[前列腺癌的上皮-间质转化：癌症干细胞还是整体癌细胞]

[Epithelial-mesenchymal transition of prostate cancer: cancer stem cells or bulk cancer cells].

作者信息

Luo Yong, Cui Xin-hao, Jiang Yong-guang, He Da-lin, Zhao Jia-hui, Zhao Lin, Chen Ya-tong, Li Ming-chuan, Lin Yun-hua

机构信息

Department of Urology, Affiliated Beijing Anzhen Hospital, Capital Medical University, Beijing 100029, China.

出版信息

Zhonghua Yi Xue Za Zhi. 2013 Jan 22;93(4):256-60.

PMID:23578503

Abstract

OBJECTIVE

To test the hypothesis that epithelial-mesenchymal transition (EMT) of prostate cancer is most likely to occur in cancer stem cells (CSC).

METHODS

The isolation of CSC from LNCaP cell line was performed by flow cytometry based on side-population (SP) phenotype. After SP sorting, LNCaP/SP and LNCaP/NSP were used for further transfection of hypoxia-inducible factor-1α (HIF-1α). Subsequently, EMT-associated proteins were detected by Western blotting. And the assays of Transwell and methyl thiazolyl tetrazolium (MTT) were used to compare invasive and proliferative potency between LNCaP/SP and LNCaP/NSP after HIF-1α induction. Eventually, xenograft experiments were performed with LNCaP/HIF-1α/SP and LNCaP/HIF-1α/NSP cells for further analysis of in vivo tumorigenesis and distant metastasis.

RESULTS

Through HIF-1α-induced EMT, LNCaP/HIF-1α/SP exhibited such remarkable EMT characteristics as a positive expression of epithelial markers (E-cadherin and CK18) and a negative expression of mesenchymal markers (vimentin, N-cadherin, fibronectin, cathepsin D, MMP-2 and uPAR). And LNCaP/HIF-1α/NSP underwent partial EMT with an abnormal expression of some mesenchymal proteins (vimentin and cathepsin D) and loss of epithelial protein (CK18) despite reservation of another important epithelial marker (E-cadherin). Further Transwell and MTT assays indicated that LNCaP/HIF-1α/SP exhibited stronger in vitro invasive and proliferative potency than LNCaP/HIF-1α/NSP cells. In animal models, the volume of subcutaneous tumor by LNCaP/HIF-1α/SP cells was much greater than that by LNCaP/HIF-1α/NSP counterparts ((1008 ± 230) vs (288 ± 145) mm(3), P < 0.01). Moreover, LNCaP/HIF-1α/SP cells also had a significantly higher rate of subcutaneous tumor incidence (80% vs 53%, P < 0.05) and bone metastasis (40% vs 0, P < 0.01) as compared with LNCaP/HIF-1α/NSP counterparts.

CONCLUSION

As the main target cells of prostatic EMT, CSCs may develop a more malignant phenotype after EMT.

摘要

目的

验证前列腺癌上皮-间质转化（EMT）最有可能发生于癌干细胞（CSC）这一假说。

方法

基于侧群（SP）表型，通过流式细胞术从LNCaP细胞系中分离CSC。SP分选后，LNCaP/SP和LNCaP/NSP用于进一步转染缺氧诱导因子-1α（HIF-1α）。随后，通过蛋白质免疫印迹法检测EMT相关蛋白。采用Transwell和甲基噻唑基四氮唑（MTT）试验比较HIF-1α诱导后LNCaP/SP和LNCaP/NSP之间的侵袭和增殖能力。最后，用LNCaP/HIF-1α/SP和LNCaP/HIF-1α/NSP细胞进行异种移植实验，进一步分析体内肿瘤发生和远处转移情况。

结果

通过HIF-1α诱导的EMT，LNCaP/HIF-1α/SP表现出显著的EMT特征，如上皮标志物（E-钙黏蛋白和细胞角蛋白18）阳性表达，间充质标志物（波形蛋白、N-钙黏蛋白、纤连蛋白、组织蛋白酶D、基质金属蛋白酶-2和尿激酶型纤溶酶原激活物受体）阴性表达。LNCaP/HIF-1α/NSP发生部分EMT，尽管保留了另一种重要的上皮标志物（E-钙黏蛋白），但一些间充质蛋白（波形蛋白和组织蛋白酶D）表达异常，上皮蛋白（细胞角蛋白18）丢失。进一步的Transwell和MTT试验表明，LNCaP/HIF-1α/SP比LNCaP/HIF-1α/NSP细胞表现出更强的体外侵袭和增殖能力。在动物模型中，LNCaP/HIF-1α/SP细胞形成的皮下肿瘤体积远大于LNCaP/HIF-1α/NSP细胞形成的皮下肿瘤体积（(1008 ± 230) vs (288 ± 145) mm(3)，P < 0.01）。此外，与LNCaP/HIF-1α/NSP细胞相比，LNCaP/HIF-1α/SP细胞皮下肿瘤发生率（80% vs 53%，P < 0.05）和骨转移率（40% vs 0，P < 0.01）也显著更高。