健脾化痰活血方对亚临床甲减大鼠甲状腺激素水平及脑单羧酸转运蛋白8的影响
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摘要
目的:研究L-T_4对亚临床甲状腺功能减退(SCH)大鼠血清及海马组织中甲状腺激素(TH)、脑单羧酸转运蛋白8(MCT8)的影响,同时比较了中药与西药联用的协同增效作用。方法:Wistar成年雄性大鼠,采用"甲状腺全切+术后L-T_4皮下注射"建立SCH模型。随机分5组:空白对照组、模型组、中药组(健脾化痰活血方13.7 g·kg~(-1)·d~(-1))、L-T_4组(5.0μg·kg~(-1)·d~(-1))、中药+L-T_4组(13.7 g·kg~(-1)·d~(-1)+5.0μg·kg~(-1)·d~(-1))。予处理因素6周,麻醉处死大鼠。放射免疫法(RIA)检测血清中总T_4(TT_4);化学发光免疫分析法(ICMA)检测血清中促甲状腺激素(TSH);光镜下观察经HE染色、Nissl染色的海马组织CA1区细胞及尼氏体形态;ELISA法检测大鼠海马中TT_4水平;RT-PTC及WB法分别检测大鼠海马中MCT8蛋白表达。结果:与空白对照组比较,模型组大鼠血清TSH水平明显升高、TT_4水平无变化、海马TT_4水平明显降低、海马MCT8蛋白灰度值及mRNA积分光密度值均显著升高、海马CA1区细胞形态明显散乱、排列不紧密,细胞不完整、尼氏体数目明显减少,说明SCH模型成立并存在脑损伤;与模型组比较,3个治疗组均可降低SCH大鼠升高的TSH水平,3个治疗组均可改善SCH大鼠血清TSH水平、提高海马组织TT_4水平、降低海马MCT8蛋白及其mRNA表达水平、增加海马CA1区尼氏体的数量,其中,中药与L-T_4联合应用疗效最优,分别优于中药组及L-T_4组。结论:健脾化痰活血方可改善SCH大鼠血清TSH水平、调节脑组织中TH及MCT8水平,与L-T_4联用有协同增效作用。
Objective: To study the effect of L-T_4 on thyroid hormone(TH) and mono carboxylic acid transporter 8(MCT8) in serum and hippocampus of subclinical hypothyroidism(SCH) ratsand compare the synergistic effect of Chinese medicine and Western medicine.Methods: In adult male rats of Wistar, the SCH model was established by "total thyroidectomy+subcutaneous injection of L-T_4 after operation". The rats were randomly divided into 5 groups: control group, model group, traditional Chinese medicine(TCM) group which was given Jianpi Huatan Huoxue Recipe 13.7 g·kg~(-1)·d~(-1), L-T_4 group(5.0 μg·kg~(-1)·d~(-1)),TCM + L-T_4 group(13.7 g·kg~(-1)·d~(-1)+5.0 μg·kg~(-1)·d~(-1)). The treatment lasted for 6 weeksand rats were killed after anesthesia. The total T_4(TT_4) in serum was detected by radioimmunoassay(RIA). The serum levels of thyroid stimulating hormone(TSH) were detected by chemiluminescent immunoassay(ICMA). The CA1 cell and Nissl body morphology of the hippocampus were observed by HE staining and Nissl staining. ELISA was used to detect the level of TT_4 in the hippocampus of rats. The expression of MCT8 protein in the hippocampus of rats was detected by RT-PTC and WB.Results: Compared with the blank control group, the level of serum TSH in the model group was significantly higher, the level of TT4 was not changed, and the level of TT4 in the hippocampus decreased significantly. In the hippocampus, the gray value of MCT8 protein and the value of mRNA integral light density increased significantly. The cell morphology of the hippocampal CA1 area was obviously scattered and not closely arranged. The cells were incomplete, and the number of Nissl body decreased obviously. It indicated that the SCH model was established with brain damage.Compared with the model group, the three treatment groups could reduce the elevated level of TSH in SCH rats. The three treatment groups can improve the serum TSH level of SCH rats, increase the TT_4 level of hippocampal tissue, decrease the expression level of MCT8 protein and mRNA in hippocampus, increase the number of Nissl bodies in hippocampus CA1 region, and the combination of Chinese medicine and L-T_4was better than that of TCM group and L-T_4 group.Conclusion: Jianpi Huatan Huoxue Recipe can improve the level of thyroid function in SCH rats, regulate the levels of TH and MCT8 in brain tissueand has synergistic synergy with L-T_4.
Objective: To study the effect of L-T_4 on thyroid hormone(TH) and mono carboxylic acid transporter 8(MCT8) in serum and hippocampus of subclinical hypothyroidism(SCH) ratsand compare the synergistic effect of Chinese medicine and Western medicine.Methods: In adult male rats of Wistar, the SCH model was established by "total thyroidectomy+subcutaneous injection of L-T_4 after operation". The rats were randomly divided into 5 groups: control group, model group, traditional Chinese medicine(TCM) group which was given Jianpi Huatan Huoxue Recipe 13.7 g·kg~(-1)·d~(-1), L-T_4 group(5.0 μg·kg~(-1)·d~(-1)),TCM + L-T_4 group(13.7 g·kg~(-1)·d~(-1)+5.0 μg·kg~(-1)·d~(-1)). The treatment lasted for 6 weeksand rats were killed after anesthesia. The total T_4(TT_4) in serum was detected by radioimmunoassay(RIA). The serum levels of thyroid stimulating hormone(TSH) were detected by chemiluminescent immunoassay(ICMA). The CA1 cell and Nissl body morphology of the hippocampus were observed by HE staining and Nissl staining. ELISA was used to detect the level of TT_4 in the hippocampus of rats. The expression of MCT8 protein in the hippocampus of rats was detected by RT-PTC and WB.Results: Compared with the blank control group, the level of serum TSH in the model group was significantly higher, the level of TT4 was not changed, and the level of TT4 in the hippocampus decreased significantly. In the hippocampus, the gray value of MCT8 protein and the value of mRNA integral light density increased significantly. The cell morphology of the hippocampal CA1 area was obviously scattered and not closely arranged. The cells were incomplete, and the number of Nissl body decreased obviously. It indicated that the SCH model was established with brain damage.Compared with the model group, the three treatment groups could reduce the elevated level of TSH in SCH rats. The three treatment groups can improve the serum TSH level of SCH rats, increase the TT_4 level of hippocampal tissue, decrease the expression level of MCT8 protein and mRNA in hippocampus, increase the number of Nissl bodies in hippocampus CA1 region, and the combination of Chinese medicine and L-T_4was better than that of TCM group and L-T_4 group.Conclusion: Jianpi Huatan Huoxue Recipe can improve the level of thyroid function in SCH rats, regulate the levels of TH and MCT8 in brain tissueand has synergistic synergy with L-T_4.
引文
[1] Zhongyan Shan, Lulu Chen, Xiaolan Lian,et al.Iodine Status and Prevalence of Thyroid Disorders After Introduction of Mandatory Universal Salt Iodization for 16 Years in China: A Cross-Sectional Study in 10 Cities[J].Thyroid Official Journal of the American Thyroid Association, 2016,26(8):1125-1130.
[2] 黄靖,郭天康,刘静,等.亚临床甲状腺功能减退大鼠骨代谢改变及左旋甲状腺素疗效[J].中华骨质疏松和骨矿盐疾病杂志,2015,8(1):43-47.
[3] 汤莎莎.促甲状腺激素对脂肪细胞促炎症因子分泌的影响[D].天津:天津医科大学,2013.
[4] 阎芳.促甲状腺激素增加肝脏甘油三酯含量的机制研究[D].济南:山东大学,2014.
[5] 姜冬青.促甲状腺激素通过PKA通路抑制3T3-L1脂肪细胞中脂肪甘油三酯脂酶的表达[D].济南:山东大学,2016.
[6] 贾秀娟,谢超,蔡东升,等.甲状腺激素影响神经元信号传导系统的实验研究[C].广州:中华医学会第六次全国内分泌学术会议论文汇编,2011:362-363.
[7] Mayerl S,Muller J,Baure R,et al.Transporters MCT8 and OATP1C1 maintain murine brain thyroid hormone homeostasis[J].Journal of clinical investigation,2014,124(5):1987-1999.
[8] Roberts LM, Woodford K,Zhou M,et al.Expression of the thyroid hormone transporters monocarboxylate transporter-8(SLC16A2) and organic ion transporter-14(SLCO1C1) at the blood-brain barrier[J].Endocrinology,2008,149(12):6251-6261.
[9] 朱帅俊.亚临床甲状腺功能减退症动物模型的建立[D].沈阳:中国医科大学,2008.
[10] 潘立文.温肾方对亚临床甲减雌鼠血清甲状腺功能的影响[J].湖北中医杂志,2011,33(10):20-21.
[11] 杨瑞霞,陈如泉.温肾方对亚临床甲状腺功能减退模型大鼠心肌组织T3及甲状腺激素受体的影响[J].中医学报,2013,28(7):989-991.
[12] 黄婧,郭天康,刘静,等.亚临床甲状腺功能减退大鼠骨代谢改变及左旋甲状腺素效[J].中华骨质疏松和骨矿盐疾病杂志,2015,8(1):43-47.
[13] Da CR,Laureanomelo R,Oliveira KC,et al.Antidepressant behavior in thyrodectomized Wistar rats is induced by hippocampal hypothyroidism[J]. Physiology & Behavior,2016,157:158-164.
[14] Liu YY,Brent GA.Functional analysis of MCT8 mutations identified in patients with X-linked psychomotor retardation and elevated serum triiodothyronine[J].Colloid and Polymer Science,2007,253(2):179.
[15] Liu YY,Brent GA.Thyroid hormone and the brain: Mechanisms of action in development and role in protection and promotion of recovery after brain injury Pharmacology and Therapeutics[J].Colloid and Polymer Science,2018,186:176-185.
[16] Jurgen Jansen,Edith CH,Friesema,et al.Genotype-Phenotype Relationship in Patients with Mutations in Thyroid Hormone Transporter MCT8[J].Endocrinology,2008,149(5):2184-2190.
[17] Stohn JP,Martinez ME,Matoin K,et al. MCT8 Deficiency in Male Mice Mitigates the Phenotypic Abnormalities Associated With the Absence of a Functional Type 3 Deiodinase[J].Endocrinology,2016,157(8):3266
[18] 陈巍,高天舒,张凤暖,等.健脾化痰活血方对中青年亚临床甲减致认知功能损伤患者疗效学观察[J].辽宁中医杂志,2017,44(6):1219-1221.
[19] 崔丹,管玉青,蒋海山,等.甲状腺激素转运体MCT8在2VO大鼠脑组织中的表达[J].南方医科大学学报,2012,32(6):913-915.
[20] 刘迪杰.妊娠期母体亚临床甲减对后代智力和脑发育相关基因表达影响的动物实验研究[D].沈阳:中国医科大学,2009.
[2] 黄靖,郭天康,刘静,等.亚临床甲状腺功能减退大鼠骨代谢改变及左旋甲状腺素疗效[J].中华骨质疏松和骨矿盐疾病杂志,2015,8(1):43-47.
[3] 汤莎莎.促甲状腺激素对脂肪细胞促炎症因子分泌的影响[D].天津:天津医科大学,2013.
[4] 阎芳.促甲状腺激素增加肝脏甘油三酯含量的机制研究[D].济南:山东大学,2014.
[5] 姜冬青.促甲状腺激素通过PKA通路抑制3T3-L1脂肪细胞中脂肪甘油三酯脂酶的表达[D].济南:山东大学,2016.
[6] 贾秀娟,谢超,蔡东升,等.甲状腺激素影响神经元信号传导系统的实验研究[C].广州:中华医学会第六次全国内分泌学术会议论文汇编,2011:362-363.
[7] Mayerl S,Muller J,Baure R,et al.Transporters MCT8 and OATP1C1 maintain murine brain thyroid hormone homeostasis[J].Journal of clinical investigation,2014,124(5):1987-1999.
[8] Roberts LM, Woodford K,Zhou M,et al.Expression of the thyroid hormone transporters monocarboxylate transporter-8(SLC16A2) and organic ion transporter-14(SLCO1C1) at the blood-brain barrier[J].Endocrinology,2008,149(12):6251-6261.
[9] 朱帅俊.亚临床甲状腺功能减退症动物模型的建立[D].沈阳:中国医科大学,2008.
[10] 潘立文.温肾方对亚临床甲减雌鼠血清甲状腺功能的影响[J].湖北中医杂志,2011,33(10):20-21.
[11] 杨瑞霞,陈如泉.温肾方对亚临床甲状腺功能减退模型大鼠心肌组织T3及甲状腺激素受体的影响[J].中医学报,2013,28(7):989-991.
[12] 黄婧,郭天康,刘静,等.亚临床甲状腺功能减退大鼠骨代谢改变及左旋甲状腺素效[J].中华骨质疏松和骨矿盐疾病杂志,2015,8(1):43-47.
[13] Da CR,Laureanomelo R,Oliveira KC,et al.Antidepressant behavior in thyrodectomized Wistar rats is induced by hippocampal hypothyroidism[J]. Physiology & Behavior,2016,157:158-164.
[14] Liu YY,Brent GA.Functional analysis of MCT8 mutations identified in patients with X-linked psychomotor retardation and elevated serum triiodothyronine[J].Colloid and Polymer Science,2007,253(2):179.
[15] Liu YY,Brent GA.Thyroid hormone and the brain: Mechanisms of action in development and role in protection and promotion of recovery after brain injury Pharmacology and Therapeutics[J].Colloid and Polymer Science,2018,186:176-185.
[16] Jurgen Jansen,Edith CH,Friesema,et al.Genotype-Phenotype Relationship in Patients with Mutations in Thyroid Hormone Transporter MCT8[J].Endocrinology,2008,149(5):2184-2190.
[17] Stohn JP,Martinez ME,Matoin K,et al. MCT8 Deficiency in Male Mice Mitigates the Phenotypic Abnormalities Associated With the Absence of a Functional Type 3 Deiodinase[J].Endocrinology,2016,157(8):3266
[18] 陈巍,高天舒,张凤暖,等.健脾化痰活血方对中青年亚临床甲减致认知功能损伤患者疗效学观察[J].辽宁中医杂志,2017,44(6):1219-1221.
[19] 崔丹,管玉青,蒋海山,等.甲状腺激素转运体MCT8在2VO大鼠脑组织中的表达[J].南方医科大学学报,2012,32(6):913-915.
[20] 刘迪杰.妊娠期母体亚临床甲减对后代智力和脑发育相关基因表达影响的动物实验研究[D].沈阳:中国医科大学,2009.