维甲酸抑制对不同分化程度甲状腺肿瘤细胞体外侵袭转移能力的影响

Beneficial effects of retinoic acid on in vitro invasiveness of human thyroid carcinoma cell lines

目的 研究维甲酸对人甲状腺肿瘤细胞体外侵袭转移能力的影响,初步探讨其分子机制.方法 对4种人甲状腺肿瘤细胞予一定浓度的维甲酸处理,以Transwell试验验证维甲酸对细胞侵袭能力的影响,黏附试验判断维甲酸作用机制环节,最后以RT-PCR及Western印迹法检测侵袭相关蛋白E-钙黏蛋白（E-cadherin）、基质金属蛋白酶2（MMP2）、尿激酶型纤溶酶原激活因子（uPA）及其受体（uPAR）的基因及蛋白表达变化.结果 维甲酸处理5 d后,4种甲状腺肿瘤细胞体外侵袭能力均低于乙醇对照组,差异有统计学意义,其中FTC-133为91±9比118±10,C643为92±17比164±21,HTH74为87±18比169±15,XTC.UC1为95±23比136±15,差异均有统计学意义（均P〈0.05）,同时发现除FTC133以外的3种肿瘤细胞的黏附能力均较对照组高.其中未分化癌细胞C643、HTH74 uPA和uPAR mRNA及蛋白表达均低于对照组,同时E-cadherin表达较对照组高;滤泡癌细胞FTC-133和Hürthle癌细胞XTC.UC1中,uPAR和MMP2 mRNA及蛋白水平低于乙醇对照组,uPA和E-cadherin表达无显著改变.结论 维甲酸可能通过上调E-cadherin表达,降低uPA、uPAR及MMP2等多种蛋白溶解酶活性,显著抑制人甲状腺肿瘤细胞体外侵袭转移能力.

Objective To investigate the anti metastatic potential of retinoic acid as an important determinant of metastatic behavior in thyroid carcinoma and understand the role of invasion associated proteins.Methods Differentiated thyroid carcinoma cell lines FTC-133 and XTC.UC1,anaplastic thyroid cancer cell lines C643 and HTH74 were studied.All cell lines were cultured with all-trans-RA （ATRA） or solvent ethanol.The in vitro invasion and adhesion potency were studied by transwell experiment and shortterm adhesion assay.The functions of invasion associated proteins,urokinase type plasminogen activator （uPA）,uPA receptor （uPAR）,MMP2 and E-cadherin were investigated by semi-quantitative RT-PCR and Western blot.Results In vitro invasion assay revealed that ATRA treatment could reduce the invasive potency in all the thyroid cancer cell lines.On Day 5 of ATRA treatment,the numbers of cells that migrated through extracellular matrix were as follows,in contrast to control group,FTC-133：91 ±9 vs 118±10,C643：92± 17 vs 164 ±21,HTH74：87±18 vs 169±15,and XTC.UC1：95 ±23 vs 136±15,respectively （all P 〈 0.05）.Short term adhesion assay suggested that ATRA increases cancer cell adhesion to extracellular matrix （ECM） in C643,HTH74 and XTC.UC1.RT-PCR and Western blot both revealed diminished expression of uPAR in all four carcinoma cell lines.In C643 and HTH74 cell lines,the expression of uPA was reduced and the expression of E-Cadherin was increased; whereas the MMP2 expression was not significantly down-regulated in ATRA treated group.In ATRA treated FTC-133 and XTC.UC1 cell lines,MMP2 expression was decreased,but no significant changes in uPA and E-Cadherin expression were observed.Conclusion The present study demonstrates the influence of ATRA on two important determinants of metastatic behavior （＂de adhesion＂ and proteolysis） in thyroid carcinoma cell lines.