RESUMO
INTRODUCTION: Although estrogen and progesterone play a key role in normal mammary development and in breast cancer, the potential for proliferation and lineage differentiation as well as origin of cells that express the estrogen receptor (ER) in normal breast epithelium are not known. Some evidence suggests that normal human mammary stem/progenitor cells are ER-, but the identity of these cells and the cellular hierarchy of breast epithelium are still subjects of controversy. It is likely that elucidation of these aspects will bring insight into the cellular origin of breast cancer subtypes. METHODS: We used fluorescence-activated cell sorting of primary human mammary epithelial cells along with in vitro and in vivo functional assays to examine the hierarchic relation between cells with aldehyde dehydrogenase enzymatic activity (ALDH+ cells) and ER+ cells in the normal human breast epithelium. We assessed the proliferation and lineage differentiation potential of these cells in vitro and in vivo. A gene reporter assay was used to separate live ER+ and ER- mammary epithelial cells. With shRNA-mediated knockdown, we investigated the role of ALDH isoforms in the functionality of mammary epithelial progenitor cells. RESULTS: We describe a cellular hierarchy in the normal human mammary gland in which ER-/ALDH+ cells with functional properties of stem/progenitor cells generate ER+ progenitor cells, which in turn give rise to cells of luminal lineage. We show that the ALDH1A1 isoform, through its function in the retinoic acid metabolism, affects the proliferation and/or early differentiation of stem/progenitor cells and is important for branching morphogenesis. CONCLUSIONS: This study presents direct evidence that ER+ cells are generated by ER-/ALDH+ stem/progenitor cells. We also show that ER+ cells are able to generate cell progeny of luminal lineage in vitro and in vivo. Loss of ALDH1A1 function impairs this process, as well as branching morphogenesis and clonogenicity in suspension culture. This latter effect is reversed by treatment with retinoic acid.
Assuntos
Aldeído Desidrogenase/metabolismo , Glândulas Mamárias Humanas/metabolismo , Receptores de Estrogênio/metabolismo , Tretinoína/farmacologia , Aldeído Desidrogenase/genética , Família Aldeído Desidrogenase 1 , Aldeído Oxirredutases/metabolismo , Diferenciação Celular , Linhagem da Célula , Proliferação de Células , Células Cultivadas , Feminino , Citometria de Fluxo , Humanos , Glândulas Mamárias Humanas/citologia , Isoformas de Proteínas/genética , Interferência de RNA , RNA Interferente Pequeno , Receptores de Estrogênio/biossíntese , Retinal Desidrogenase , Células-Tronco/citologia , Células-Tronco/enzimologia , Tretinoína/metabolismoRESUMO
Tumors may be initiated and maintained by a cellular subcomponent that displays stem cell properties. We have used the expression of aldehyde dehydrogenase as assessed by the ALDEFLUOR assay to isolate and characterize cancer stem cell (CSC) populations in 33 cell lines derived from normal and malignant mammary tissue. Twenty-three of the 33 cell lines contained an ALDEFLUOR-positive population that displayed stem cell properties in vitro and in NOD/SCID xenografts. Gene expression profiling identified a 413-gene CSC profile that included genes known to play a role in stem cell function, as well as genes such as CXCR1/IL-8RA not previously known to play such a role. Recombinant interleukin-8 (IL-8) increased mammosphere formation and the ALDEFLUOR-positive population in breast cancer cell lines. Finally, we show that ALDEFLUOR-positive cells are responsible for mediating metastasis. These studies confirm the hierarchical organization of immortalized cell lines, establish techniques that can facilitate the characterization of regulatory pathways of CSCs, and identify potential stem cell markers and therapeutic targets.
Assuntos
Neoplasias da Mama/patologia , Mama/citologia , Metástase Neoplásica/patologia , Células-Tronco/patologia , Aldeído Desidrogenase/genética , Animais , Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Linhagem Celular , Linhagem Celular Tumoral , Feminino , Citometria de Fluxo , Perfilação da Expressão Gênica , Homeostase , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Análise de Sequência com Séries de Oligonucleotídeos , RNA Mensageiro/genética , RNA Neoplásico/genética , RNA Neoplásico/isolamento & purificação , Receptores de Interleucina-8A/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células-Tronco/citologia , Células-Tronco/fisiologiaRESUMO
We conducted this study to examine whether the expression of c-erbB-2 and p53 is the prognostic indicator for patients with early-stage breast cancer in which axillary lymph node metastasis is absent. We examined 326 patients with early-stage breast cancer in which axillary lymph node metastasis is absent. Tissue microarrays were constructed. Following this, immunohistochemical staining was done for estrogen receptor (ER), progesterone receptor (PR), c-erbB2, and p53. The results were as follows: (1) expression of c-erbB-2 was correlated with other clinicopathologic factors (eg, patient's age, presence of menopause, tumor size, histologic and nuclear grade, and presence of hormone receptors such as ER and PR); and (2) expression of p53 was correlated with survival rate, patient's age, presence of menopause, and tumor size. However, these results were not statistically significant. In conclusion, our results indicate that expression of c-erbB-2 and p53 did not have any prognostic value in patients with early-stage breast cancer in which axillary lymph node metastasis is absent.
