Role of spinal Nrf2/HO-1 signaling pathway in hydrogen-induced reduction of inflammatory pain in rats

Objective To evaluate the role of spinal nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1) signaling pathway in hydrogen-induced reduction of inflammatory pain in rats. Methods Sixty-four SPF healthy adult male Sprague-Dawley rats, weighing 200-250 g, were divided into 4 groups (n=16 each) using a random number table: control group (group C), inflammatory pain group (group IP), inflammatory pain plus hydrogen-rich saline group (group IP+ H2) and inflammatory pain plus hydrogen-rich saline plus Nrf2 inhibitor all-trans retinoic acid (ATRA) group (group IP+ H2+ ATRA). Chronic inflammatory pain was induced by injecting complete Freund′s adjuvant (CFA) 100 μl into the plantar surface of the left hind paw in IP group and IP+ H2 group.The equal volume of normal saline was given instead in group C. Hydrogen-rich saline 5 ml/kg was injected intraperitoneally once a day for 7 consecutive days starting from 1 day after injecting CFA in group IP+ H2 and group IP+ H2+ ATRA, and the equal volume of normal saline was given instead in the other groups.ATRA 7 mg/kg was injected intraperitoneally once a day for 2 consecutive days starting from 2 days before injecting CFA.The mechanical paw withdrawal threshold (MWT) and thermal paw withdrawal latency (TWL) were measured at 1 day before establishing the model (T0) and 1, 3 and 7 days after establishing the model (T1-3). Six rats were sacrificed after the last measurement of pain threshold on day 7 after establishing the model, and the L4-6 lumbar segments of the spinal cord were removed for determination of the expression of Nrf2, HO-1 and glial fibrillary acidic protein (GFAP) by Western blot. Results Compared with group C, the MWT was significantly decreased and the TWL was shortened at T1-3, and the expression of Nrf2, HO-1 and GFAP was up-regulated in IP and IP+ H2 groups (P 0.05). Compared with group IP+ H2, the MWT was significantly decreased and the TWL was shortened at T1-3, the expression of Nrf2 and HO-1 was down-regulated, and the expression of GFAP was up-regulated in group IP+ H2+ ATRA (P<0.05). Conclusion Activation of spinal Nrf2/HO-1 signaling pathway is involved in hydrogen-induced reduction of inflammatory pain in rats. Key words: NF-E2-related factor 2; Heme oxygenase-1; Spinal cord; Hydrogen; Pain; Inflammation