Research article

Immortalized, premalignant epithelial cell populations contain long-lived, label-retaining cells that asymmetrically divide and retain their template DNA

Karen M Bussard¹, Corinne A Boulanger¹, Frances S Kittrell², Fariba Behbod³, Daniel Medina² and Gilbert H Smith^1*

• * Corresponding author: Gilbert H Smith gs4d@nih.gov

Author Affiliations

¹ Mammary Biology and Tumorigenesis Laboratory, National Cancer Institute, National Institutes of Health, Building 37, Room 1112, 37 Convent Drive, Bethesda, MD 20892, USA

² Department of Cell Biology, Baylor College of Medicine, One Baylor Plaza, Houston, TX 77030, USA

³ Department of Pathology and Laboratory Medicine, University of Kansas Medical Center, 3901 Rainbow Blvd, Lied G015, Kansas City, KS 66160, USA

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Breast Cancer Research 2010, 12:R86 doi:10.1186/bcr2754

See related Editorial by Zeps & Hemmings, http://breast-cancer-research.com/content/13/1/101

Published: 21 October 2010

Abstract

Introduction

During selective segregation of DNA, a cell asymmetrically divides and retains its template DNA. Asymmetric division yields daughter cells whose genome reflects that of the parents, simultaneously protecting the parental cell from genetic errors that may occur during DNA replication. We hypothesized that long-lived epithelial cells are present in immortal, premalignant cell populations, undergo asymmetric division, retain their template DNA strands, and cycle both during allometric growth and during pregnancy.

Methods

The glands of 3-week-old immune-competent Balb/C female mice were used intact or cleared of host epithelium and implanted with ductal-limited, lobule-limited, or alveolar-ductal progenitor cells derived from COMMA-D1 pre-malignant epithelial cells. 5-Bromo-2-deoxyuridine (5-BrdU) was administered to identify those cells that retain their template DNA. Nulliparous mice were then either injected with [³H]-thymidine (³H-TdR) to distinguish 5-BrdU label-retaining cells that enter the cell cycle and euthanized, or mated, injected with ³H-TdR, and euthanized at various days after coitus. Sections were stained for estrogen receptor-α (ER-α) or progesterone receptor (PR) with immunohistochemistry. Cells labeled with both 5-BrdU and ³H-TdR were indicative of label-retaining epithelial cells (LRECs).

Results

Cells that retained a 5-BrdU label and cells labeled with [³H]-thymidine were found in all mice and were typically detected along the branching epithelium of mature mouse mammary glands. Cells containing double-labeled nuclei (LRECs) were found in the intact mammary glands of both pregnant and nulliparous mice, and in mammary glands implanted with premalignant cells. Double-labeled cells (³H-TdR/5-BrdU) represent a small portion of cells in the mammary gland that cycle and retain their template DNA (5-BrdU). Some label-retaining cells were also ER-α or PR positive. LRECs distributed their second label (³H-TdR) to daughter cells, and this effect persisted during pregnancy. LRECs, and small focal hyperplasia, were found in all immortalized premalignant mammary-implant groups.

Conclusions

The results indicate that a subpopulation of long-lived, label-retaining epithelial cells (LRECs) is present in immortal premalignant cell populations. These LRECs persist during pregnancy, retain their original DNA, and a small percentage express ER-α and PR. We speculate that LRECs in premalignant hyperplasia represent the long-lived (memory) cells that maintain these populations indefinitely.