RESUMEN
Objective To investigate the expression of marrow-derived thyroid stimulating hormone β (TSHβ) splice variant in thyroid of mouse with autoimmune thyroiditis induced by thyroglobulin(Tg) immunization,and to analyze whether TSHβ splice variant participated in the pathological process of autoimmune thyroiditis.Methods Using random number table,forty-eight mice(24 females and 24 males) of 7 to 8 weeks old with body mass 20 to 25 g were randomly divided into 4 groups(12 females and 12 males in each group) based on body weight and gender.The control group:fed with deionized water; the Tg-treated group(TG):fed with deionized water,and immunized subcutaneously with 0.1 mg Tg at 8 weeks old,boost immunized twice at 11 and 15 weeks old,respectively; the high iodine-treated group (HI):fed with deionized water containing 0.05％ sodium iodide (NaI); the Tg combined with HI group (TG + HI):fed with deionized water containing 0.05％ NaI,and immunized the same way as the TG group did.Peripheral blood was collected after 8 weeks treatment,which was used for determination of total tetraiodothyronine (TT4),free tetraiothyronine (FT4),total triiodothyronine (TT3) and free triiodothyronine (FT3) with chemiluminescence immunoassay (CIA); thyroid glands were collected to examine the expression of TSHβ splice variant with SYBR Green fluorescent quantitative real-time PCR,and frozen sections were HE stained for observation of histopathological changes of thyroid cells under light microscopy.Results Under naked eyes,the thyroid gland enlarged significantly,and looked dark red in HI and TG + HI groups.Under an optical microscope,thyroid follicular epithelial cells presented cuboidal,with abundant cytoplasm,presented abundant glial in follicular cavity,without lymphocyte infiltration in the control group; in TG group,the thyroid follicular epithelial cells presented cuboidal,with some single scattered lymphocytes; in HI group,colloid volume expansion appeared in thyroid follicles,thyroid follicular epithelial cells presented low cuboidal or flat,with few single scattered lymphocytes; in TG + HI group,most colloid accumulative large follicles presented in thyroid,thyroid follicular epithelial cells presented flat,some destructive thyroid follicular structure and infiltrating lymphocytes appeared.The differences of FT3,TT4,FT4 and TSHβ splice variant between groups were statistically significant(F =4.00,12.54,31.92,214.29,all P ＜ 0.05).Compared to the control group,the serum TT3(nmol/L:0.92 ± 0.07 vs.1.30 ± 0.33,t =-2.24),TT4(nmol/L:1.30 ± 0.33 vs.95.60 ± 14.10,t =-3.02),FT4(pmoL/L:54.07 ± 3.67 vs.154.80 ± 0.01,t =-54.87) and the thyroids' TSHβ(× 10-3:4.11±0.32 vs.8.38 ± 0.22,t =-19.11) were higher in TG group(all P ＜ 0.05) ; the serum TT4(nmol/L:67.75 ± 11.91 vs.45.50± 3.85,t =3.55,P ＜ 0.05) was lower in HI group; the serum FT4(pmol/L:54.07 ± 3.67 vs.139.46 ± 30.00,t =-5.65) and the thyroids' TSHβ splice variant (× 10-3:4.11 ± 0.32 vs.5.33 ± 1.47,t =-5.95) were higher in TG + HI group (all P ＜ 0.05).Conclusions High iodine has aggravated thyroiditis of BALB/c mice induced by Tg immunization; the level of thyroid TSHβ in mice with autoimmune thyroiditis is higher; all of these results indicated that TSHβ is involved in the pathogenesis of autoimmune thyroiditis.
RESUMEN
ObjectiveTo find out if the immune system derived thyroid stimulating hormone(TSH) β splice variant(TSHβ-Ⅴ) would be regulated by circulating thyroid hormone levels to get a further understanding of the function and mechanism of this TSHβ-Ⅴ in thyroid homeostasis.MethodsA total of 20 weaning Balb/c mice (half male and half female) were selected and randomly divided into two groups according to their body mass and gender(n =10).Mice of control group were fed with common diet and deionized water.Mice of the low-iodine(LI) group were fed with low-iodine diet(containing iodine 20 - 40 μg/kg,iodine-intake about 0.25 μg/d) and deionized water.The experimental period was 3 months.At the end of the experiment,mice were executed and the blood was collected to observe the levels of TSH and thyroid hormone by chemiluminescence immunoassay (CIA) ; bone marrow (BM),peripheral blood(PBL),thyroid gland and pituitary were collected to assay the TSHβ-Ⅴ mRNA expression by real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR).ResultsThe serum free thyroxine(FT4) and total thyroxine(TT4) levels in LI group of mice[(0.47 ± 0.70)nmol/L,(2.41 ± 0.28)pmol/L] were significantly lower than that of the control group of mice [(55.2 ± 3.68) nmol/L, (32.72 ± 1.02) pmol/L,t =43.81,86.04 、all P ＜ 0.01 ] and the serum total triiodothyronine(TT3) and free triiodothyronine(FT3) reduction in LI group of mice[ (0.76 ± 0.08)nmol/L,(4.01 ± 0.40)pmol/L] were significantly lower than that of the control group of mice [ (1.10 ± 0.06)nmol/L,(5.40 ± 0.38)pmol/L,t =9.81,7.5 1,P ＜ 0.01 ].Iodine insufficiency strongly elevated the serum TSH in LI group of mice[ (35.67 ± 17.39)mU/L] than that in control group of mice[ (0.24 ± 0.10)mU/L,t =- 6.11,P ＜ 0.01 ].The mRNA levels of TSH β-Ⅴ in BM (9.62 ± 0.60) and in PBL( 9.25 ± 0.83 ) of LI group of mice were lower than those in control group of mice (7.69 ± 0.36,7.11 ± 0.41,t =6.77,5.64,P ＜ 0.01),while the mRNA level of TSH β-Ⅴ in pituitary of LI group of mice (1.99 ± 0.61) was increased compared with that in control group of mice (5.75 ± 0.98,t =- 8.02,P＜ 0.01).Compared with control group of mice(9.12 ± 0.62),the level of thyroid TSH β-Ⅴ mRNA in LI group of mice (9.32 ± 0.91 ) was not significantly changed (t =0.45,P ＞ 0.05).There was no detectable native TSHβ in BM,PBL and thyroid.The mRNA level of native TSHβ in pituitary in LI group of mice( - 7.17 ± 1.78) was dramatically elevated compared to that in control group of mice( - 1.43 ± 0.51,t =- 7.60,P ＜ 0.01 ).ConclusionsThe mRNA levels of TSHβ-Ⅴ are suppressed in BM and PBL in low iodinediet induced hypothyroidism mice,which suggest that immune system derived TSHβ-Ⅴ may be more important thannative TSHβ in immune-thyroid regulation.