Docetaxel-induced human breast ccarcinoma drug-resistant BT549/Docetaxel stem cell isolation and adenosine triphosphate binding cassette transporter G2 gene expression level

Abstract

Objective To isolate docetaxel-induced human breast carcinoma drug-resistant stem cells MCF7/Docetaxel and study its biological characteristics. Methods By suspension culture combined with docetaxel resistant cell line sorting method, BT549/Docetaxel stem cells were isolated, and BT549, and BT549/Docetaxel stem cells were enriched by suspension culture method. The cells were labeled with CD44 and CD24, and detected by flow cytometry. Reverse transcriptase-polymerase chain reaction(RT-PCR) was used to detect the expression of adenosine triphosphate binding cassette transporter G2(ABCG2)in BT549 and BT549/Docetaxel stem cells. Results The population doubling time of BT549/Docetaxel and BT549 stem cells was(39.22±0.65)and(29.20 ±0.50) h with the difference being statistically significant(P< 0.05). Flow cytometric analysis showed CD44+/CD24-/low ratio in BT549 microspheres cells was(13.8±0.5)%, higher than that of BT549 cells in monolayer culture[(2.8±0.6)%, P<0.01].CD44+/CD24-/low ratio in BT549/Docetaxel microspheres cells was(19.1±0.3)%, higher than that in the monolayer culture[(5.0± 0.4)%, P< 0.01].CD44+/CD24-/low ratio in BT549/Docetaxel microspheres cells was significantly higher than that in BT549 microspheres cells(P<0.05). The drug content in BT549 stem cells was (46.9±0.5) ng/106 cells, and that in BT549/Docetaxel stem cells was(7.1±0.3) ng/106 cells (P< 0.01). After culture with normal medium for 4 h, the drug content in BT549 cells was(28.3± 0.6)ng/106 cells, and that in BT549/Docetaxel cells was(3.5±0.1)ng/106 cells(P< 0.01). The expression ABCG2 m RNA in BT549/Docetaxel stem cells was significantly higher than that in BT549 stem cells(P< 0.01). Conclusion The BT549/Docetaxel stem cells were successfully isolated with stable growth and drug resistance. ABCG2 gene expression in BT549/Docetaxel stem cells was significantly increased. Key words: Docetaxel; Drug resistance; Breast cancer stem cell