摘要
目的研究氯化钴(CoCl2)诱导的低氧对甲状腺乳头状癌NPA细胞上皮间质转化的影响,初步探讨HIF-1α在其中的机制。方法 MTT检测CoCl2(50、100、150、200、250μmol/L)对细胞活力的影响,倒置显微镜观察低氧(150μmol/L的CoCl2分别作用0、12、24、48 h)对细胞形态的影响,Western blot检测低氧(150μmol/L的CoCl2分别作用0、3、6、12、24 h)对HIF-1α、E-cadherin和Vimentin表达的影响,Transwell法检测低氧(150μmol/L的CoCl2分别作用0、6 h)对细胞侵袭、转移的影响,用免疫荧光对HIF-1α蛋白进行亚细胞定位。结果低氧模拟剂CoCl2可抑制NPA细胞活力。随着CoCl2低氧处理时间的延长,NPA细胞逐渐获得间质细胞的形态特征,HIF-1α、Vimentin表达在0～6 h逐渐增加,随后开始下降,E-cadherin表达持续降低,低氧0 h与6 h相比各蛋白表达均具有统计学差异(P<0.01)。低氧组侵袭、转移细胞数目较对照组明显增加(P<0.01),同时低氧组细胞HIF-1α发生核转位。结论 CoCl2模拟化学性低氧可促进NPA细胞上皮间质转化及侵袭、转移,其分子机制可能涉及HIF-1α介导的信号通路对E-cadherin、Vimentin基因表达的调控。
Objective To determine the effect of CoCl2 induced hypoxia on the epithelial-mesenchymal transition in human papillary thyroid carcinoma and investigate the role of HIF-1a in this process. Methods Cell vitality was evaluated by MTF assay in human papillary thyroid carcinoma NPA cells after the treatment of 50, 100, 150, 200 and 250 μmol/L COCl2. Morphological changes of NPA cells were observed by inverted microscopy after 150 μmol/L CoCl2 treatment for 0, 12, 24 and 48 h. The expression levels of HIF-1a, E-cadherin and Vimentin were analyzed by Western blotting in NPA cells after 150 μmoL/L CoCl2 treatment for O, 3, 6, 12, and 24 h. The invasion and metastasis potential in NPA ceils after 150 μmol/L CoCl2treatment for 0 and 6 h was detected by Transwell assay. Immunofluorescence was used to detect the location of HIF-1a in NPA cells. Results CoCl2 treatment resulted in suppression of NPA cell vitality in a dose-dependent manner. The longer CoCl2wastreated, NPA cells gradually acquired the morphological characteristics of mesenehymal cells. The expression of HIF-1a, E-cadherin and Vimentin was gradually enhanced in 0 to 6 h after treatment, and then declined, especially in E-cadherin. Significant differences were found in the expression levels of these proteins at 0 h and 6 h （ P 〈 0. 01 ）. There were more cells that penetrated through the Transwell membrane in hypoxia group than in normoxic group （P 〈 0. 01 ）. HIF-1a translocated to the nucleus in hypoxia group, while it didn＇t occur in normoxia group. Conclusion CoCl2 treatment, which mimics chemical hypoxia improves the epithelial-mesenchymal transition, invasion and migration of NPA cells, which might be due to that HIF-1a mediated signal pathway regulates E-cadherin and Vimentin.
出处
《第三军医大学学报》
CAS
CSCD
北大核心
2012年第2期150-153,共4页
Journal of Third Military Medical University
基金
国家自然科学基金面上项目(81072183)
重庆市自然科学基金重点项目(CSTC,2011BA5038)
重庆医科大学重点科研项目(XBZD201002)~~
关键词
低氧诱导因子-1Α
上皮间质转化
甲状腺乳头状癌
hypoxia-inducible factor-1a
epithelial-mesenchymal transition
papillary thyroid carcinoma
作者简介
[通信作者]刘智敏，E-mail：liuzm9999@yahoo．com．cn
