| Objective: 1. To establish rat model disturbed by low-dose 2,3′,4,4′,5-pentachlorobiphenyl(PCB118) exposure. 2. To investigate the effects of PCB118 on thyroid structure and function. 3. To explore the possible mechanism of PCB118-induced thyroid dysfunction.Methods: 1. Forty adult male Wistar rats(200 ± 10 g) were randomly assigned to one of the four treatment groups comprising ten animals each. PCB118 was dissolved in corn oil. Rats in the three experimental groups were administered PCB118 at 10, 100, or 1000 μg/kg/day i.p. for 5 consecutive days per week for 13 weeks, whereas control rats were injected with corn oil(0.5m L/kg) on the same treatment schedule. Body weights were recorded daily. At the end of treatment, all animals were exsanguinated for serum hormone analysis, Serum was isolated from whole blood and stored at-70 °C for further analysis, and the thyroid glands were removed for histological analyses. 2. Thyroids were removed and fixed overnight in 4% paraformaldehyde, following paraffin-embedded thyroids sections were stained with hematoxylin and eosin(HE) and observed via light microscope. 3. Thyroid specimens were fixed in 5% glutaraldehyde, post-fixed in 1% osmium tetroxide and sectioned on an ultramicrotome for transmission electron microscopy. 4. Serum concentrations of the hormones FT3, FT4, TG and TSH, TGAb, TPOAb were determined by radioimmunoassay, Specific operation reference instruction manual of the corresponding RIA kit. 5. Total RNA was isolated using TRIzol reagent. Quantitative real-time PCR was performed to measure the expression levels of NIS, TG, TPO, AKT and Fox O3 a m RNAs in the rat thyroid.6. After isolate total protein, we detected the change of AKT, p-AKT and p-Fox O3 a expression in the rat thyroid by and western blotting.Result: 1. No sign of toxicity or morbidity was observed among the experimental rats during or following exposure to PCB118. Similarly, no significant change in body weight was observed in PCB118 or vehicle-treated animals. 2. Treatment with PCB118 lead to distinct histopathological changes in the rats thyroid and generally dose-dependent. Compared with the thyroid follicles, colloid and interfollicular areas in control rats, all PCB118-treated rats exhibited hyperplasia and expansion of the follicles, shedding of epithelial cells, deficient luminal colloid, collapsed follicles, mesenchymal fibrosis and interstitial vascular proliferation. Fibrinoid necrosis or even disappearance of the follicular structure was also observed in thyroid sections from the 100 and 1000 μg/kg/d dosage groups. 3. Thyrocytes in vehicle-treated control rats showed an abundance of neatly arranged microvilli on the apical membrane. The morphology of the mitochondria and rough endoplasmic reticulum(RER) was normal and the apical cell areas exhibited numerous secretory vesicles and colloid droplets. In PCB118-treated rats, dose-dependent changes in cellular and organelle ultrastructure were observed. In all PCB-treated groups, the follicular cells had many vacuoles and few microvilli, the perinuclear gap was wider than that in follicular cells from control rats, and secretory vesicles were sparse at the apical pole. Swollen mitochondria and dilated RER cisternae were present in the basal pole. There was a general loss of subcellular organization and of cellular contents. 4. Along with increasing doses of PCB118, serum FT3, FT4, TSH and TG decreased. Significant reductions in FT3 only occurred in 1000 μg/kg/d dosage group(P<0.05). Levels of FT4, TG and TSH were dramatically reduced and generally dose-dependent(P<0.05). It was also found that following with increasing dose of PCB118, the levels of serum TGAb, TPOAb and TPO m RNA in thyroid tissue reached highest concentration in 100 μg/kg/d dosage group, but decreased in 1000 μg/kg/d dosage group.5. Expression levels of NIS and TG m RNAs in the thyroid were significantly lower in the two higher PCB118 dosage groups(P<0.05), but not in the lowest dosage group(P>0.05), compared with the vehicle-treated controls. Expression level of TPO m RNAs in the thyroid were significantly higher in PCB118100 μg/kg/d dosage group(P<0.05), compared with the control group. 6.(1) Expression level of AKT m RNA in the thyroid were significantly higher in the highest PCB118 dosage group(P<0.05), but not in 10-100 μg/kg/d dosage groups(P>0.05), compared with the vehicle-treated controls.There is no significant difference on expression level of Fox O3 a m RNA in the thyroid among all PCB-treated groups and the vehicle-treated controls.(2) Expression levels of AKT protein in the thyroid were significantly higher in the two higher PCB118 dosage groups(P<0.05), but not in the lowest dosage group(P>0.05), compared with the control group. Expression level of p-AKT protein only in the highest dosage group was significantly higher than that in controls(P < 0.05). Expression levels of p-Fox O3 a protein increased progressively with increasing PCB118 dose and were significantly higher in all three dosage groups compared with the control group(P<0.05).Conclusion: 1. Chronic low-dose PCB118 exposure can destroy the structure of the thyroid gland and dramatically induce thyroid dysfunction in a dose-dependent manner. 2. Chronic low-dose PCB118 exposure can initiate the autoimmunity of thyroid. 3. Chronic low-dose PCB118 exposure can reduce the expression of TH biosynthesis-associated gene and generally dose dependent. 4. Chronic low-dose PCB118 exposure can activate PI3K/AKT signal transduction pathway, and then increase the phosphorylation level of Fox O3 a. |
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