EMT-related protein expression in polyploid giant cancer cells and their daughter cells with different passages after triptolide treatment
Our previous work has demonstrated that paclitaxel can induce the formation of polyploid giant cancer cells (PGCCs) and inhibit tumor growth by reprogramming ovarian cancer epithelial cells to a benign fibroblastic state via epithelial–mesenchymal transition. Here, triptolide (TPL) was used to treat the breast and ovarian cancer lines. The morphologic characteristics and EMT-related protein expression were studied in different generation of cancer cells after TPL treatment. When BT-549 and HEY cells reached 80–90% confluence, TPL was added to BT-549 for 48 h and HEY for 9 h at a concentration of 40 ng/ml. Scattered PGCCs survived from TPL treatment and generated daughter cells, and then were cultured in medium without TPL for at least ten generation. Western blot analysis and immunocytochemical staining were performed to detect the expression levels and subcellular location of EMT-related proteins in control cells and different generation of TPL-induced PGCCs with daughter cells. Furthermore, wound-healing, transwell, cell counting kit-8, and MTT assay were used to compare the alternation of migration, invasion, and proliferation among control cells and different generation of TPL-induced PGCCs with daughter cells. Scattered PGCCs survived from the treatment of TPL and produced small-sized daughter cells 20–30 days after treatment. Compared to the control cells, the first generation of TPL-induced PGCCs with their daughter cells differentially expressed EMT-related proteins including fibronectin, E-cadherin, vimentin, and Twist, and had lower migration, invasion, and proliferation abilities. The abilities of migration, invasion, and proliferation of TPL-induced PGCCs with their daughter cells gradually enhanced as the passages increasing, and markedly exceeded the control cells in the tenth generation. TPL-induced PGCCs with their daughter cells gradually obtain the abilities of invasion and metastasis in vitro as the number of passage increasing, which can be used to mimick the cancer cells subjected to anti-cancer drugs in vivo and may provide some new insights to explore the mechanism of cancer invasion, metastasis and relapse after chemotherapy.
KeywordsTriptolide Polyploid giant cancer cells Epithelial–mesenchymal transition
SZ designed the study; collected, analyzed, and interpreted data; contributed to manuscript writing; and approved the manuscript before submission. XW, FF and CL collected and analyzed data and approved the manuscript before submission. JD and KL collected, analyzed, and interpreted data, contributed to manuscript writing, and approved the manuscript before submission. YL and MZ collected data, gave constructive comments on the manuscript, revised the paper and approved the manuscript before submission.
This work was supported in part by Grants from the National Natural Science Foundation of China (81672426), and the foundation of committee on science and technology of Tianjin (17ZXMFSY00120 and 17YFZCSY00700).
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Conflict of interest
The authors declare that they have no conflict of interest.
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