肾虚型慢性再生障碍性贫血小鼠模型建立

发布时间：2017-12-28 08:17

本文关键词：肾虚型慢性再生障碍性贫血小鼠模型建立　出处：《中国中西医结合杂志》2017年09期 　论文类型：期刊论文

【摘要】：目的建立肾虚型慢性再生障碍性贫血(chronic aplastic anemia,CAA)小鼠模型。方法将70只小鼠行体能测试,筛选出体能相近的小鼠随机分成肾阴虚组(20只)、肾阳虚组(20只)及正常组(10只),其余随机分成苯组(10只)及玉米油组(10只)。肾阴虚组每日灌胃甲状腺素片水溶液(350 mg/kg),共21天;肾阳虚组每日皮下注射氢化可的松(25 mg/kg),共14天。正常组皮下注射生理盐水(25 mg/kg),共14天,并灌胃生理盐水(350 mg/kg),共21天。在肾虚模型建立的基础上,给予皮下注射苯试剂和玉米油混合液2 m L/kg,每周3次,共25次,建立肾虚型CAA小鼠模型。相应药物介入1 d起,观察小鼠日常生活状态,通过体能测试监测游泳时间;采用ELISA试剂盒测定血清T3、T4、尿17-羟皮质类固醇(17-hydroxy-cortico steroid,17-OHCS)、环磷酸腺苷(c AMP)、环磷酸腺苷/环磷酸鸟苷(c AMP/c GMP)水平;血细胞分析仪检测白细胞、血红蛋白及血小板数量;电子显微镜下计数骨髓有核细胞;HE染色观察骨髓病理特征。结果药物干预25 d后,与正常组比较,肾阴虚组游泳耗时延长、肾阳虚组游泳耗时缩短(P0.01)。药物干预22 d后,肾阴虚组小鼠T3、T4、17-OHCS、c AMP、c AMP/c GMP水平上升;肾阳虚组T3、T4、17-OHCS、c AMP/c GMP水平下降(P0.05,P0.01)。苯试剂介入第12次时,与正常组比较,肾阴虚+苯组、肾阳虚+苯组及苯组白细胞、血小板计数降低(P0.05,P0.01)。苯试剂介入第18次时,与正常组比较,肾阴虚+苯组、肾阳虚+苯组及苯组白细胞、血红蛋白、血小板计数均下降(P0.05,P0.01)。药物介入75 d,肾阴虚+苯组、肾阳虚+苯组及苯组骨髓有核细胞数均下降(P0.05)。药物介入75 d,苯试剂导致骨髓增生明显低下,血白细胞、血红蛋白及血小板计数均有不同程度减少,非造血细胞比例增高。结论灌胃甲状腺素法联合皮下注射苯试剂法可成功建立肾阴虚型CAA小鼠模型;皮下注射氢化可的松法联合皮下注射苯试剂法可成功建立肾阳虚型CAA小鼠模型。
[Abstract]:Objective to establish a mouse model of chronic aplastic anemia (CAA) with kidney deficiency. Methods a total of 70 mice were selected for physical performance test, and the mice with similar physical strength were randomly divided into 20 groups: kidney yin deficiency group (20 cases), kidney yang deficiency group (20 cases) and normal group (10). The rest were randomly divided into benzene group (n = 10) and corn oil group (10 rats). The kidney yin deficiency group was injected with water solution of thyroxine tablets (350 mg/kg) daily for 21 days, and the hypodermic injection of hydrocortisone (25 mg/kg) daily for 14 days in the kidney yang deficiency group. Normal saline (25 mg/kg) was subcutaneously injected into the normal group for 14 days, and the saline (350 mg/kg) was administered to the stomach (350 mg/kg), for a total of 21 days. On the basis of the establishment of kidney deficiency model, subcutaneous injection of benzene reagents and corn oil mixture 2 m L/kg, 3 times a week, a total of 25 times, the establishment of kidney deficiency type CAA mice model. The corresponding drug intervention 1 D, observed the state of daily life, through the physical fitness test monitoring swimming time; serum T3, T4, 17- in urine hydroxycorticosteroid using ELISA Kit (17-hydroxy-cortico steroid, 17-OHCS), cyclic adenosine monophosphate (C AMP), cyclic adenosine monophosphate / cyclic guanosine monophosphate (C AMP/c GMP); the number of blood cell analyzer detection of white blood cell, hemoglobin and platelet count under electron microscope; bone marrow cell; bone marrow pathology HE staining. Results after 25 D, compared with the normal group, the swimming time of the kidney yin deficiency group was prolonged and the time of swimming in the kidney yang deficiency group was shortened (P0.01). After 22 D intervention, the levels of T3, T4, 17-OHCS, C AMP and C AMP/c GMP increased in the kidney yin deficiency group, while the levels of T3, T4, GMP, and gamma in the kidney yang deficiency group decreased. Compared with the normal group, the kidney yin deficiency + benzene group, kidney yang deficiency + benzene group and benzene group white blood cell and platelet count decreased (P0.05, P0.01) compared with the normal group for Twelfth times. When benzene reagent was intervened eighteenth times, compared with the normal group, the kidney yin deficiency + benzene group, kidney yang deficiency + benzene group and benzene group white blood cell, hemoglobin and platelet count decreased (P0.05, P0.01). Drug intervention 75 D, kidney yin deficiency + benzene group, kidney yang deficiency + benzene group and the number of nucleated cells in the benzene group were decreased (P0.05). Drug intervention for 75 D, benzene reagents caused bone marrow hyperplasia was significantly lower, white blood cells, hemoglobin and platelet count decreased to varying degrees, the proportion of non hematopoietic cells increased. Conclusion the CAA mice model of kidney yin deficiency can be successfully established by intragastric administration of thyroxine combined with subcutaneous injection of benzene reagent. Subcutaneous injection of hydrocortisone combined with subcutaneous injection of benzene reagent can successfully establish a CAA mouse model of kidney yang deficiency.
【作者单位】：浙江大学医学院附属第一医院急诊科;杭州市余杭区中医院肿瘤内科;浙江中医药大学第一临床医学院;浙江省丽水市中医医院;浙江中医药大学动物实验研究中心;浙江中医药大学附属第一医院血液科;
【基金】：国家自然科学基金资助项目(No.81373634) 浙江省高等教育质量工程-中青年学科带头人资助项目(No.GK2011,No.GK2012)
【分类号】：R259;R-332
【正文快照】： 慢性再生障碍性贫血(chronic aplastic anemia,CAA)具有病情反复、疗程较长、可缓解却难于治愈等特点,中医学将其纳入“虚劳”范畴,根据临床表现,通常分为肾阴虚、肾阳虚、肾阴阳两虚3型。中医药在治疗上以补肾为本,辅以健脾、活血化瘀、清热解毒等法,能明显增强机体免疫功能,

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