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SirT1 modulates the estrogen-insulin-like growth factor-1 signaling for postnatal development of mammary gland in mice.

Li H, Rajendran GK, Liu N, Ware C, Rubin BP, Gu Y - Breast Cancer Res. (2007)

Bottom Line: The effect of exogenous estrogen was also examined by subcutaneous implantation of a slow-releasing pellet in the subscapular region.SirT1 deficiency deregulates the expression of IGF-1 binding protein-1 and attenuates the effect of IGF-1 signals, including estrogen-stimulated local IGF-1 signaling for the onset of ductal morphogenesis.These findings suggest that the enzymatic activity of SirT1 may influence both normal growth and malignant growth of mammary epithelial cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Radiation Oncology, University of Washington School of Medicine, Seattle, WA 98195, USA.

ABSTRACT

Introduction: Estrogen and insulin-like growth factor-1 (IGF-1) play important roles in mammary gland development and breast cancer. SirT1 is a highly conserved protein deacetylase that can regulate the insulin/IGF-1 signaling in lower organisms, as well as a growing number of transcription factors, including NF-kappaB, in mammalian cells. Whether SirT1 regulates the IGF-1 signaling for mammary gland development and function, however, is not clear. In the present study, this role of SirT1 was examined by studying SirT1-deficient mice.

Methods: SirT1-deficient (SirT1(ko/ko)) mice were generated by crossing a new strain of mice harboring a conditional targeted mutation in the SirT1 gene (SirT1(co/co)) with CMV-Cre transgenic mice. Whole mount and histology analyses, immunofluorescence staining, immunohistochemistry, and western blotting were used to characterize mammary gland development in virgin and pregnant mice. The effect of exogenous estrogen was also examined by subcutaneous implantation of a slow-releasing pellet in the subscapular region.

Results: Both male and female SirT1(ko/ko) mice can be fertile despite the growth retardation phenotype. Virgin SirT1(ko/ko) mice displayed impeded ductal morphogenesis, whereas pregnant SirT1(ko/ko) mice manifested lactation failure due to an underdeveloped lobuloalveolar network. Estrogen implantation was sufficient to rescue ductal morphogenesis. Exogenous estrogen reversed the increased basal level of IGF-1 binding protein-1 expression in SirT1(ko/ko) mammary tissues, but not that of IkappaB alpha expression, suggesting that increased levels of estrogen enhanced the production of local IGF-1 and rescued ductal morphogenesis. Additionally, TNFalpha treatment enhanced the level of the newly synthesized IkappaB alpha in SirT1(ko/ko) cells. SirT1 deficiency therefore affects the cellular response to multiple extrinsic signals.

Conclusion: SirT1 modulates the IGF-1 signaling critical for both growth regulation and mammary gland development in mice. SirT1 deficiency deregulates the expression of IGF-1 binding protein-1 and attenuates the effect of IGF-1 signals, including estrogen-stimulated local IGF-1 signaling for the onset of ductal morphogenesis. These findings suggest that the enzymatic activity of SirT1 may influence both normal growth and malignant growth of mammary epithelial cells.

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Estrogen implantation stimulates ductal elongation and site branching. (a) Whole mount analysis of the mammary tissues from the virgin wild-type (+/+) and SirT1ko/ko mice implanted with estrogen pellets (+E2) on day 14 and day 21. Scale bars = 200 μm. (b) Western blot analysis of the expression of insulin-like growth factor-1 binding protein-1 (IGFBP-1) and IκBα, with actin as a loading control, in mammary tissues from the virgin wild-type (+/+) and SirT1ko/ko mice implanted with estrogen pellets on day 21.
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Figure 4: Estrogen implantation stimulates ductal elongation and site branching. (a) Whole mount analysis of the mammary tissues from the virgin wild-type (+/+) and SirT1ko/ko mice implanted with estrogen pellets (+E2) on day 14 and day 21. Scale bars = 200 μm. (b) Western blot analysis of the expression of insulin-like growth factor-1 binding protein-1 (IGFBP-1) and IκBα, with actin as a loading control, in mammary tissues from the virgin wild-type (+/+) and SirT1ko/ko mice implanted with estrogen pellets on day 21.

Mentions: To test whether the increased levels of estrogen in pregnant mice were sufficient to induce ductal morphogenesis in SirT1ko/ko mice, a single estrogen pellet was implanted in the subscapular region of each virgin SirT1ko/ko mouse and the mammary tissues were analyzed 14 and 21 days after the implantation. By day 14, wild-type mice displayed ductal side branching reminiscent of the morphological changes during early pregnancy (Figure 4a, left panel). In contrast, SirT1ko/ko mice showed TEBs and ductal elongation, which are the characteristic features of ductal morphogenesis in pubertal wild-type mice. By day 21, ductal elongation and side branching were restored in SirT1ko/ko mice and the extent of ductal morphogenesis was indistinguishable between wild-type and SirT1ko/ko female mice (Figure 4a, right panel). These observations clearly indicated that exogenous estrogen alone is sufficient to rescue ductal morphogenesis in virgin SirT1ko/ko mice. Furthermore, the presence of transitional TEBs on day 14 and of ductal side branching on day 21 suggested that the characteristic features of mammary gland development at puberty and during early pregnancy could be coupled in response to increasing levels of estrogen. Increased levels of estrogen, either during pregnancy or through implantation, were therefore sufficient to stimulate the differentiation of epithelial progenitor cells and ductal morphogenesis in virgin SirT1ko/ko mice.


SirT1 modulates the estrogen-insulin-like growth factor-1 signaling for postnatal development of mammary gland in mice.

Li H, Rajendran GK, Liu N, Ware C, Rubin BP, Gu Y - Breast Cancer Res. (2007)

Estrogen implantation stimulates ductal elongation and site branching. (a) Whole mount analysis of the mammary tissues from the virgin wild-type (+/+) and SirT1ko/ko mice implanted with estrogen pellets (+E2) on day 14 and day 21. Scale bars = 200 μm. (b) Western blot analysis of the expression of insulin-like growth factor-1 binding protein-1 (IGFBP-1) and IκBα, with actin as a loading control, in mammary tissues from the virgin wild-type (+/+) and SirT1ko/ko mice implanted with estrogen pellets on day 21.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC1851382&req=5

Figure 4: Estrogen implantation stimulates ductal elongation and site branching. (a) Whole mount analysis of the mammary tissues from the virgin wild-type (+/+) and SirT1ko/ko mice implanted with estrogen pellets (+E2) on day 14 and day 21. Scale bars = 200 μm. (b) Western blot analysis of the expression of insulin-like growth factor-1 binding protein-1 (IGFBP-1) and IκBα, with actin as a loading control, in mammary tissues from the virgin wild-type (+/+) and SirT1ko/ko mice implanted with estrogen pellets on day 21.
Mentions: To test whether the increased levels of estrogen in pregnant mice were sufficient to induce ductal morphogenesis in SirT1ko/ko mice, a single estrogen pellet was implanted in the subscapular region of each virgin SirT1ko/ko mouse and the mammary tissues were analyzed 14 and 21 days after the implantation. By day 14, wild-type mice displayed ductal side branching reminiscent of the morphological changes during early pregnancy (Figure 4a, left panel). In contrast, SirT1ko/ko mice showed TEBs and ductal elongation, which are the characteristic features of ductal morphogenesis in pubertal wild-type mice. By day 21, ductal elongation and side branching were restored in SirT1ko/ko mice and the extent of ductal morphogenesis was indistinguishable between wild-type and SirT1ko/ko female mice (Figure 4a, right panel). These observations clearly indicated that exogenous estrogen alone is sufficient to rescue ductal morphogenesis in virgin SirT1ko/ko mice. Furthermore, the presence of transitional TEBs on day 14 and of ductal side branching on day 21 suggested that the characteristic features of mammary gland development at puberty and during early pregnancy could be coupled in response to increasing levels of estrogen. Increased levels of estrogen, either during pregnancy or through implantation, were therefore sufficient to stimulate the differentiation of epithelial progenitor cells and ductal morphogenesis in virgin SirT1ko/ko mice.

Bottom Line: The effect of exogenous estrogen was also examined by subcutaneous implantation of a slow-releasing pellet in the subscapular region.SirT1 deficiency deregulates the expression of IGF-1 binding protein-1 and attenuates the effect of IGF-1 signals, including estrogen-stimulated local IGF-1 signaling for the onset of ductal morphogenesis.These findings suggest that the enzymatic activity of SirT1 may influence both normal growth and malignant growth of mammary epithelial cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Radiation Oncology, University of Washington School of Medicine, Seattle, WA 98195, USA.

ABSTRACT

Introduction: Estrogen and insulin-like growth factor-1 (IGF-1) play important roles in mammary gland development and breast cancer. SirT1 is a highly conserved protein deacetylase that can regulate the insulin/IGF-1 signaling in lower organisms, as well as a growing number of transcription factors, including NF-kappaB, in mammalian cells. Whether SirT1 regulates the IGF-1 signaling for mammary gland development and function, however, is not clear. In the present study, this role of SirT1 was examined by studying SirT1-deficient mice.

Methods: SirT1-deficient (SirT1(ko/ko)) mice were generated by crossing a new strain of mice harboring a conditional targeted mutation in the SirT1 gene (SirT1(co/co)) with CMV-Cre transgenic mice. Whole mount and histology analyses, immunofluorescence staining, immunohistochemistry, and western blotting were used to characterize mammary gland development in virgin and pregnant mice. The effect of exogenous estrogen was also examined by subcutaneous implantation of a slow-releasing pellet in the subscapular region.

Results: Both male and female SirT1(ko/ko) mice can be fertile despite the growth retardation phenotype. Virgin SirT1(ko/ko) mice displayed impeded ductal morphogenesis, whereas pregnant SirT1(ko/ko) mice manifested lactation failure due to an underdeveloped lobuloalveolar network. Estrogen implantation was sufficient to rescue ductal morphogenesis. Exogenous estrogen reversed the increased basal level of IGF-1 binding protein-1 expression in SirT1(ko/ko) mammary tissues, but not that of IkappaB alpha expression, suggesting that increased levels of estrogen enhanced the production of local IGF-1 and rescued ductal morphogenesis. Additionally, TNFalpha treatment enhanced the level of the newly synthesized IkappaB alpha in SirT1(ko/ko) cells. SirT1 deficiency therefore affects the cellular response to multiple extrinsic signals.

Conclusion: SirT1 modulates the IGF-1 signaling critical for both growth regulation and mammary gland development in mice. SirT1 deficiency deregulates the expression of IGF-1 binding protein-1 and attenuates the effect of IGF-1 signals, including estrogen-stimulated local IGF-1 signaling for the onset of ductal morphogenesis. These findings suggest that the enzymatic activity of SirT1 may influence both normal growth and malignant growth of mammary epithelial cells.

Show MeSH
Related in: MedlinePlus