槲皮素调控p38MAPK信号通路促进大鼠急性脊髓损伤修复的研究

发布时间：2018-07-11 15:01

  本文选题：槲皮素 + 脊髓损伤　； 参考：《南方医科大学》2017年硕士论文

【摘要】：目的:通过使用黄酮类化合物槲皮素治疗大鼠急性脊髓损伤(spinal cord injury,SCI),验证其治疗效果,并探讨其可能的作用机制。方法:取128只成年健康雌性SD大鼠,随机等分为对照组、干预组、治疗组和假手术组。对照组、干预组及治疗组大鼠使用自制脊髓损伤打击器建模:常规麻醉并消毒,摘除T9-T11椎板后暴露相应脊髓节段,采用10g×25mm致伤势能对脊髓进行垂直打击建模。治疗组及干预组大鼠于术后1h给予腹腔注射槲皮素(25μmol/kg,12h/次),持续7日;干预组大鼠在注射槲皮素后立即在对侧腹腔注射p38MAPK特异性激活剂anisomycin;假手术组大鼠仅行椎板切除术。对照组及假手术组大鼠术后予以与治疗组等量、等频率的生理盐水腹腔注射。大鼠建模成功后按摩膀胱排尿3次/日,直到恢复反射性膀胱;术后7日内所有大鼠行氨苄青霉素腹腔注射预防感染。术后1d、3d、7d、14d、21d及28d对各组大鼠进行BBB评分。术后3d、7d、14d及28d时每组随机取8只大鼠麻醉并行心脏灌注,灌注成功后切取长约1cm的脊髓组织,纵切为均匀的左右两半,取其中一半保存于-80℃冰箱,以备行Western blot蛋白检测,另一半组织在4%多聚甲醛中浸泡24h用于免疫组化或HE染色。术后28d对各组大鼠脊髓组织进行HE染色并在显微镜下观察。术后3d、7d、14d及28d对各组大鼠脊髓组织的胶质纤维酸性蛋白(glial fibrillary acidic protein,GFAP)和神经丝蛋白-200(neurofilament protein-200,NF-200)进行免疫组化染色,显微镜下观察,统计分析各组的阳性细胞数目。术后3d、7d、14d及28d对各组大鼠脊髓组织的p38MAPK、磷酸化p38MAPK(phosphorylation p38MAPK,p-p38MAPK)蛋白的表达情况进行Western blot检测,术后28d对各组大鼠脊髓组织的GFAP和NF-200蛋白进行Western blot检测。结果:术后7d、14d、21d及28d,治疗组大鼠BBB评分显著高于对照组与干预组(P0.05)。术后3d和7d各脊髓损伤组的p38MAPK磷酸化水平明显高于假手术组(P0.05),治疗组的磷酸化水平在术后3d、7d及14 d时明显低于对照组与干预组(P0.05)。各脊髓损伤组在术后各时间点NF-200和GFAP阳性细胞数均高于假手术组(P0.05),治疗组的NF-200阳性细胞数较对照组和干预组显著增高(P0.05);术后7d、14d和28d,治疗的GFAP阳性细胞数明显低于对照组与干预组(P0.05)。结论:槲皮素通过抑制急性脊髓损伤后p38MAPK信号通路的激活,促进了 NF-200的表达及神经元轴突再生,抑制了 GFAP表达及星形胶质细胞的增生、阻碍了胶质瘢痕的形成,修复受损脊髓的神经传导作用,从而促进急性脊髓损伤后运动功能的恢复。
[Abstract]:Aim: to investigate the therapeutic effect of quercetin on acute spinal cord injury (spinal cord injura) in rats and its possible mechanism. Methods: 128 adult female SD rats were randomly divided into control group, intervention group, treatment group and sham operation group. Control group, intervention group and treatment group used self-made spinal cord injury batter model: routine anesthesia and disinfection, after removing T9-T11 vertebral lamina, the corresponding spinal cord segment was exposed. The injury induced by 10 g 脳 25mm could be used to model the spinal cord vertical strike. Rats in the treatment group and the intervention group were intraperitoneally injected with quercetin (25 渭 mol / kg, 12 h / time) for 7 days, and the rats in the intervention group were injected with p38 MAPK specific activator anisomycin in the contralateral abdominal cavity immediately after injection of quercetin, while rats in the sham operation group were treated with laminectomy only. Rats in the control group and sham operation group were given intraperitoneal injection of saline with the same amount and frequency after operation as the treatment group. After successful modeling, the bladder urination was massaged 3 times a day until the reflex bladder was restored, and all rats received ampicillin intraperitoneal injection to prevent infection within 7 days after operation. BBB scores were evaluated on the 1st day, 3rd day, 7th day, 14 d, 21 d and 28 d after operation. 8 rats in each group were anesthetized and perfused on the 14th and 28th days after the operation. The spinal cord tissue about 1cm was cut out and divided into two parts, half of which was stored in the refrigerator at -80 鈩，

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