Extraction and identification of exosomes from drug-resistant breast cancer cells and their potential role in cell-to-cell drug-resistance transfer.
- Author:
Jinjin XU
1
;
Wenjing LI
;
Shanliang ZHONG
;
Xiujuan LI
;
Zhiyuan CHEN
;
Qing HU
;
Jinhai TANG
;
Jianhua ZHAO
2
Author Information
- Publication Type:Journal Article
- MeSH: Antibiotics, Antineoplastic; pharmacology; Antineoplastic Agents; pharmacology; Cell Survival; Coculture Techniques; DNA-Binding Proteins; metabolism; Doxorubicin; pharmacology; Drug Resistance, Neoplasm; Endosomal Sorting Complexes Required for Transport; metabolism; Exosomes; metabolism; pathology; Green Fluorescent Proteins; metabolism; Humans; MCF-7 Cells; pathology; Taxoids; pharmacology; Transcription Factors; metabolism
- From: Chinese Journal of Oncology 2014;36(3):165-170
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo explore whether docetaxel-resistant cells (MCF-7/Doc) and doxorubicin-resistant cells (MCF-7/ADM) can secrete Exosomes and their potential role in cell-cell drug-resistance transfer.
METHODSExosomes were extracted from the cell culture supernatants of MCF-7/Doc and MCF-7/ADM cells by fractionation ultracentrifugation, and were identified by transmission electron microscopy and Western blot analysis. GFP-MCF-7/S, a breast cancer parental sensitive cell line stably expressing green fluorescent protein (GFP), was constructed by recombinant lentiviral vector with GFP. Then the resistance experiment of cells and the experiment of resistance transfer by exosomes were designed to observe the phenomenon of cell-to-cell drug-resistance transfer.
RESULTSSimilar to the breast cancer parental sensitive cells (MCF-7/S), the breast cancer resistant sublines could secrete exosomes, which exhibited round or elliptic shape ranging from 30 to 100 nm in diameter with intact membrane, and only expressed the protein marker of exosomes, Tsg101, did not express the endoplasmic reticulum marker calnexin. After MCF-7/S, MCF-7/DOC and MCF-7/ADM cells we cocultured with GFP-MCF-7/S cells for 72 h, there were no significant differences in the expression of fluorescence-labeled cells among the four groups. When treated by the drug ADM or DOC for 24 hours, the MCF-7/DOC+GFP-MCF-7/S group was in favor of a significant higher survival rate of fluorescence-labeled cells compared with the MCF-7/S+GFP-MCF-7/S group (65.5% vs. 25.5%, P < 0.001), and so did the MCF-7/ADM+GFP-MCF-7/S group (53.6% vs. 25.4%, P < 0.001). The exosomes extracted from MCF-7/S, MCF-7/DOC and MCF-7/ADM cells were cultured with the GFP-MCF-7/S cells for 48 h. Among these groups, no significant differences in the expression of fluorescence-labeled cells were found. After treated by the drug ADM or DOC for 24 hours, the exosomes extracted from MCF-7/DOC+GFP-MCF-7/S group was associated with a significant higher survival rate of fluorescence-labeled cells compared with the exosomes extracted from MCF-7/S+GFP-MCF-7/S group (59.9% vs. 32.4%, P < 0.001), and so did the exosomes extracted from the MCF-7/ADM)+GFP-MCF-7/S group (58.3% vs. 27.2%, P < 0.001).
CONCLUSIONOur results suggest that drug-resistance can be transferred between breast cancer cells, and exosomes are probably the transporter of the drug resistance.
