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ER-α36-mediated rapid estrogen signaling positively regulates ER-positive breast cancer stem/progenitor cells.

Deng H, Zhang XT, Wang ML, Zheng HY, Liu LJ, Wang ZY - PLoS ONE (2014)

Bottom Line: We found that 17-β-estradiol (E2β) treatment increased the population of ER-positive breast cancer stem/progenitor cells while failed to do so in the cells with knocked-down levels of ER-α36 expression.Cells with forced expression of recombinant ER-α36, however, responded strongly to E2β treatment by increasing growth in vitro and tumor-seeding efficiency in vivo.We concluded that ER-α36-mediated rapid estrogen signaling plays an important role in regulation and maintenance of ER-positive breast cancer stem/progenitor cells.

View Article: PubMed Central - PubMed

Affiliation: Departments of Medical Microbiology and Immunology, Creighton University Medical School, Omaha, Nebraska, United States of America ; Jiangda Pathology Center, Jianghan University, Wuhan, Hubei, P. R. China.

ABSTRACT
The breast cancer stem cells (BCSC) play important roles in breast cancer occurrence, recurrence and metastasis. However, the role of estrogen signaling, a signaling pathway important in development and progression of breast cancer, in regulation of BCSC has not been well established. Previously, we identified and cloned a variant of estrogen receptor α, ER-α36, with a molecular weight of 36 kDa. ER-α36 lacks both transactivation domains AF-1 and AF-2 of the 66 kDa full-length ER-α (ER-α66) and mediates rapid estrogen signaling to promote proliferation of breast cancer cells. In this study, we aim to investigate the function and the underlying mechanism of ER-α36-mediated rapid estrogen signaling in growth regulation of the ER-positive breast cancer stem/progenitor cells. ER-positive breast cancer cells MCF7 and T47D as well as the variants with different levels of ER-α36 expression were used. The effects of estrogen on BCSC's abilities of growth, self-renewal, differentiation and tumor-seeding were examined using tumorsphere formation, flow cytometry, indirect immunofluorence staining and in vivo xenograft assays. The underlying mechanisms were also studied with Western-blot analysis. We found that 17-β-estradiol (E2β) treatment increased the population of ER-positive breast cancer stem/progenitor cells while failed to do so in the cells with knocked-down levels of ER-α36 expression. Cells with forced expression of recombinant ER-α36, however, responded strongly to E2β treatment by increasing growth in vitro and tumor-seeding efficiency in vivo. The rapid estrogen signaling via the AKT/GSK3β pathway is involved in estrogen-stimulated growth of ER-positive breast cancer stem/progenitor cells. We concluded that ER-α36-mediated rapid estrogen signaling plays an important role in regulation and maintenance of ER-positive breast cancer stem/progenitor cells.

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Estrogen expands the population of ER-positive breast cancer stem/progenitor cells.ER-positive breast cancer MCF7 and T47D cells were used. The tumorsphere formation assay and flow cytometry analysis of the CD44− and CD24+ cells were used to assess the population of ER-positive breast cancer stem/progenitor cells. (A). Estrogen treatment increases the population of the CD44−/CD24+ cells in MCF7 and T47D cells. The monolayer (parental) and tumorspheres of MCF7 and T47D cells were treated with vehicle (ethanol) or 0.1 nM of E2β for five days. The population of CD44−/CD24+ cells in these cells were analyzed after staining with fluorochrome-conjugated antibodies. The representative results are shown on the upper panel. Lower panel: the columns represent the means of three experiments; bars, SE. *, P<0.05 for vehicle treated cells vs cells treated with E2β. (B). Estrogen treatment increases the size of tumorspheres from MCF7 and T47D cells. A representative tumorsphere from MCF7 and T47D cells treated with vehicle or 0.1 nM E2β for seven days. (C). Estrogen treatment increases the number of tumorspheres and cells from dissociated tumorspheres derived from MCF7 and T47D cells. The columns represent the means of three experiments; bars, SE. *, P<0.05 for cells treated with vehicle vs cells treated with E2β.
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pone-0088034-g001: Estrogen expands the population of ER-positive breast cancer stem/progenitor cells.ER-positive breast cancer MCF7 and T47D cells were used. The tumorsphere formation assay and flow cytometry analysis of the CD44− and CD24+ cells were used to assess the population of ER-positive breast cancer stem/progenitor cells. (A). Estrogen treatment increases the population of the CD44−/CD24+ cells in MCF7 and T47D cells. The monolayer (parental) and tumorspheres of MCF7 and T47D cells were treated with vehicle (ethanol) or 0.1 nM of E2β for five days. The population of CD44−/CD24+ cells in these cells were analyzed after staining with fluorochrome-conjugated antibodies. The representative results are shown on the upper panel. Lower panel: the columns represent the means of three experiments; bars, SE. *, P<0.05 for vehicle treated cells vs cells treated with E2β. (B). Estrogen treatment increases the size of tumorspheres from MCF7 and T47D cells. A representative tumorsphere from MCF7 and T47D cells treated with vehicle or 0.1 nM E2β for seven days. (C). Estrogen treatment increases the number of tumorspheres and cells from dissociated tumorspheres derived from MCF7 and T47D cells. The columns represent the means of three experiments; bars, SE. *, P<0.05 for cells treated with vehicle vs cells treated with E2β.

Mentions: To examine the effects of estrogen signaling on growth of ER-positive breast cancer stem/progenitor cells, we used the well-characterized ER-positive breast cancer MCF7 and T47D cells as models. Both MCF7 and T47D cells were treated with or without 0.1 nM of E2β for five days. The CD44+/CD24− stem-like cell populations in these cells were assessed with flow cytometry. We found that estrogen treatment significantly increased the CD44+/CD24− cell population in both MCF7 and T47D cells (Figure 1A). We then cultured both MCF7 and T47D cells in the tumorsphere medium and under suspension conditions to form tumorspheres. We found that E2β treatment also increased the CD44+/CD24− cell populations in tumorspheres from these cells (Figure 1A). We also found that E2β treatment markedly increased the size and number of the tumorspheres as well as the number of cells in the tumorspheres (Figure 1B and C). Our results thus suggested that estrogen signaling increases the population of ER-positive breast cancer stem/progenitor cells.


ER-α36-mediated rapid estrogen signaling positively regulates ER-positive breast cancer stem/progenitor cells.

Deng H, Zhang XT, Wang ML, Zheng HY, Liu LJ, Wang ZY - PLoS ONE (2014)

Estrogen expands the population of ER-positive breast cancer stem/progenitor cells.ER-positive breast cancer MCF7 and T47D cells were used. The tumorsphere formation assay and flow cytometry analysis of the CD44− and CD24+ cells were used to assess the population of ER-positive breast cancer stem/progenitor cells. (A). Estrogen treatment increases the population of the CD44−/CD24+ cells in MCF7 and T47D cells. The monolayer (parental) and tumorspheres of MCF7 and T47D cells were treated with vehicle (ethanol) or 0.1 nM of E2β for five days. The population of CD44−/CD24+ cells in these cells were analyzed after staining with fluorochrome-conjugated antibodies. The representative results are shown on the upper panel. Lower panel: the columns represent the means of three experiments; bars, SE. *, P<0.05 for vehicle treated cells vs cells treated with E2β. (B). Estrogen treatment increases the size of tumorspheres from MCF7 and T47D cells. A representative tumorsphere from MCF7 and T47D cells treated with vehicle or 0.1 nM E2β for seven days. (C). Estrogen treatment increases the number of tumorspheres and cells from dissociated tumorspheres derived from MCF7 and T47D cells. The columns represent the means of three experiments; bars, SE. *, P<0.05 for cells treated with vehicle vs cells treated with E2β.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3928099&req=5

pone-0088034-g001: Estrogen expands the population of ER-positive breast cancer stem/progenitor cells.ER-positive breast cancer MCF7 and T47D cells were used. The tumorsphere formation assay and flow cytometry analysis of the CD44− and CD24+ cells were used to assess the population of ER-positive breast cancer stem/progenitor cells. (A). Estrogen treatment increases the population of the CD44−/CD24+ cells in MCF7 and T47D cells. The monolayer (parental) and tumorspheres of MCF7 and T47D cells were treated with vehicle (ethanol) or 0.1 nM of E2β for five days. The population of CD44−/CD24+ cells in these cells were analyzed after staining with fluorochrome-conjugated antibodies. The representative results are shown on the upper panel. Lower panel: the columns represent the means of three experiments; bars, SE. *, P<0.05 for vehicle treated cells vs cells treated with E2β. (B). Estrogen treatment increases the size of tumorspheres from MCF7 and T47D cells. A representative tumorsphere from MCF7 and T47D cells treated with vehicle or 0.1 nM E2β for seven days. (C). Estrogen treatment increases the number of tumorspheres and cells from dissociated tumorspheres derived from MCF7 and T47D cells. The columns represent the means of three experiments; bars, SE. *, P<0.05 for cells treated with vehicle vs cells treated with E2β.
Mentions: To examine the effects of estrogen signaling on growth of ER-positive breast cancer stem/progenitor cells, we used the well-characterized ER-positive breast cancer MCF7 and T47D cells as models. Both MCF7 and T47D cells were treated with or without 0.1 nM of E2β for five days. The CD44+/CD24− stem-like cell populations in these cells were assessed with flow cytometry. We found that estrogen treatment significantly increased the CD44+/CD24− cell population in both MCF7 and T47D cells (Figure 1A). We then cultured both MCF7 and T47D cells in the tumorsphere medium and under suspension conditions to form tumorspheres. We found that E2β treatment also increased the CD44+/CD24− cell populations in tumorspheres from these cells (Figure 1A). We also found that E2β treatment markedly increased the size and number of the tumorspheres as well as the number of cells in the tumorspheres (Figure 1B and C). Our results thus suggested that estrogen signaling increases the population of ER-positive breast cancer stem/progenitor cells.

Bottom Line: We found that 17-β-estradiol (E2β) treatment increased the population of ER-positive breast cancer stem/progenitor cells while failed to do so in the cells with knocked-down levels of ER-α36 expression.Cells with forced expression of recombinant ER-α36, however, responded strongly to E2β treatment by increasing growth in vitro and tumor-seeding efficiency in vivo.We concluded that ER-α36-mediated rapid estrogen signaling plays an important role in regulation and maintenance of ER-positive breast cancer stem/progenitor cells.

View Article: PubMed Central - PubMed

Affiliation: Departments of Medical Microbiology and Immunology, Creighton University Medical School, Omaha, Nebraska, United States of America ; Jiangda Pathology Center, Jianghan University, Wuhan, Hubei, P. R. China.

ABSTRACT
The breast cancer stem cells (BCSC) play important roles in breast cancer occurrence, recurrence and metastasis. However, the role of estrogen signaling, a signaling pathway important in development and progression of breast cancer, in regulation of BCSC has not been well established. Previously, we identified and cloned a variant of estrogen receptor α, ER-α36, with a molecular weight of 36 kDa. ER-α36 lacks both transactivation domains AF-1 and AF-2 of the 66 kDa full-length ER-α (ER-α66) and mediates rapid estrogen signaling to promote proliferation of breast cancer cells. In this study, we aim to investigate the function and the underlying mechanism of ER-α36-mediated rapid estrogen signaling in growth regulation of the ER-positive breast cancer stem/progenitor cells. ER-positive breast cancer cells MCF7 and T47D as well as the variants with different levels of ER-α36 expression were used. The effects of estrogen on BCSC's abilities of growth, self-renewal, differentiation and tumor-seeding were examined using tumorsphere formation, flow cytometry, indirect immunofluorence staining and in vivo xenograft assays. The underlying mechanisms were also studied with Western-blot analysis. We found that 17-β-estradiol (E2β) treatment increased the population of ER-positive breast cancer stem/progenitor cells while failed to do so in the cells with knocked-down levels of ER-α36 expression. Cells with forced expression of recombinant ER-α36, however, responded strongly to E2β treatment by increasing growth in vitro and tumor-seeding efficiency in vivo. The rapid estrogen signaling via the AKT/GSK3β pathway is involved in estrogen-stimulated growth of ER-positive breast cancer stem/progenitor cells. We concluded that ER-α36-mediated rapid estrogen signaling plays an important role in regulation and maintenance of ER-positive breast cancer stem/progenitor cells.

Show MeSH
Related in: MedlinePlus