[Screening of a sub-clone of human breast cancer cells with high metastasis potential].
OBJECTIVE
To screen a sub-clone of human breast cancer cell of the MCF-7 line with high metastasis potential.


METHODS
Human breast cancer cells of the MCF-7 line were injected subcutaneously into 10 severe combined immunodeficiency (SCID) mice. Sixty-eight days after the mice were killed and their lungs were taken out. Primary cell culture was conducted. When the cells were passed on to the third generation a sub-clone was screened from the lung tissue and termed LM-MCF-7. Microscopy was performed on the lung tissues. The growth curve was drawn. Flow cytometry was used to examine the cell cycle. Chromosome analysis was done. Immunohistochemistry was used to detect the expression of breast cancer specific antigen CAI5-3. Western blotting was used to detect the protein expression of the protein associated with tumor metastasis: nm23 (a metastasis-suppressing gene), myosin light chain kinase (MLCK, a kinase related to cell movement), survivin, bcl-2 and p27 (a gene related to cell cycle). LM-MCF-7 cells were injected into other SCID mice and these mice were killed 30 days later to observe the metastasis of cancer so as to detect the tumorigenic ability of the LM-MCF-7 cells.


RESULTS
When the cells from the mouse lung tissues were passed on to the third generation a sub-clone with high metastasis potential was screened and termed LM-MCF-7. The morphology of the new cell line was typically epithelioid. Flow cytometry showed that the DNA relatively contents were 53.40% of the LM-MCF-7 cells were in the G(0)/G(1) phase, a lower percentage than that of the MCF-7 cells, and 17.10% in the S phase and 23.20% in the G(2+)M phase, both percentages higher than those of the MCF-7 cells. The proliferating time of the LM-MCF-7 cell population was about 20 +/- 14 hours, much shorter than that of the parent strain cells. The chromosomes of the LM-MCF-7 cells, numbering 16-123, showed the morphology characteristic c of human chromosomes. The marker of human breast cancer CA15-3 was detected in both MCF-7 and LM-MCF-7 cells. The protein expression of nm23 and p27 was down-regulated, but the protein expression of MLCK, bcl-2 and survivin was up-regulated in LM-MCF-7 cells in comparison with those in MCF-7 cells. The tumorigenesis rate of LM-MCF-7 cells was 100% (5/5), with a latent period of 5.0 +/- 0.0 d, and the tumor metastasized to lung, kidney, spleen, bone marrow, lymph node and heart.


CONCLUSIONS
A human breast cancer line, LM-MCF-7 cell line, with high metastasis potential has been derived from the human breast cancer cells of MCF-7 line.
Ye
Li-hong
LH
College of Life Sciences, Nankai University, Tianjin 300071, China. yelihong@nankai.edu.cn
Wu
Lian-ying
LY
Guo
Wei
W
Ma
Hong-tao
HT
Zhang
Xiao-dong
XD
D000818:Animals; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000378:metabolism / Q000473:pathology*; D045744:Cell Line, Tumor; D002999:Clone Cells / Q000378:metabolism / Q000473:pathology; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000378:metabolism; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D008325:Mammary Neoplasms, Experimental / Q000378:metabolism / Q000473:pathology; D051379:Mice; D008819:Mice, Nude; D016513:Mice, SCID; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism; D054778:NM23 Nucleoside Diphosphate Kinases / Q000378:metabolism; D009362:Neoplasm Metastasis / Q000473:pathology*; D009363:Neoplasm Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D014183:Transplantation, Heterologous; D025521:Tumor Suppressor Proteins / Q000378:metabolism
D054778:NM23 Nucleoside Diphosphate Kinases; D009363:Neoplasm Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D025521:Tumor Suppressor Proteins; D050760:Cyclin-Dependent Kinase Inhibitor p27; D009219:Myosin-Light-Chain Kinase