Study On The Effects Of Ammonium Perchlorate On Thyroid-specific Gene And Thyroid Hormone Levels Of Workers

Ammonium perchlorate (AP) is a kind of inorganic white crystalline powder. AP is the most prevalent form of perchlorate compounds. AP is primarily for use by the defense industry and aerospace industry, has been used in solid propellants, and is a solid constituent of rocket and missile. Now AP is also widely used as components in fireworks, air bag inflators, and flares, and as additives in many other manufacturing operations, including electroplating, production of paints and tanning and leather finishing. AP is a specific particle generated from workplace and other occupational sites. In the process of production and manufacturing operations, people could suffer the health risk of AP dust through various exposure routes. Some studies have reported that AP can alter the structure of animal thyroid gland and disturb thyroid normal function. Sodium/iodide symporter (NIS) at the basolateral membrane of the thyroid follicles is a membrane glycoprotein that mediates active iodide uptake into the thyroid follicular cells. Studies have demonstrated that AP is a strong inhibitor of NIS. AP can competitively inhibit thyroidal iodide uptake and therefore affect the synthesis and secretion of thyroid hormone. This thyroid status may finally disturb thyroid homeostasis and impair thyroid function.Several thyroid-specific genes and proteins such as NIS,Tg,TPO involved in biosynthesis of thyroid hormones play an important role in thyroid homeostasis. NIS mediates active iodide uptake into the thyroid follicular cells, this is the first step in thyroid hormone biosynthesis. In addition, Tg is a thyroid protein that is the precursor to iodine-containing thyroid hormones and is typically present in the colloid of thyroid gland follicles. Its primary function is to serve as a macromolecular substrate and matrix for the coupling of iodide to its tyrosine residues during thyroid hormone biosynthesis. TPO is the enzyme can oxidize I- into I+, and its role in thyroid hormone biosynthesis is responsible for iodinating tyrosyl groups of Tg and binding these groups to form hormonal molecules T3 and T4. Some studies have demonstrated that AP may not only competitively inhibit thyroidal iodide uptake but also affect biosynthesis of thyroid hormones directly. The mode of action of AP may include regulations of various thyroid-specific genes. However, more details of molecular mechanism of AP toxicity are still unclear, and the current studies are short of modulation of various thyroid-specific genes. Meanwhile, AP is a primary type of occupational hazards in workplace. Occupational group could suffer the health risk of AP dust through various exposure routes such as inhalation and skin exposure. However, in the literature, there have been a limited number of reports of health effects induced by AP, and few preliminary animal studies on the thyroid toxicity of AP. in the present study, based on animal experiment and occupational health surveillance, we aim to explore the effects of AP on thyroid-specific genes expression and thyroid hormone level of workers so as to better understand the mechanism of AP and provide important information to protect workers and to control occupational health risks of AP. Objective:To investigate the effects of AP on body weight gain, organ coefficient, thyroid hormone level, thyroid morphous and its function.Methods:(1) animal treatment:thirty healthy male Sprague-Dawley rats (28-day old, weighing 80-100 g) were purchased from Tongji Medical College Animal Laboratory. The animals were allowed free access to food and water at all times. The rats were randomly divided into six groups, each group containing five rats. Three groups as AP-treated group were given different doses of Ammonium perchlorate (AP) 130 mg/kg b.wt (low),260 mg/kg b.wt (moderate),520 mg/kg b.wt(high) respectively in deionized water every day by oral administration for 90 days. The concentration of AP in deionized water was adjusted weekly for these groups, based on measured body weight, to achieve the desired dosage levels. Other three groups are control group (normal feed), iodine-deficient group (fed by iodine-deficient diet, iodine content:50μg/kg) and high iodine-AP combined group (administered with 520 mg/kg b.wt AP and 10 mg/L iodine solution). (2) Measurement indexes:After 90-day treatment, all rats were sacrificed. The thyroid gland, heart, liver, spleen, lung kidney and testis of rat were removed immediately and weighed and calculated organ coefficient. Blood samples were collected from all animal. Serum was separated via centrifugation of clotted blood sample and stored at -80℃for later thyroid hormone analysis. Histological samples of thyroid gland from rats were used to prepare HE staining in order to observe histological alteration.Results:(1) body weight gain and organ coefficient:comparing with control group,90-day treatment with 130,260 mg/kg b.wt AP did not result in significant change of body weight, but an significant decrease in body weight was observed in high dose AP group (520 mg/kg b.wt) and iodine-deficient group (P<0.05 and P<0.01 respectively). AP can induce the enlargement of thyroid. Significant alteration of the organ coefficient of thyroid in high dose AP group and iodine-deficient group were observed in the present study(P<0.01). In addition, organ coefficient of heart and spleen in high dose AP group were significantly lower than control group(P<0.05). Organ coefficient of testis in high dose AP group and iodine-deficient group were significantly higher than control group(P<0.05). (2) Thyroid hormone level:comparing with control group, FT3 levels of AP-treated groups have a decreasing trend but all failed to reach statistically significant levels. FT4 levels were significantly decreased in moderate dose AP group (260 mg/kg b.wt),high dose AP group (520 mg/kg b.wt) and iodine-deficient group(P<0.05, P<0.01 and P<0.01 respectively). TSH level of high dose AP group (520 mg/kg b.wt) and iodine-deficient group were significantly higher than control group(P<0.05). (3) Histological alteration of thyroid gland:comparing with control group, thyroid gland section of low dose AP group display a slight shrinking follicular, lightly stained colloid and lightly increased follicular epithelial height. Moderate dose AP group display an obvious shrinking follicular, colloid depletion, extensive increased follicular epithelial height and follicular interstitial hyperemia. High dose AP group display a follicular atresia, colloid depletion, interstitial collagen fiber hyperplasia and severe follicular interstitial hyperemia. Iodine-deficient group display an obvious shrinking follicular, colloid depletion and follicular epithelial hyperplasia. High iodine-AP combined group display an enlarged follicular cavity and normal colloid.Conclusions:(1) AP can affect the growth and development of rat and obviously retard the body weight gain of rat. (2) AP can decrease serum FT4 level and feedback to increase TSH level. AP can disturb normal biosynthesis of thyroid hormones. (3) AP can induce enlargement of thyroid gland and alter the structure of thyroid cell. Histological alterations display a follicular epithelial hyperplasia, obvious shrinking follicular, follicular interstitial hyperemia and colloid depletion. Objective:In order to explore the molecular mechanism of AP toxicity on thyroid, the present study investigate the effects of AP on thyroid-specific gene NIS, Tg, TPO and thyroid transcription factor TTF-1, Pax8 and Foxel mRNA expression.Methods:Using real-time quantitative PCR to measure mRNA expression levels of NIS, Tg, TPO, TTF-1, Pax8 and Foxel. The protein levels of NIS, Tg, TPO were determined by western blotting.Results:(1) NIS gene expression levels have an increasing trend in AP-treated groups and iodine-deficient group, but all differences were not statistically significant(P>0.05). Tg gene expression levels were obviously decreased in AP-treated groups. Tg mRNA levels in all AP-treated groups were significantly lower than that of control (P<0.01). Tg gene expression levels in iodine-deficient group and high iodine-AP combined group did not change significantly with comparing to control group(P>0.05). TPO mRNA levels also have a decreasing trend in AP-treated groups and the level was significantly lower in high dose AP group than that in control (P<0.05). No statistically significant alteration of TPO mRNA level in iodine-deficient group and high iodine-AP combined group was observed 1 in this study. TTF-1 mRNA levels in high dose AP group and iodine-deficient group were significantly lower than control group(P<0.05). Foxel mRNA levels in moderate and high dose AP group were significantly higher than control group(P<0.01, P<0.05). Pax8 mRNA levels in all treatment groups did not appear significant change with comparing to control group(P>0.05). (2) NIS protein level in all treatment groups did not appear significant change with comparing to control group(P>0.05). Tg protein level in low, moderate and high dose AP groups have a decreasing trend but only high dose AP group was significantly lower than control group(P<0.05). TPO protein level in low, moderate and high dose AP groups also have a decreasing trend but only high dose AP group was significantly lower than control group(P<0.05).Conclusions:(1) AP can suppress mRNA and protein expression of Tg and TPO. This may indicate that AP can affect biosynthesis process of thyroid hormones directly via disturbing thyroid-specific gene expression. (2) AP suppressing thyroid-specific gene Tg and TPO expression possibly via affecting thyroid transcription factor TTF-1 and Foxel.Objective:To explore effects of AP on thyroid hormone level, the present study perform an occupational field investigation in A, B, C, D four AP factories and aim to provide some useful information to protect workers and to control occupational health risks of AP.Methods:Using FC-2 dust sample collector to collect AP dust and monitor AP dust level in the air of workplace. Workers health examination includes individual health state, personal occupational history, previous medical history, smoking history. Laboratory examination includes blood test (WBC, RBC, Hb), liver and renal function (ALT, BUN, Cr), thyroid hormone level (FT3, FT4, TSH).Results:AP is a primary type of occupational hazards in workplace. The results of AP dust monitoring show that the AP dust concentrations in the air of group batching workplace of factory A, size reduction workplace and mesh screening workplace of factory B, size reduction workplace and mesh screening workplace of factory C are 51.63mg/m3, 61.09mg/m3,55.88mg/m3,55.13mg/m3,26.34mg/m3. These data were all higher than the threshold value of 15mg/mJ from OSHA's PEL-TWA. Respiratory system symptom detection rate of AP dust-exposed group is significantly higher than that of control group (P <0.01). Blood test, liver and renal function in AP dust-exposed group did not appear significant change with comparing to control group(P>0.05). Serum FT3 level in AP dust-exposed group of factory A was significantly lower than that of control group (P< 0.05).Conclusions:(1) AP dust concentrations in the air of some workplace are higher than threshold value of PEL-TWA. AP factory should reinforce the dust-proof measures and enhance the personal protection for workers. (2) Respiratory system symptom detection rate of AP dust-exposed group significantly increased, this indicated that AP dust can impair the respiratory system. (3) AP did not appear obvious effects on blood system, liver and renal function. (4) AP dust exposure can significantly decrease FT3 level in AP dust-exposed group of factory A, this demonstrate that AP in some extent, can affect thyroid function of workers. AP factory should enhance protective measures to protect workers and to control occupational health risks of AP.