天麻素对坐骨神经痛模型大鼠TNF-α/STAT3通路及痛觉敏感性的影响

Effects of gastrodin on TNF-α/STAT3 pathway and pain sensitivity in sciatica model rats

目的:探究天麻素对坐骨神经痛大鼠TNF-α/转录激活因子3(STAT3)通路的调控作用及对痛觉敏感性的影响。方法:SD大鼠随机分为假手术组、模型组、天麻素组(120 mg/kg)、TNF-α过表达的重组腺病毒组(Ad-TNF-α,4μl/只)、空腺病毒组(Ad-GFP,4μl/只)、天麻素+Ad-TNF-α组(120 mg/kg+4μl/只),除假手术组外,其余各组均采用坐骨神经压迫法建立大鼠坐骨神经痛模型,每组10只。各组均于造模成功后开始给药,天麻素经腹腔注射给药(1次/d),Ad-GFP及Ad-TNF-α经鞘内注射给药(2次/周),各组连续给药14 d后,观察大鼠一般行为活动;检测热辐射刺激缩爪反应潜伏期(PWL)及机械缩足痛敏阈值(PWT);ELISA检测痛觉敏感相关物质-降钙素基因相关肽(CGRP)、前列腺素E2(PGE2)水平;HE染色检测脊髓病理形态变化;免疫荧光双标记法检测TNF-α与星形胶质活化蛋白(GFAP)共表达水平及TNF-α、STAT3、电压门控钠通道亚型(Nav1.6)与神经元结构蛋白微管相关蛋白2(MAP2)阳性共表达水平;Western blot检测TNF-α、STAT3及其下游蛋白Nav1.6、趋化因子配体2(CCL2)、IL-6、环氧化酶-2(COX-2)的表达。结果:与假手术组相比,模型组大鼠神经纤维排列紊乱且结构破坏严重,CGRP、PGE2水平、TNF-α与GFAP、TNF-α与MAP2、Nav1.6与MAP2阳性共表达水平、TNF-α、STAT3、Nav1.6、CCL2、IL-6、COX-2蛋白表达升高(P<0.05),PWL、PWT降低(P<0.05)。天麻素组神经纤维排列稍整齐且结构破坏缓解,且上述指标变化与模型组相反(P<0.05);Ad-TNF-α组大鼠神经纤维排列紊乱且结构破坏进一步加重,上述指标变化趋势与模型组一致且比模型组严重(P<0.05)。天麻素+Ad-TNF-α组上述指标变化与天麻素组相反,差异有统计学意义(P<0.05)。Ad-GFP组与模型组比较上述指标差异无统计学意义(P>0.05)。结论:天麻素可能通过抑制TNF-α/STAT3通路激活下调Nav1.6表达,降低坐骨神经痛模型大鼠痛觉敏感性。

Objective:To explore regulatory effect of gastrodin on TNF-α/signal transducer and activator of transcription 3(STAT3)pathway and its effect on pain sensitivity in sciatica rats.Methods:SD rats were randomly divided into sham operation group,model group,gastrodin group(120 mg/kg),recombinant adenovirus overexpressing TNF-α group(Ad-TNF-α,4 μl/rat),empty adenovirus group(Ad-GFP,4 μl/rat),gastrodin+Ad-TNF-α group(120 mg/kg+4 μl/rat),sciatica model was established by sciatic nerve compression in all groups except sham operation group,with 10 rats in each group. All groups were given drugs after successful modeling,gastrodin was injected intraperitoneally(once a day),and Ad-GFP and Ad-TNF-α were injected intrathecally(twice a week),after 14 days of continuous administration,general behavior of rats were observed;paw withdrawal latency(PWL)and paw withdrawal threshold(PWT)were measured;levels of calcitonin gene-related peptide(CGRP)and prostaglandin E2(PGE2)were detected by ELISA;HE staining was used to detect pathological changes of spinal cord;co-expression level of TNF-α and glial fibrillary acidic protein(GFAP)and co-expression levels of TNF-α,STAT3,voltage-gated sodium channel subtype(Nav1.6)and neuronal structural protein microtubule-associated protein 2(MAP2)were detected by immunofluorescence double labeling method;Western blot was used to detect expressions of TNF-α,STAT3 and its downstream protein Nav1.6,chemokine ligand 2(CCL2),IL-6 and cyclooxygenase-2(COX-2).Results:Compared with those in sham operation,arrangement of nerve fibers in model group was disordered and the structure was destroyed seriously,levels of CGRP and PGE2,co-expression levels of TNF-α and GFAP,TNF-α and MAP2,Nav1.6 and MAP2,expressions of TNF-α,STAT3,Nav1.6,CCL2,IL-6,COX-2 protein were increased(P<0.05),PWL and PWT were decreased(P<0.05). In gastrodin group,nerve fibers was arranged in order and structure damage was relieved,and changes of the above indexes were opposite to those in model group(P<0.05);in Ad-TNF-α group,arrangement of nerve fibers was disordered and structural damage was further aggravated,change trend of the above indexes was consistent with that in model group and was more serious than that in model group(P<0.05). The above indexes in gastrodin+Ad-TNF-α group were contrary to those in gastrodin group,and the difference was statistically significant(P<0.05). There was no significant difference in the above indexes between Ad-GFP group and model group(P>0.05).Conclusion:Gastrodin may reduce the pain sensitivity of sciatica rats by inhibiting activation of TNF-α/STAT3 pathway and down-regulating expression of Nav1.6.