Electroacupuncture Attenuates Inflammatory Pain by Activating AMPK/autophagy and Inhibiting iNOS and IL-1β in Inflamed Tissues

Abstract

BackgroundElectroacupuncture (EA) produces analgesic effects on inflammatory pain partially via activating adenosine monophosphate (AMP)-activated protein kinase (AMPK) pathway in the spinal cord. However, it is unclear whether EA activates AMPK in peripheral tissues in inflammatory pain. This study was aimed at determining whether EA promotes autophagy by activating AMPK to inhibit the expression of inflammatory mediators IL-1β and iNOS in inflamed skin tissues. MethodsIn CFA-induced inflammatory pain in mice, mechanical allodynia and thermal hyperalgesia were tested 2 hours after EA treatment. The AMPK antagonist Compound C was injected intraperitoneally 30 minutes before EA treatment. The analgesic effects of AMPK activator 5-aminoimidazole-4-carboxyamide ribonucleoside (AICAR) were determined and its effects on autophagy, IL-1β and iNOS expression were detected. Also, the effects of the autophagy inhibitor 3-methyladenine (3-MA) on EA analgesia and iNOS/IL-1β expression in inflamed skin tissues were examined. The phosphorylation of AMPK (Thr172) and total AMPK proteins, LC3BII/I, autophagy substrate protein p62, IL-1β and iNOS were detected using Western blotting. Co-labeling of macrophages (CD68) with IL-1β and iNOS was detected using immunofluorescence. In addition, after NR8383 macrophages were treated with CFA, the effects of AICAR and Compound C on autophagy were determined using stubRFP-sensGFP-LC3 Lentivirus..ResultsEA reduced CFA-induced inflammatory pain, activated AMPK and autophagy, and inhibited iNOS and IL-1β expression in inflamed skin tissues. AICAR also attenuated CFA-induced hyperalgesia, promoted autophagy and inhibited iNOS and IL-1β expression in vivo and in vitro. In addition, the AMPK inhibitor Compound C reversed the effect of EA on autophagy. Pretreatment with 3-MA, an inhibitor of autophagy, inhibited the effect of EA on inflammatory pain and expression of iNOS and IL-1β in inflamed skin tissues. ConclusionsEA treatment alleviated inflammatory pain by activation of AMPK, enhancing autophagy, and inhibiting iNOS and IL-1β expression in the inflamed skin tissues.