Time-lapse Imaging of Primary Preneoplastic Mammary Epithelial Cells Derived from Genetically Engineered Mouse Models of Breast Cancer

被引:3
作者
Nakles, Rebecca E. [1 ]
Millman, Sarah L. [1 ]
Cabrera, M. Carla [1 ]
Johnson, Peter [1 ,2 ]
Mueller, Susette [1 ,2 ]
Hoppe, Philipp S. [3 ]
Schroeder, Timm [3 ]
Furth, Priscilla A. [1 ,2 ,4 ,5 ,6 ]
机构
[1] Georgetown Univ, Dept Oncol, Washington, DC 20057 USA
[2] Georgetown Univ, Lombardi Comprehens Canc Ctr, Washington, DC 20057 USA
[3] German Res Ctr Environm Hlth, Helmholtz Zentrum Munchen, Munich, Germany
[4] Georgetown Univ, Dept Med, Washington, DC 20057 USA
[5] Dankook Univ, Dept Nanobiomed Sci, Yongin, South Korea
[6] Dankook Univ, WCU Res Ctr Nanobiomed Sci, Yongin, South Korea
来源
JOVE-JOURNAL OF VISUALIZED EXPERIMENTS | 2013年 / 72期
基金
新加坡国家研究基金会;
关键词
Cancer Biology; Issue; 72; Medicine; Cellular Biology; Molecular Biology; Anatomy; Physiology; Oncology; Mammary Glands; Animal; Epithelial Cells; Mice; Genetically Modified; Primary Cell Culture; Time-Lapse Imaging; Early Detection of Cancer; Models; Genetic; primary cell culture; preneoplastic mammary epithelial cells; genetically engineered mice; time-lapse imaging; BRCA1; animal model; GENERATION; BRCA1;
D O I
10.3791/50198
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
Time-lapse imaging can be used to compare behavior of cultured primary preneoplastic mammary epithelial cells derived from different genetically engineered mouse models of breast cancer. For example, time between cell divisions (cell lifetimes), apoptotic cell numbers, evolution of morphological changes, and mechanism of colony formation can be quantified and compared in cells carrying specific genetic lesions. Primary mammary epithelial cell cultures are generated from mammary glands without palpable tumor. Glands are carefully resected with clear separation from adjacent muscle, lymph nodes are removed, and single-cell suspensions of enriched mammary epithelial cells are generated by mincing mammary tissue followed by enzymatic dissociation and filtration. Single-cell suspensions are plated and placed directly under a microscope within an incubator chamber for live-cell imaging. Sixteen 650 mu m x 700 mu m fields in a 4x4 configuration from each well of a 6-well plate are imaged every 15 min for 5 days. Time-lapse images are examined directly to measure cellular behaviors that can include mechanism and frequency of cell colony formation within the first 24 hr of plating the cells (aggregation versus cell proliferation), incidence of apoptosis, and phasing of morphological changes. Single-cell tracking is used to generate cell fate maps for measurement of individual cell lifetimes and investigation of cell division patterns. Quantitative data are statistically analyzed to assess for significant differences in behavior correlated with specific genetic lesions.
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页数:10
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