Experimental Study Of Tamoxifen In The Treatment Of Differentiated Thyroid Cancer

1.The expression of the estrogrn receptor in thyroid tumour tissuesand the clinical significancesObjective: To investigate the expression of estrogen receptor (ER) inthyroid tissues and explore the clinical significance.Method: 64 cases of thyroid abnornal tissues including 31 thyroidcarcinomas, 8 adenomas, 3 Harshimoto's thyroiditis and 22 thyroidnodules as well as 10 cases of normal thyroid tissues diagnosedpathologically were from the first affiliated hospital of Nanjing MedicalUniversity, from Febrary 2002 through Febrary 2003. The breastcarcinoma tissues were used as the ER positive control. The specimenenbeded with paraffin wax were treated by immunohistochemicalmethod of Avidin-biotin-eroxidase Complex (ABC).Results: 1 of 10 normal thyroid tissues was ER positive. Of 64 cases inthyroid abnormal tissuses, 28(43.8ï¼…) were ER positive, 19 of 39(48.7ï¼…)thyroid tumours were ER positive and 9 of 25 (36.0ï¼…) non-thyroidcarcinomas were ER positive; The incidence of ER positive in thethyroid diseases was more than that in the normal thyroid tissues(P＜0.01). 11 of 22(50ï¼…) thyroid tumour were ER positive and 1 of9(11.1ï¼…) neck metastases were ER positive, which was differentsignificantly. The presence of ER was significantly associated withneither the age nor the sex in this study.Conclusion: The ER expression in thyroid tumours is more than that inthe normal thyroid tissue, which means that the development of thyroidtumours may relate to the ER over-expression and to the estrogen.2.The Proliferation of Papillary Thyroid Carcinoma Cells wasPromoted byβ-estrodiol, which was reversed by TamoxifenObjective: To investigate the effects ofβ-estrodiol on proliferation ofthyroid papillary carcinoma cells, and the mechanisms possiblyinvolved.Methods: The primary cultured papillary thyroid carcinoma cells wereincubated with 0, 10^-11, 10^-10, 10^-9,10^-8, 10^-7 mol/Lβ-estrodiol and 10^-10,10^-9, 10^-8, 10^-7, 10^-6 mol/L TAM for 1, 3, 5, 7d and were harvested. Thecell viabilities were measured by dimethylthiazol-diphenyltetrazoliumbromide (MTT) assay. The cell cycles of the stimulated cells wereobserved by flow cytometer, and The expression of Cyclin D1 mRNAwas analyzed by semi-quantitative RT-PCR.Results:β-estrodiol obviously promoted the proliferation of thyroid carcinoma cells in a dose-dependent and time-dependent patterns(P＜0.05), which was reversed by estrogen antagonist tamoxifen. Afterthe treatment ofβ-estrodiol, the proportion of cells in G₀/G₁ phasesdecreased and cells in S phase increased significantly(P＜0.05). TheCyclin D1 mRNA expression increased under 10^-8 mol/Lβ-estrodiolstimulation, which were inhibited by TAM.Conclusion:β-estrodiol can promote the growth of ER positivepapillary thyroid carcinoma cells by facilitating the G₁-S phase transitionand TAM inhibits the growth of ER positive thyroid cancer cells byarresting the G₁-S phase transition.3. Tamoxifen Inhibits the Growth of ER Negative Thyroidcarcinomas in vitro and in vivoObjectives: To investigate the effects of tamoxifen (TAM) on thegrowth of estrogen receptor (ER) negative thyroid carcinomas and themechanisms involved.Methods: In vitro, papillary thyroid carcinoma cells(IHH-4) andfollicular thyroid carcinoma cells(FTC-133) were treated withtamoxifen in different concentration of 10, 8, 6, 4, 2, 0μmol/L anddifferent lengths of time, using the dimethylthiazol-dipheltetrazoliumbromide (MTT) method to observe the cell proliferation. Cell cycle and apoptosis were analysed with flow cytometric DNA studies aftertreated with 8μmol/L. In vivo, the papillary thyroid carcinoma cells wereinjected into the nude mice subcutaneously, and the mice had been feedwith TAM (5mg/kg/d) daily since the second week for 5 weeks. Thevolumes were calculated weekly. Finally the mice were sacrificed andthe tumors were weighed separately.Results: In vitro, TAM inhibited the proliferation of the IHH-4 cell lineand the FTC-133 cell line in dose-dependent and time-dependentpatterns; TAM inhibited the growth of the IHH-4 and FTC-133 cell linesby blocking the cell cycle in G₁ phase. Immunohistochemistry andRT-PCR studies showed the ER negative in both cell lines. In vivo, TAMalso inhibited the diameter and weight of IHH-4 xenografts in nudemice.Conclusion: TAM inhibits the growth of ER negative thyroid carcinomain vitro and in vivo. TAM acts independently of estrogen receptor andmay do by arresting the cell cycle in the G₁ phase.4. Tamoxifen induced apoptosis of thyroid carcinoma cell lineFTC-133 related to up-regulation of baxObjectives: Tamoxifen (TAM) inhibits the growth of estrogen receptorpositive cells and the growth of the ertrogen receptor negative cells. Theaim of this project is to investigate the effects of tamoxifen (TAM) on the growth of the follicular thyroid carcinoma cell line FTC-133 and themechanisms involved.Methods: The follicular thyroid carcinoma cells FTC-133 wereincubated with 0, 2, 4, 6, 8, 10μmol/L Tamoxifen, the proliferation wasevaluated by dimethylthiazol-dipheltetra- zolium bromide(MTT) methodat 24, 48, 72h. Apoptosis of the thyroid carcinoma cells was detectedwith flow cytometric (FCM) ANNEXIN-V/PI pathway after treaded by8μmol/L Tamoxifen for 24h. The expressions of Bax and Bcl-2 wereobserved with real-time PCR at mRNA level and Western blot at proteinlevel after FTC-133 incubated with 8μmol/L TAM after 4, 8, 16, 24 h.Results- TAM inhibited the proliferation of the FTC-133 cells indose-dependent and time-dependent patterns; TAM induced apoptosis ofFTC-133 cell. The expression of Bax gene was up-regulated andexpression of Bcl-2 gene was no change significantly either mRNA levelor protein level by TAM. There is no expression of estrogen receptor inthe thyroid carcinoma cell line FTC-133 by Western blot and RT-PCRstudies.Conclusion: TAM induces apoptosis of thyroid carcinoma cellsFTC-133, which could related to the up-regulation of the bax.