Immortalization protocols used in cell culture models of human breast morphogenesis

Abstract

Defining the key players in normal breast differentiation is instrumental to understanding how morphogenesis becomes defective during breast cancer progression. During the past 2 decades much effort has been devoted to the development of technologies for purification and expansion of primary human breast cells in culture and optimizing a relevant microenvironment, which may help to define the niche that regulates breast differentiation and morphogenesis. In contrast to the general property of cancer, normal human cells have a finite lifespan. After a defined number of population doublings, normal cells enter an irreversible proliferation-arrested state referred to as replicative senescence. To overcome this obstacle for continuous long-term studies, replicative senescence can be bypassed by treatment of cells with chemical agents such as benzopyrene, by radiation or by transfection with viral oncogenes or the gene for human telomerase (human telomerase reverse transcriptase, hTERT). A drawback of some of these protocols is a concurrent introduction of chromosomal changes, which sometimes leads to a transformed phenotype and selection of a subpopulation, which may not be representative of the tissue of origin. In recent years, we have sought to establish immortalized primary breast cells, which retain crucial characteristics of their original in situ tissue pattern. This review discusses various approaches to immortalization of breast-derived epithelial and stromal cells and the application of such cell lines for studies on human breast morphogenesis.

Mammary Transplantation of Stromal Cells and Carcinoma Cells in C57BL/6J Mice

“…Expression of specific oncogenes and tumor suppressors can be modified in stromal cells or epithelial cells through stable expression of siRNAs for example, prior to transplantation. Because primary cells senesce with continual passaging, genetic manipulation of cultured cells is easier if the cells are immortalized, for example through spontaneous immortalization or expression of an oncogene 12,13,14,15 . Immortalization of different stromal cell types has been achieved with macrophages and endothelial cells 16,17,18 .…”

Prostate cell cultures as in vitro models for the study of normal stem cells and cancer stem cells

“…In fact, it has been reported that some human mammary epithelial immortalized cell lines exhibited stem-cell-like characteristics and could be used as models of normal breast morphogenesis. 53,54 Although it has been suggested that the study of CSCs should be performed using primary cancer cells rather than cancer cell lines, 55 recent evidence using SP analysis 43,44,56 and cell surface marker, such as CD133, [57][58][59] or clonal analysis, 60,61 shows that long-term cultured cell lines may retain a hierarchical proliferation or differentiation pattern as a potential of CSCs.…”

Selection for EGFR gene amplification in a breast epithelial cell line with basal-like phenotype and hereditary background

“…Over the years, highly useful cell culture models have been developed to answer important questions in breast cancer biology, such as cancer progression as well as the general biology of the normal breast (reviewed in Gudjonsson et al 2004;Lacroix and Leclercq 2004). When cultured in reconstituted basement membrane (rBM) matrix, normal luminal epithelial cells generate polarized acini mimicking epithelial cells in situ.…”