miR-16对神经病理性疼痛模型大鼠脑干中缝核SERT表达作用的研究

目的 通过研究miR-16对神经病理性疼痛模型大鼠脑干中缝核中5羟色胺转运体(SERT)表达的作用,探讨miR-16通过作用于5羟色胺(5HT)再摄取影响神经病理性疼痛的可能机制。 方法 选用雄性6周龄SD大鼠40只,采用慢性坐骨神经结扎法(chronic constriction injury,CCI)制备大鼠病理性神经痛模型。将大鼠随机分为4组:假手术组(S组),仅分离坐骨神经鞘不结扎;坐骨神经损伤组(CCI组),松弛结扎坐骨神经;坐骨神经损伤+氟西汀组(CCI/F组);对照组(C组),不做任何处理,每组10只。术前1 d及术后第1、3、5、7、14天测定机械缩足痛阈值(MWT);14 d后处死大鼠,取大鼠脑干中缝核组织,通过实时定量PCR法(qRT-PCR)检测miR-16表达情况;western blot检测SERT蛋白表达情况;组织匀浆后,ELISA法检测皮质组织内5羟色胺(5HT)浓度。 结果 与术前相比,CCI组大鼠第1天MWT即出现下降,与S组相比差异具有统计学意义(t=5.167,P<0.001),CCI/F组与S组相比MWT显著升高,差异具有统计学意义(t=4.664,P<0.001),而S组及C组未见明显变化。qRT-PCR实验显示,与S组相比CCI组大鼠脑干中缝核组织匀浆中内源性miR-16表达显著降低(t=11.840,P<0.001),CCI/F组升高(t=28.640,P<0.001);western blot显示CCI组大鼠脑干中缝核SERT蛋白表达升高,CCI/F组SERT表达降低;ELISA实验显示CCI组皮质中5HT水平较S组显著降低(t=17.920,P<0.001),而CCI/F组5HT水平升高(t=7.232,P=0.002),差异均具有统计学意义。 结论 miR-16能够通过抑制脑干中缝核SERT表达,减少5HT再摄取影响病理性神经痛的维持。 

Effect of miR-16 on SERT expression in the rapheal nuclei of brain stem of neuropathic pain rat

Objective To explore the mechanism of the affection of miR-16 on 5HT reuptake in the neuropathtic pain by studying the miR-16 regulating SERT expression. Methods Forty adult male SD rats were randomly divided into control group (C group),sham operation group (S group),chronic constriction injury group (CCI group),chronic constriction injury plus fluoxetine group (CCI/F group),10 rats per group.Left sciatic nerve of rats in CCI group were freed and loosely compressed by sutures,while rats in the S group were only freed the neurilemma.The mechanism withdraw threshold (MWT) was test in the preoperative day and the day 1,3,5,7,14 after surgery.Fourteen days after surgery,the rats were sacrificed,and the rapheal nuclei of brain stem were taken.The miR16 expression was tested by qRT-PCR,and SERT was tested by western blot.The 5HT level in the tissue homogenate was analyzed by ELISA assay. Results Compared with the S group,the MWT in the CCI group was significantly declined in the first day after surgery (t=5.167,P<0.001),but the value was increased in the CCI/F group (t=28.640,P<0.001).SERT in the rapheal nuclei of CCI group was increased but decreased in the CCI/F group as shown by western blot.Compared with the S group,5HT in the rapheal nuclei of CCI group was significantly decreased (t=17.92,P<0.001),but increased in the CCI/F group (t=7.232,P=0.002). Conclusion miR-16 can regulate the expression of SERT in the rapheal nuclei,thereby affect the reuptake of 5HT to influence the neuropathic pain.