Effects of intrathecal DREAM-shRNA on expression of p-CREB in spinal dorsal horn in a rat model of neuropathic pain

Abstract

Objective To investigate the effects of intrathecal (IT) DREAM-short hairpin RNA (DREAM-shRNA) on expression of phosphorylated cyclic AMP response element binding protein (p-CREB) in the spinal dorsal horn in a rat model of neuropathic pain. Methods Adult male SD rats weighing 280-320 g were anesthetized with intraperitoneal 10% chloral hydrate. Neuropathic pain was induced by chronic constrictive injury (CCI) to sciatic nerve. IT catheters were placed according to the method described by Yaksh on 3rd day after CCI. Twenty-four rots in which IT catheter was successfully implanted were randomly divided into 4 groups (n = 6 each) : group Ⅰ sham operation (group S) ; group Ⅱ neuropathic pain (group NP) ; group Ⅲ RNA interference (group RNAi) and group Ⅳ blank vector (group BV). Lentivius with DREAM-shRNA 5 μl was injected IT in group RNAi, and blank vector 5 μl in group BV, and once a day for 7 days, starting from the day 8 after CCI. The mechanical pain threshold was measured at day 1 before CCI (T0 ,baseline) and day 7-14 after CCI (T1-8). The animals were killed on 15th day after CCI. The L4-6 lumbar segment of the spinal cord was removed for determination of the expression of green fluorescent protein (GFP) and p-CREB by immuno-fluorescent method.Results The mechanical pain threshold was significantly decreased as compared with the baseline at T0 in all 4 groups and returned to the baseline levels at T5-8 in group S and RNAi, but remained low in group NP and BY. The mechanical pain threshold was significantly lower after CCI/sham operation and significanty higher at T8 in group RNAi than in the other 3 groups. The expression of p-CREB in the spinal dorsal horn was up-regulated in group NP, RNAi and BV as compared with group S, and in group NP and BV as compared with group RNAi. The green fluorescence was observed in group RNAi but not in the other 3 groups. Conclusion IT DREAM-shRNA can ameliorate neuropathic pain in rats through inhibiting the expression of p-CREB in the spinal dorsal horn. Key words: RNA interference; Transcription factors ; Injections, spinal; Neuralgia; DNA-binding protein, cyclic AMP responsive ; Spinal cord