温肾方对甲状腺功能减退症大鼠神经信号传导及基因表达的影响

【作者】 刘祖发；

【导师】 陈如泉；

【作者基本信息】 湖北中医学院 ， 中医内科学， 2004， 博士

【摘要】 目的 甲状腺功能减退症是一种常见的内分泌综合症侯群，可导致全身多个系统病变，引起各种合并症。甲状腺功能减退症合并脑功能损害临床较为常见，这种甲减性脑损害，随着病情加重，几乎所有病例都会出现，表现为不同程度神经精神症状。目前，临床对甲减性脑损害并无专门的治疗，现代医学仅是针对甲状腺功能减退本身，采用甲状腺素替代治疗，忽视对脑功能损害自身的治疗，疗效不如人意。发挥中医优势，运用中医药治疗甲减合并症，有利于提高本病的疗效。温肾方临床效果良好，不仅可改善甲减本身的征状，而且对神经精神症状改善明显。我们在温肾方治疗甲减性脑损害的临床的基础上，以甲状腺功能减退症动物模型为实验对象，运用现代医学的理论和试验方法，对温肾方治疗甲减性脑损害的作用及其机理进行了研究，从而为临床提供了理论与实验依据。 方法 将受试大鼠平均分为五组：正常对照组、模型组、甲状腺片组、中药小剂量组、中药大剂量组。后四组予以造模，模型大鼠按1.35ml／200g体重／只，每日灌服0.04％他巴唑混悬液，共计造模45天。造模完成后开始灌服相应药物，连续15天。然后各实验组分别进行如下指标的检测：采用放射免疫法测定血清T₃、T₄、FT₃、FT₄、TSH水平；选用水迷宫法、穿梭箱法观察药物对模型大鼠行为学障碍的影响；运用逆转录-多聚酶联反应（RT-PCR）、蛋白免疫印迹法、免疫组织化学的方法、Lowery法及改良Takai法，分别检测模型大鼠海马NO、nNOS含量及nNOSmRNA表达、Goa蛋白含量及GoamRNA表达、PKC活性测定。 结果 1．模型组大鼠造模后出现肾阳虚的症状，反映甲减程度的临床指标血清T₃、T₄、FT₃、FT₄的含量显著低于正常对照组，血清TSH的含量显著高于正常对照组。中药组尤其中药大剂量组不仅能明显改善试验大鼠甲减症状，而且能明显升高T₃、T₄、FT₃、FT₄水平，降低TSH含量，但不强于甲状腺片组。2．温肾方能显著改善模型大鼠行为学障碍，Morris水迷宫实验表现为游泳逃避潜伏期及游泳路径总长明显缩短，同时穿梭箱实验也表现为电击次数（被动逃避次数）及电击时间明显减少，且中药大剂量组优于甲状腺片组。3．温肾方能明显升高模型大鼠NO生成和释放的含量，调高NO信号系统的基因表达；能明显下调模型大鼠GoamRNA表达水平，降低Goa蛋白含量；能明显下调模型大鼠PKC活性，其作用强于甲状腺片组。 结论 温肾方对甲减模型大鼠不仅能明显改善实验大鼠甲减症状，而且能明显升高T₃、T₄、FT₃、FT₄水平，降低TSH含量，但不强于甲状腺片组。温肾方能显著改善模型大鼠行为学障碍，能明显升高模型大鼠NO生成和释放的含量，调高NO信号系统的基因表达；能明显下调模型大鼠GoamRNA表达水平，降低Goa蛋白含量；能明显下调模型大鼠PKC活性，其作用均强于甲状腺片组。表明温肾方对甲减性脑损害治疗作用显著。温肾方大剂量组对模型大鼠行为学障碍改善优于甲状腺片组，进一步表明温肾方对甲减性脑损害的治疗作用，除增加内源性甲状腺素分泌外，还能直接作用于脑，如改善脑信号传导系统，是通过多途经发挥作用的。更多还原

【Abstract】 Objective Hypothyroidism is a common disease of endocrine system, which could to lead to pathological changes and syndromes of many systems. It is common in clinical that hypothyroidism merging the impairment of brain and with the development of hypothyroidism, every patient have the impairment of brain, showing neurotic and psychiatric symptoms. Now only thyroxine sodium is used in treatment of hypothyroidism itself and has no specially therapeutic methods for the impairment of brain caused by hypothyroidism in clinic, and the effect is dissatisfaction. So it is therapeutic current with Chinese medicine for prevention and treatment of the impairment of brain caused by hypothyroidism by giving full play to the advantages of Traditional Chinese Medicine. It was confirmed that the therapeutic effect of Wenshentang in clinic is well. It could improve the symptoms not only hypothyroidism itself but also the impairment of brain caused by hypothyroidism. We study the function of Wenshentang on the impairment of brain caused by hypothyroidism, and its mechanism under the basic theory and experimental methods of modern medicine with the hypothyroidism animal model in order to provide a theory and experimental base for the clinical practice.Methods The rats were randomly divided into 5 groups, such as normal control group, model group, thyroxin group, small dosage group of Wenshentang and the large dosage of Wenshentang. The rats of the latter 4 groups were made model by infusing 0.04% tapazole suspension 1.35%/200g/day for 45 days. After that, the rats were treated with corresponding medicines for 15 days. Then the following detection were taken out: radioimmunoassay on serum FT3, FT4, T3, T4 and TSH; water labyrinth and shuttling box methods on the behavior changes; RT-PCR, Western blot, Lowery, improved Takai and immunohistochemical method on the content or expression of NOmRNA, Go a mRNA and the activity of protein kinase C (PKC) in the hippocampus of hypothyroidism rats.Results (1) In the rats of model group, the behavior changes of kidney yang-insuf f iciency in hypothyroidism were appeared, and the indexes which may imply the severe degree of hypothyroidism presented the decreasing of FT3, FT4, T3, T4 and the increasing of TSH compared with the normal control group. Wenshentang, especially with large dosage had not only improved the symptoms of hypothyroidism, but also increased the content of FT3, FT4, T3, T4 and decreased the content of TSH, but the effect was not better than that of thyroxin group. (2) Wenshentang significantly improved the behavior disorder of the hypothyroidsm rats. The evasive latent period and length of swimming were significantlyshorten in the experiment of Morris water labyrinth, at the same time the times and the hours of get electric shock were significantly decreased in the experiment of shuttling box. The effect of large dosage group was better than that of thyroxin group. (3) Wenshentang significantly increased the content or expression of NOmRNA, decreased the content or expression of Go a mRNA and downregulation the activity of PKC of the hypothyroidsm rats, and the effect was better than that of thyroxin group.Conclusion In the rats of model group, the behavior changes of kidney yang-insufficiency in hypothyroidism were appeared , and the indexes which may imply the severe degree of hypothyroidism presented the decreasing of FT3, FT4, T3, T4 and the increasing of TSH compared with the normal control group. Wenshentang had not only improved the symptoms of hypothyroidism rats, but also increased the content of FT3, FT4, T3, T4 and decreased the content of TSH, but the effect were not excel the thyroxin group. Wenshentang significantly increased the content or expression of NOmRNA, decreased the content or expression of Go a mRNA and down regulation the activity of PKC of the hypothyroidism rats, and the effect were excel the thyroxin group. Wenshentang significantly improved the behavior disorder of the hypothyroidism rats which implied that Wenshentang have signific更多还原