氟过量与碘氟过量对甲状腺功能和形态影响的实验研究

【作者】 国秀娟；

【导师】 滕卫平；

【作者基本信息】 中国医科大学 ， 内科学， 2002， 硕士

【摘要】 前言 碘与氟是人体必需的两种微量元素，机体对碘、氟的摄取主要取决于外环境。本课题组于实现全民食盐碘化(universal salt iodated USI)法规3年后对辽宁、河北三个不同碘摄入量农村社区(人群尿碘中位数分别为103μg/L、374μg/L、615μg/L)进行甲状腺疾病的流行病学调查，结果显示：碘摄入量增加导致甲状腺功能减退症、自身免疫甲状腺炎显著增加，乳头状甲状腺癌患病率增加。同时我们发现，在高碘地区存在高氟的因素，氟过量可引起机体不同程度的代谢紊乱和中毒表现，如氟斑牙、氟骨症，氟过量对甲状腺的影响是国际学者长期争论的问题，有关于此的流行病学调查和动物实验研究结果亦不一致，氟对甲状腺影响机制尚不明确。本实验拟通过对大鼠碘氟代谢，甲状腺激素水平测定及形态学的观察来探讨氟过量及碘氟过量对甲状腺的影响，以明确氟在甲状腺损伤中的作用，更好地了解微量元素之间的复杂关系和对人类健康的影响，为防治措施提供理论依据。 实验材料 一、实验动物 选用我校实验动物部提供的5周龄Wistar大鼠160只，体重125～165g，雌雄各半，随机分8组。实验期间饲以普通饲料和去离子水。 二、主要仪器 电子天平；自动体重秤；不锈钢代谢笼；熔蜡箱；水浴锅；温箱；pH计；-20℃冰箱；-70℃冰箱；石蜡切片机；离心机；透射电镜；光学显微镜；显微图像分析仪；万能显微镜。 实验方法 一、动物分组与处理 将5周龄WISthe大鼠160只，体重125J，随机分为8组，每组20只，雌雄各半。选用动物部提供的普通饲料，分别自由饮用不同浓度的氟化钠配制的去离子水，碘过量组为同一浓度，用碘酸钾配制。分组如下： 对照组：去离子水 氟过量二组：含氟15ppm的去离子水 （相当于3倍高氟） 氟过量11组：含氟30ppm的去离子水 （相当于6倍高氟） 氟过量皿组：含氟60ppm的去离子水 （相当于12倍高氟） 碘过量组：含碘1200pglL的去离子水 （相当于6倍高碘） 碘氟过量互组：含碘1200pgiL、氟15ppm的去离子水 （相当于6倍高碘3倍高氟） 碘氟过量D组：含碘 1200pg／L、氟 30ppm的去离子水 （相当于6倍高碘在倍高氟） 碘氟过量皿组：含碘1200Pg／L、氟60PPm的去离子水 （相当于6倍高碘＊ 倍高氟） 实验早期第75天处死一半动物，每组10只，后期第150天处死剩余大鼠。 二、标本收集及处理 二．尿 2．体重 3．血清 ·2· 4．甲状腺及其湿重 5．常规组织形态学观察 O）石蜡切片的制作过程 固定；脱水；透明；浸蜡；包埋；切片。 （2）HE染色 脱蜡；水化；苏木精染色；二％盐酸酒精分化；返蓝；伊红染色；水洗；脱水；透明；封片。 6．透射电镜超微结构观察 三、血清垂体——甲状腺激素水平 ’IT、’IT用免疫放射法（RIAXhH采用固相免疫放射分析法（IRMA）。 四、尿碘尿氟测定 尿碘采用砷钵催化分光光度法，尿氟采用氟离子选择电极法。 五、图像采集与分析 六、数据统计学处理 应用SPSS10．0统计软件进行单因素方差分析＊值检验，进一步比较用 SNK检验＊检验L结果用 k。SD表示，以 P＜0．05为有统计学意义。 实验结果 一、尿碘测定结果 碘过量组及碘氟过量组75天上50天尿碘水平与对照组比较明显升高帅＜0．05人碘氟过量各组与碘过量组比较无显著差异（P＞0．05）。 二、尿氟测定结果’ 氟过量皿组75天，碘氟过量皿组的尿氟水平与对照组比较明显升高帅＜0．05入其 150天水平与对照组比较明显增加h＜0·05入与 75天水平比较亦明显升高h＜0刀5入碘氟过量 I、匝组 ·3·与对照组比较无明显差异帅＞0．05人 三、血清垂体——甲状腺激素测定结果 二．冗 75天＊ 天各处理组大鼠 ’IT 7k平与对照组比较无明显差异h＞0．05入碘氟过量各组与碘过量组比较无明显差异h＞0．05人 150天氟过量 K、皿组几水平分别比同浓度 75天水平明显降低（p＜0．05）。 2。h 75天时各对照组 h水平与对照组比较无明显差异仲＞0．05入150天碘过量组 rIT水平与对照组比较无明显差异h＞0·05入碘氟过量各组兀水平与对照组比较无明显差异帅＞0．0幻，150天氟过量各组 ’IT水平与对照组比较明显降低帅＜0．05X较其 75天水平亦明显降低（p＜0．05）。 3．TSH 75天＊50天各处理组大鼠hH水平与对照组比较无明显差异h＞0．05X碘氟过量各组与碘过量比较亦无明显差异h＞0．05h碘过量组 150天水平与其 75天比较无显著差异讣＞0．05八 4．甲状腺?更多还原

【Abstract】 Iodine and fluorine are two kinds of essential trace elements for human being. The intake of them is more dependent on the environment surrounded. Our government has established the law of universal salt iodated ( USI) from 1996. In order to acknowledge the situation of thyroid disease in population, we have taken an cross - section study in three different iodine intake regions. The results show the increasing incidence of hypothyroidism and thyroid carcinoma. Due to the u-biquitous nature of exposures to high iodine and fluorine in China and the issue of whether fluorine affected thyroid is still controversial, we hope to investigate the action of excess fluorine on thyroid. According to the concentrations of fluorine and iodine in the studied area, the animal experiment was designed to evaluate the effects of single and combined excess of fluorine and iodine on the thyroid gland function and structure in rats to investigate the mechanism to explore complex relationship between the trace elements and to afford the theory for therapy."iMaterials and Methods1. animals and the treatmentOne hundred and sixty Wistar rats of both sexes, 5 weeks old (125 -165g body weight) provided and treated by the laboratory animal department of China Medical University, were divided randomly into 8 groups and were individually housed and fed normal diets. Rats of three groups had free access to deionized drinking water with three different concentrations of fluorine respectively ( 15ppmF, 30ppmF, 60ppmF). One excess iodine group ( 1200ug/L I) , and other threegroups were given respectively three different excess fluorine added the equivalent concentration of excess iodine as above - mentioned. Two groups were given the deionized water without fluorine and iodine as controls.Rats were anaesthetized with ether half - and -.half after feeding for 75 days and 150 days. Blood collected by cardiac puncture before the rats were killed, was transferred to a test tube with no anticoagulant to obtain serum by centrifugation, then was immediately frozen at - 70℃ until analysis. The thyroid gland removed immediately after the death of rats was measured the weight and was stored under two different conditions as follows: (1) fixed in 4% PFA (0. 1M PBS, pH7.4 at 4℃ ) for light microscopy studies. (2) fixed in 2.5% glut-araldehyde (pH7.3 at 4℃ ) for electron microscopy studies.2. Assay methodsSerum T3 and T4 concentrations were determined by the RIA method and TSH concentration by the IRMA method. All kits were purchased from DPC (USA). Urinary iodine was measured according to the Ce/As method and urinary fluorine by fluorine ion selective e-lectrode.3. Thyroid Morphology and morphological examination Thyroid lobe from each rat was embedded in paraffin. Sections(3um) were stained with hematoxylin and eosin. We measured the area of colloid lumens, the heights of epithelial cells and the heights of cell nucleus using Metamorph/DP10/BX51.4. Transmission electron microscopyThyroid tissue were prepared for electron microscopy using standard procedures and the ultrathin sections were examined and photographed by a JEOL 1200 EX electron microscope.5. Statistical AnalysisData were analyzed using SPSS 10. 0 for Windows software. The results were expressed as the mean ± SD. Comparisons between groups were performed by one - way ANOVA, followed by the SNK test, to detect significance of difference among multiple groups p <0.05 indicated a statistically significant difference.Results1. Urinary iodineAt two time points, the levels of urinary iodine in the excess iodine group and all excess, iodine - fluorine groups were significantly higher than in the control group.2. Urinary fluorineAt the 75th day, the levels of urinary fluorine in excess fluorine III group and excess iodine - fluorine III group were significantly higher than in the control group, and the levels at the 150th day were significantly higher than in the control group or their levels at the 75th day.3. Serum TT3, TT4and更多还原