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Hormone-sensing cells require Wip1 for paracrine stimulation in normal and premalignant mammary epithelium.

Tarulli GA, De Silva D, Ho V, Kunasegaran K, Ghosh K, Tan BC, Bulavin DV, Pietersen AM - Breast Cancer Res. (2013)

Bottom Line: In the absence of Wip1, hormone-receptor-positive cells have significantly reduced transcription of RANKL (receptor activator of nuclear factor kappa-B ligand) and IGF2 (insulin-like growth factor 2), paracrine stimulators of alveolar development.In the MMTV-neu model, HER2/neu activates STAT5 in alveolar progenitor cells independent of Wip1, but HER2/neu does not override the defect in STAT5 activation in Wip1-deficient hormone-sensing cells, and paracrine stimulation remains attenuated.Furthermore, our findings highlight that hormone-sensing cells convert not only estrogen and progesterone but also prolactin signals into paracrine instructions for mammary gland development.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Introduction: The molecular circuitry of different cell types dictates their normal function as well as their response to oncogene activation. For instance, mice lacking the Wip1 phosphatase (also known as PPM1D; protein phosphatase magnesium-dependent 1D) have a delay in HER2/neu (human epidermal growth factor 2), but not Wnt1-induced mammary tumor formation. This suggests a cell type-specific reliance on Wip1 for tumorigenesis, because alveolar progenitor cells are the likely target for transformation in the MMTV(mouse mammary tumor virus)-neu but not MMTV-wnt1 breast cancer model.

Methods: In this study, we used the Wip1-knockout mouse to identify the cell types that are dependent on Wip1 expression and therefore may be involved in the early stages of HER2/neu-induced tumorigenesis.

Results: We found that alveolar development during pregnancy was reduced in Wip1-knockout mice; however, this was not attributable to changes in alveolar cells themselves. Unexpectedly, Wip1 allows steroid hormone-receptor-positive cells but not alveolar progenitors to activate STAT5 (signal transducer and activator of transcription 5) in the virgin state. In the absence of Wip1, hormone-receptor-positive cells have significantly reduced transcription of RANKL (receptor activator of nuclear factor kappa-B ligand) and IGF2 (insulin-like growth factor 2), paracrine stimulators of alveolar development. In the MMTV-neu model, HER2/neu activates STAT5 in alveolar progenitor cells independent of Wip1, but HER2/neu does not override the defect in STAT5 activation in Wip1-deficient hormone-sensing cells, and paracrine stimulation remains attenuated. Moreover, ERK (extracellular signal-regulated kinase) activation by HER2/neu in hormone-sensing cells is also Wip1 dependent.

Conclusions: We identified Wip1 as a potentiator of prolactin and HER2/neu signaling strictly in the molecular context of hormone-sensing cells. Furthermore, our findings highlight that hormone-sensing cells convert not only estrogen and progesterone but also prolactin signals into paracrine instructions for mammary gland development. The instructive role of hormone-sensing cells in premalignant development suggests targeting Wip1 or prolactin signaling as an orthogonal strategy for inhibiting breast cancer development or relapse.

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Wip1-knockout animals have reduced alveolar development during pregnancy. (A, B) Carmine-alum-stained whole mounts of mammary glands from virgin (iparous) or 3-days (3d), 7-days (7d), or 14-days (14d) pregnant wild-type (WT, blue box) or Wip1-knockout (KO, green box) mice. Images are representative of three to five animals. Scale bar, 500 μm. (C, D) Hematoxylin and eosin-stained tissue sections of mammary glands from WT (blue box) or Wip1 KO (green box) virgin and pregnant mice, as indicated earlier. Black arrows, distention of lumens present in WT alveolar lobules at 14 days of pregnancy, which are reduced in Wip1 KO sections. Insets are enlarged regions to visualize bilayered epithelium. Dotted lines, location of basement membrane. Images are representative of three animals/group. Scale bar, 100 μm.
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Figure 1: Wip1-knockout animals have reduced alveolar development during pregnancy. (A, B) Carmine-alum-stained whole mounts of mammary glands from virgin (iparous) or 3-days (3d), 7-days (7d), or 14-days (14d) pregnant wild-type (WT, blue box) or Wip1-knockout (KO, green box) mice. Images are representative of three to five animals. Scale bar, 500 μm. (C, D) Hematoxylin and eosin-stained tissue sections of mammary glands from WT (blue box) or Wip1 KO (green box) virgin and pregnant mice, as indicated earlier. Black arrows, distention of lumens present in WT alveolar lobules at 14 days of pregnancy, which are reduced in Wip1 KO sections. Insets are enlarged regions to visualize bilayered epithelium. Dotted lines, location of basement membrane. Images are representative of three animals/group. Scale bar, 100 μm.

Mentions: To elucidate the role of Wip1 in mammary epithelium, we assessed mammary gland development in Wip1-deficient mice at adulthood and during pregnancy. We first examined the morphology of the ductal system by carmine staining of whole mammary glands (Figure 1A, B). The mammary ducts of adult virgin females were indistinguishable between wild-type (WT) and Wip1-knockout (Wip1 KO) mice. Because the mammary gland responds to fluctuations in hormone levels across the estrus cycle by generating and regressing side branches and alveoli on a small scale, we compared each Wip1 KO gland with a control gland from a WT mouse in the same estrus stage (metoestrus). Examination of the ductal architecture at the cellular level with hematoxylin and eosin (H&E) staining of tissue sections (Figure 1C, D) revealed morphologically normal bilayered ducts with proper lumens in the Wip1 KO. To evaluate the effect of loss of Wip1 on alveolar development during pregnancy, animals were timed-mated, and glands were collected at 3, 7, and 14 days of pregnancy. In WT mammary glands, the formation of alveoli becomes evident with carmine whole-mount staining at 7 days of pregnancy, with a further increase in number and size of the alveolar lobules by day 14 of pregnancy (Figure 1A). In contrast, generation of alveolar lobules in Wip1 KO glands is substantially delayed. Analyses of tissue sections show that the initiation of mammary alveolar development can already be detected with H&E in 3-day pregnant WT mice, whereas this is observed only in 7-day pregnant Wip1 KO animals (Figure 1C, D). In WT mammary glands at 14 days of pregnancy, distended lumens become apparent in the developing alveoli, but in the absence of Wip1, the alveolar architecture still resembles that of the WT at 7 days of pregnancy (Figure 1C, D). It is noteworthy that Wip1 KO animals are eventually able to nurse their pups, indicating that alveolar development progresses all the way to functional lactation, but our analyses show an obvious delay in alveologenesis during the initial phase of pregnancy.


Hormone-sensing cells require Wip1 for paracrine stimulation in normal and premalignant mammary epithelium.

Tarulli GA, De Silva D, Ho V, Kunasegaran K, Ghosh K, Tan BC, Bulavin DV, Pietersen AM - Breast Cancer Res. (2013)

Wip1-knockout animals have reduced alveolar development during pregnancy. (A, B) Carmine-alum-stained whole mounts of mammary glands from virgin (iparous) or 3-days (3d), 7-days (7d), or 14-days (14d) pregnant wild-type (WT, blue box) or Wip1-knockout (KO, green box) mice. Images are representative of three to five animals. Scale bar, 500 μm. (C, D) Hematoxylin and eosin-stained tissue sections of mammary glands from WT (blue box) or Wip1 KO (green box) virgin and pregnant mice, as indicated earlier. Black arrows, distention of lumens present in WT alveolar lobules at 14 days of pregnancy, which are reduced in Wip1 KO sections. Insets are enlarged regions to visualize bilayered epithelium. Dotted lines, location of basement membrane. Images are representative of three animals/group. Scale bar, 100 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 1: Wip1-knockout animals have reduced alveolar development during pregnancy. (A, B) Carmine-alum-stained whole mounts of mammary glands from virgin (iparous) or 3-days (3d), 7-days (7d), or 14-days (14d) pregnant wild-type (WT, blue box) or Wip1-knockout (KO, green box) mice. Images are representative of three to five animals. Scale bar, 500 μm. (C, D) Hematoxylin and eosin-stained tissue sections of mammary glands from WT (blue box) or Wip1 KO (green box) virgin and pregnant mice, as indicated earlier. Black arrows, distention of lumens present in WT alveolar lobules at 14 days of pregnancy, which are reduced in Wip1 KO sections. Insets are enlarged regions to visualize bilayered epithelium. Dotted lines, location of basement membrane. Images are representative of three animals/group. Scale bar, 100 μm.
Mentions: To elucidate the role of Wip1 in mammary epithelium, we assessed mammary gland development in Wip1-deficient mice at adulthood and during pregnancy. We first examined the morphology of the ductal system by carmine staining of whole mammary glands (Figure 1A, B). The mammary ducts of adult virgin females were indistinguishable between wild-type (WT) and Wip1-knockout (Wip1 KO) mice. Because the mammary gland responds to fluctuations in hormone levels across the estrus cycle by generating and regressing side branches and alveoli on a small scale, we compared each Wip1 KO gland with a control gland from a WT mouse in the same estrus stage (metoestrus). Examination of the ductal architecture at the cellular level with hematoxylin and eosin (H&E) staining of tissue sections (Figure 1C, D) revealed morphologically normal bilayered ducts with proper lumens in the Wip1 KO. To evaluate the effect of loss of Wip1 on alveolar development during pregnancy, animals were timed-mated, and glands were collected at 3, 7, and 14 days of pregnancy. In WT mammary glands, the formation of alveoli becomes evident with carmine whole-mount staining at 7 days of pregnancy, with a further increase in number and size of the alveolar lobules by day 14 of pregnancy (Figure 1A). In contrast, generation of alveolar lobules in Wip1 KO glands is substantially delayed. Analyses of tissue sections show that the initiation of mammary alveolar development can already be detected with H&E in 3-day pregnant WT mice, whereas this is observed only in 7-day pregnant Wip1 KO animals (Figure 1C, D). In WT mammary glands at 14 days of pregnancy, distended lumens become apparent in the developing alveoli, but in the absence of Wip1, the alveolar architecture still resembles that of the WT at 7 days of pregnancy (Figure 1C, D). It is noteworthy that Wip1 KO animals are eventually able to nurse their pups, indicating that alveolar development progresses all the way to functional lactation, but our analyses show an obvious delay in alveologenesis during the initial phase of pregnancy.

Bottom Line: In the absence of Wip1, hormone-receptor-positive cells have significantly reduced transcription of RANKL (receptor activator of nuclear factor kappa-B ligand) and IGF2 (insulin-like growth factor 2), paracrine stimulators of alveolar development.In the MMTV-neu model, HER2/neu activates STAT5 in alveolar progenitor cells independent of Wip1, but HER2/neu does not override the defect in STAT5 activation in Wip1-deficient hormone-sensing cells, and paracrine stimulation remains attenuated.Furthermore, our findings highlight that hormone-sensing cells convert not only estrogen and progesterone but also prolactin signals into paracrine instructions for mammary gland development.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Introduction: The molecular circuitry of different cell types dictates their normal function as well as their response to oncogene activation. For instance, mice lacking the Wip1 phosphatase (also known as PPM1D; protein phosphatase magnesium-dependent 1D) have a delay in HER2/neu (human epidermal growth factor 2), but not Wnt1-induced mammary tumor formation. This suggests a cell type-specific reliance on Wip1 for tumorigenesis, because alveolar progenitor cells are the likely target for transformation in the MMTV(mouse mammary tumor virus)-neu but not MMTV-wnt1 breast cancer model.

Methods: In this study, we used the Wip1-knockout mouse to identify the cell types that are dependent on Wip1 expression and therefore may be involved in the early stages of HER2/neu-induced tumorigenesis.

Results: We found that alveolar development during pregnancy was reduced in Wip1-knockout mice; however, this was not attributable to changes in alveolar cells themselves. Unexpectedly, Wip1 allows steroid hormone-receptor-positive cells but not alveolar progenitors to activate STAT5 (signal transducer and activator of transcription 5) in the virgin state. In the absence of Wip1, hormone-receptor-positive cells have significantly reduced transcription of RANKL (receptor activator of nuclear factor kappa-B ligand) and IGF2 (insulin-like growth factor 2), paracrine stimulators of alveolar development. In the MMTV-neu model, HER2/neu activates STAT5 in alveolar progenitor cells independent of Wip1, but HER2/neu does not override the defect in STAT5 activation in Wip1-deficient hormone-sensing cells, and paracrine stimulation remains attenuated. Moreover, ERK (extracellular signal-regulated kinase) activation by HER2/neu in hormone-sensing cells is also Wip1 dependent.

Conclusions: We identified Wip1 as a potentiator of prolactin and HER2/neu signaling strictly in the molecular context of hormone-sensing cells. Furthermore, our findings highlight that hormone-sensing cells convert not only estrogen and progesterone but also prolactin signals into paracrine instructions for mammary gland development. The instructive role of hormone-sensing cells in premalignant development suggests targeting Wip1 or prolactin signaling as an orthogonal strategy for inhibiting breast cancer development or relapse.

Show MeSH
Related in: MedlinePlus