinese Academy of Medical Sciences & Peking Union Medical College Hospital, Beijing 100730, China. Electronic address: pumchxuli@163.com. FAU - Xu, Jijun AU - Xu J AD - Department of Pain Management, Anesthesiology Institute, Cleveland Clinic, Cleveland, OH 44195, United States; Department of Inflammation and Immunity, Lerner Research Institute, Cleveland Clinic, Cleveland, OH 44195, United States. Electronic address: xuj3@ccf.org. FAU - Huang, Yuguang AU - Huang Y AD - Department of Anesthesiology, Chinese Academy of Medical Sciences & Peking Union Medical College Hospital, Beijing 100730, China. LA - eng GR - K08 CA228039/CA/NCI NIH HHS/United States PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20200204 PL - Netherlands TA - Brain Behav Immun JT - Brain, behavior, and immunity JID - 8800478 RN - 0 (Receptors, N-Methyl-D-Aspartate) SB - IM EIN - Brain Behav Immun. 2020 Oct 5;:. PMID: 33032856 MH - Animals MH - *Central Nervous System Sensitization MH - Hyperalgesia MH - Ischemia MH - Keratinocytes/metabolism MH - Male MH - Pain Measurement MH - Phosphorylation MH - Rats MH - Rats, Sprague-Dawley MH - *Receptors, N-Methyl-D-Aspartate/metabolism MH - Spinal Cord/metabolism PMC - PMC8922412 MID - NIHMS1783823 OTO - NOTNLM OT - Animal model OT - Central sensitization OT - Chronic post-ischemia pain OT - Complex regional pain syndrome OT - Keratinocyte OT - NMDA receptor subunit 2B OT - Peripheral sensitization COIS- Declaration of Competing Interest None. EDAT- 2020/02/08 06:00 MHDA- 2021/04/28 06:00 PMCR- 2022/03/15 CRDT- 2020/02/08 06:00 PHST- 2019/09/20 00:00 [received] PHST- 2020/01/27 00:00 [revised] PHST- 2020/02/03 00:00 [accepted] PHST- 2020/02/08 06:00 [pubmed] PHST- 2021/04/28 06:00 [medline] PHST- 2020/02/08 06:00 [entrez] PHST- 2022/03/15 00:00 [pmc-release] AID - S0889-1591(19)31215-2 [pii] AID - 10.1016/j.bbi.2020.02.003 [doi] PST - ppublish SO - Brain Behav Immun. 2020 Jul;87:579-590. doi: 10.1016/j.bbi.2020.02.003. Epub 2020 Feb 4.