论文部分内容阅读
目的观察鞘内注射针对磷酸二酯酶4B(PDE4B)基因的小干扰RNA(siRNA)对L5脊神经结扎(SNL)大鼠的热痛觉及机械痛觉高敏的影响。方法雄性SD大鼠70只,随机均分为假手术组(Sham组)、生理盐水组(NS组)、单纯载体组(V组)、错配siRNA组(siRNA-M组)和PDE4B-siRNA组(siRNA-B组),分别于神经结扎术后即刻以及术后1、3、5、7d鞘内注射生理盐水、生理盐水、LipofectamineRNAiMAX、错配siRNA 2μg和PDE4B-siRNA 2μg,容量均为10μl。PDE4B-siRNA、错配siRNA与载体均在注射前30min混匀。于术前1d及术后2、4、6、8d分别测定五组大鼠的机械缩足阈值(MWT)和热缩足潜伏期(TWL);术后4、8d行为学测试结束后每组各处死6只大鼠,取腰段脊髓,采用实时定量PCR(real-time PCR,RT-PCR)和western bolt分别测定PDE4B的mRNA和蛋白表达。结果与术前1d比较,术后2、4、6、8dNS组、V组、siRNA-M组和siRNA-B组大鼠MWT明显缩短、TWL明显减小(P<0.05)。与NS组比较,术后2、4、6、8dsiRNA-B组大鼠MWT明显延长、TWL明显增加(P<0.05)。与Sham组比较,术后4、8dNS组、V组、siRNA-M组、siRNA-B组大鼠脊髓组织的PDE4B mRNA、PDE4B蛋白表达升高(P<0.05)。与NS组比较,术后4、8dB组大鼠脊髓PDE4B mRNA、PDE4B蛋白表达明显降低(P<0.05)。结论鞘内注射PDE4B-siRNA可显著抑制L5SNL大鼠腰段脊髓PDE4B的mRNA和蛋白表达,有效缓解机械和热痛觉高敏。
Objective To investigate the effects of intrathecal injection of small interfering RNA (siRNA) targeting phosphodiesterase 4B (PDE4B) gene on thermal hyperalgesia and mechanical hyperalgesia in L5 spinal nerve ligation (SNL) rats. Methods Seventy male Sprague-Dawley rats were randomly divided into four groups: Sham group, NS group, V group, siRNA-M group and PDE4B-siRNA group (SiRNA-B group) were injected intrathecally saline, saline, Lipofectamine RNAiMAX, mismatch siRNA 2μg and PDE4B-siRNA 2μg, respectively, both immediately after nerve ligation and at 1,3,5,7th postoperative day 10 μl. PDE4B-siRNA, mismatch siRNA and vector were mixed 30min before injection. The MWT and TWL of rats in each group were measured at 1 day before operation and at 2, 4, 6, 8 days after operation. Six rats were sacrificed and the lumbar spinal cord was taken. The mRNA and protein expression of PDE4B were measured by real-time PCR (RT-PCR) and western bolt respectively. Results Compared with the preoperative 1d, the MWT, the TWL of the rats in the NS, V, siRNA-M and siRNA-B groups were significantly decreased at 2, 4, 6 and 8 days after operation (P <0.05). Compared with NS group, MWT of rats in 2, 4, 6 and 8dsiRNA-B groups were significantly prolonged and TWL increased significantly (P <0.05). Compared with Sham group, the expression of PDE4B mRNA and PDE4B protein in spinal cord tissue of rats in NS group, V group, siRNA-M group and siRNA-B group increased on the 4th and 8th day after operation (P <0.05). Compared with NS group, the expression of PDE4B mRNA and PDE4B protein in spinal cord of rats in 4 and 8 dB group decreased significantly (P <0.05). Conclusion Intrathecal injection of PDE4B-siRNA significantly inhibited PDE4B mRNA and protein expression in the lumbar spinal cord of rats with L5SNL, effectively relieving mechanical and thermal hyperalgesia.