The long-term objective of my program has been to develop and characterize a human mammary epithelial cell (HMEC) system for use in a wide variety of studies on human cell biology and carcinogenesis. The goal of our laboratory has been to understand the normal growth control processes in HMEC, and to determine how these processes may be altered as a result of immortal and malignant transformation. This work has been guided by the desire to facilitate widespread use of human epithelial cells for molecular and cellular biology studies. Therefore, the HMEC system is relatively easy to use, can provide large quantities of standardized cell populations, and is relatively well-characterized.
To address this goal, our work since 1976 has provided an HMEC culture system that has:
a) long-term active growth of finite lifespan HMEC;
b) immortally transformed lines derived from normal HMEC exposed to chemical carcinogens, oncogenes, p53 inactivation and/or hTERT;
c) malignant transformants of the cell lines following exposure to specific oncogenes.
Detailed information on the derivation, characterization, and methods for growth of these cells, as well as information on how other labs may obtain these cells, can be found on my web site: http://www.lbl.gov/~mrgs/mindex.html
Our laboratory’s long-term emphasis on extensive development and characterization of one human epithelial cell type has enabled us to gain a unique overview of human cellular growth, aging, and transformation. Our studies have now led us to produce a new model of the senescence barriers encountered by cultured normal finite lifespan HMEC as they grow, senesce, overcome senescence barriers, and gain immortality (Garbe et al. 2007). These ongoing studies have indicated that normal cultured HMEC cease proliferation due to a stress-associated senescence barrier (stasis), rather than telomere attrition. Normal HMEC prior to this barrier (pre-stasis) display significant biological differences compared to the finite post-stasis HMEC that are commonly used as “normal”, but have overcome the stasis barrier due to silencing of the cyclin-dependent kinase inhibitor p16INK4a, consequent to growth in stress-inducing serum-free media. In turn, both pre- and post-stasis finite HMEC show many significant differences compared to the non-malignant immortally transformed lines. Although non-malignant, such immortal lines show greater resemblance to tumor-derived immortal lines than finite cultures, and are likely most reflective of HMEC in vivo at the stage of DCIS.
Based on this new model, we have recently improved culture conditions for growth of normal pre-stasis HMEC. Active growth has been achieved for ~60 population doublings using simple culture conditions. Additionally, the growing populations display phenotypes reflective of the multiple types of normal HMEC in vivo, e.g., luminal, myoepithelial, and progenitor. These HMEC cultures can now be used to better understand normal HMEC biology, and the factors that may promote or inhibit early stage carcinogenesis.
A summary of our most work can be found at our web site: http://www.lbl.gov/~mrgs/mindex.html
Garbe, J, Holst, CR, Bassett, E, Tlsty, T, Stampfer, MR, Inactivation of p53 function in cultured human mammary epithelial cells turns the telomere-length dependent senescence barrier from agonescence into crisis, Cell Cycle 6: 1927-1936, 2007.
Li Y, Pan J, Li J-L, Lee J-H, Tunkey C, Saraf K, Garbe J, Jelinsky S, Stampfer MR, Haney, SA, Transcriptional changes associated with breast cancer occur as normal human mammary epithelial cells overcome senescence barriers and become immortalized, Mol Can 6: 7-24 2007.
Stampfer, M, Garbe, J, Nijjar, T, Wigington, D, Swisshelm, K, Yaswen, P, Loss of p53 function accelerates acquisition of telomerase activity in indefinite lifespan human mammary epithelial cell lines, Oncogene 22: 5238-5251, 2003.
Olsen CL, Gardie, B, Yaswen, P, Stampfer, MR, Raf-1-induced growth arrest in human mammary epithelial cells is p16-independent and is overcome in immortal cells during conversion, Oncogene 21: 6328-6339 2002.
Romanov, SR, Kozakiewicz, K, Holst, CR, Stampfer, MR, Haupt, LM, Tlsty, TD, Normal human mammary epithelial cells spontaneously escape senescence and acquire genomic changes, Nature 409: 633-637, 2001.
Stampfer, MR, Garbe, J, Levine, G, Lichtsteiner, S, Vasserot, AP, Yaswen, P, hTERT expression can induce resistance to TGFß growth inhibition in p16INK4A(-) human mammary epithelial cells, Proc Natl Acad Sci (USA) 98: 4498-4503, 2001.
Brenner, AJ, Stampfer, MR, Aldaz, M, Increased p16 expression with first senescence arrest in human mammary epithelial cells and extended growth capacity with inactivation, Oncogene 17: 199-205, 1998.
Stampfer, MR, Bodnar, A, Garbe, J, Wong, M, Pan, A, Villeponteau, B, Yaswen, P, Gradual phenotypic conversion associated with immortalization of cultured human mammary epithelial cells, Mol Biol Cell 8: 2391-2405, 1997.
Stampfer, MR, Yaswen, P, Alhadeff, M, Hosoda, J, TGFß induction of extracellular matrix associated proteins in normal and transformed human mammary epithelial cells in culture is independent of growth effects, J Cellular Physiology 155: 210-221, 1993.
Stampfer, MR, Pan, C-H, Hosoda, J, Bartholomew, J, Mendelsohn, J, and Yaswen, P, Blockage of EGF receptor signal transduction causes reversible arrest of normal and immortal human mammary epithelial cells with synchronous reentry into the cell cycle, Exp Cell Res 208: 175-188, 1993.