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Hormonal induction and roles of Disabled-2 in lactation and involution.

Tao W, Moore R, Smith ER, Xu XX - PLoS ONE (2014)

Bottom Line: Loss of Dab2 had subtle effects on lactation, but Dab2-deficient mammary glands showed a strikingly delayed cell clearance during involution.However, Dab2 deletion did not affect Smad2 phosphorylation; rather TGF-beta-stimulated MAPK activation was enhanced in Dab2-deficient cells.We conclude that Dab2 expression is induced by hormones and Dab2 plays a role in modulating TGF-beta signaling to enhance apoptotic clearance of mammary epithelial cells during involution.

View Article: PubMed Central - PubMed

Affiliation: Sylvester Comprehensive Cancer Center, and Department of Cell Biology, Graduate Program in Molecular Cell and Developmental Biology, University of Miami Miller School of Medicine, Miami, Florida, United States of America.

ABSTRACT
Disabled-2 (Dab2) is a widely expressed endocytic adaptor that was first isolated as a 96 KDa phospho-protein, p96, involved in MAPK signal transduction. Dab2 expression is lost in several cancer types including breast cancer, and Dab2 is thought to have a tumor suppressor function. In mammary epithelia, Dab2 was induced upon pregnancy and further elevated during lactation. We constructed mutant mice with a mosaic Dab2 gene deletion to bypass early embryonic lethality and to investigate the roles of Dab2 in mammary physiology. Loss of Dab2 had subtle effects on lactation, but Dab2-deficient mammary glands showed a strikingly delayed cell clearance during involution. In primary cultures of mouse mammary epithelial cells, Dab2 proteins were also induced by estrogen, progesterone, and/or prolactin. Dab2 mammary epithelial cells were refractory to growth suppression induced by TGF-beta. However, Dab2 deletion did not affect Smad2 phosphorylation; rather TGF-beta-stimulated MAPK activation was enhanced in Dab2-deficient cells. We conclude that Dab2 expression is induced by hormones and Dab2 plays a role in modulating TGF-beta signaling to enhance apoptotic clearance of mammary epithelial cells during involution.

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Dab2 deficient mammary glands have delayed involution.A control (dab2 heterozygous) and dab2 knockout (dab2 (f/df):Sox2-Cre) group of six 6-month old mice were mated, became pregnant, gave birth, and nursed an equal number (six) of pups per mouse. At day 12 of lactation, the mice underwent forced involution by removal of the pups. Mammary glands were harvested accordingly for histological analysis. Consistent results were obtained in two independent experiments using deletion by either Meox2-Cre or Sox2-Cre. (A) Representative H&E images show the morphology of the mammary glands. (B) Images of higher magnification are shown for day 3 and 5 of involution. Accumulating apoptotic cells in the lumen are indicated by a green arrow. (C) Mammary tissues of heterozygous and dab2 knockout at day 3 of forced involution were analyzed by transmission electron microscopy. An arrow indicates the presence of cells in the interior of the dab2 knockout mammary lumens.
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pone-0110737-g005: Dab2 deficient mammary glands have delayed involution.A control (dab2 heterozygous) and dab2 knockout (dab2 (f/df):Sox2-Cre) group of six 6-month old mice were mated, became pregnant, gave birth, and nursed an equal number (six) of pups per mouse. At day 12 of lactation, the mice underwent forced involution by removal of the pups. Mammary glands were harvested accordingly for histological analysis. Consistent results were obtained in two independent experiments using deletion by either Meox2-Cre or Sox2-Cre. (A) Representative H&E images show the morphology of the mammary glands. (B) Images of higher magnification are shown for day 3 and 5 of involution. Accumulating apoptotic cells in the lumen are indicated by a green arrow. (C) Mammary tissues of heterozygous and dab2 knockout at day 3 of forced involution were analyzed by transmission electron microscopy. An arrow indicates the presence of cells in the interior of the dab2 knockout mammary lumens.

Mentions: Despite the induction of Dab2 in wildtype mammary glands, the Dab2-deficient females progressed through pregnancy, lactation, and nursing without any obvious problems. However, we consistently observed that the kinetics of mammary regression were retarded in the Dab2-deficient mammary glands, in which cells with condensed nuclei persisted and cell clearance was delayed (Figure 5). Lactating female mice were separated from their pups 12 days after birth to initiate forced mammary involution, and mammary tissues were analyzed. Accumulation of cells and debris was evidenced in the alveolar lumens of Dab2-deficient mammary glands at day 2 of involution, compared to controls (Figure 5A). In the heterozygous control group, epithelial alveoli regressed greatly by day 3, while adipose cells repopulated the glands. In contrast, Dab2-deficient mammary tissues were still composed mostly of epithelial components at this stage, and few adipocytes were present. Images at higher magnification showed that the lumens harbored a large number of rounded cells with condensed nuclei at day 3 of involution in the Dab2-deficient mammary glands (Figure 5B, arrow). Such cells were present but scarce in control mammary glands. However, by day 5, the differences became minimal, and Dab2-deficiency seemed only to delay but not incapacitate epithelial regression in mammary involution (Figure 5A, B). We have repeatedly detected the delayed mammary involution in multiple (at least 5) independent experiments using groups (3–6 mice in each groups) of control and Dab2- mice over a period of 2 years. Thus, the impact of Dab2 in mammary involution, although transient, is robust and consistent.


Hormonal induction and roles of Disabled-2 in lactation and involution.

Tao W, Moore R, Smith ER, Xu XX - PLoS ONE (2014)

Dab2 deficient mammary glands have delayed involution.A control (dab2 heterozygous) and dab2 knockout (dab2 (f/df):Sox2-Cre) group of six 6-month old mice were mated, became pregnant, gave birth, and nursed an equal number (six) of pups per mouse. At day 12 of lactation, the mice underwent forced involution by removal of the pups. Mammary glands were harvested accordingly for histological analysis. Consistent results were obtained in two independent experiments using deletion by either Meox2-Cre or Sox2-Cre. (A) Representative H&E images show the morphology of the mammary glands. (B) Images of higher magnification are shown for day 3 and 5 of involution. Accumulating apoptotic cells in the lumen are indicated by a green arrow. (C) Mammary tissues of heterozygous and dab2 knockout at day 3 of forced involution were analyzed by transmission electron microscopy. An arrow indicates the presence of cells in the interior of the dab2 knockout mammary lumens.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4216001&req=5

pone-0110737-g005: Dab2 deficient mammary glands have delayed involution.A control (dab2 heterozygous) and dab2 knockout (dab2 (f/df):Sox2-Cre) group of six 6-month old mice were mated, became pregnant, gave birth, and nursed an equal number (six) of pups per mouse. At day 12 of lactation, the mice underwent forced involution by removal of the pups. Mammary glands were harvested accordingly for histological analysis. Consistent results were obtained in two independent experiments using deletion by either Meox2-Cre or Sox2-Cre. (A) Representative H&E images show the morphology of the mammary glands. (B) Images of higher magnification are shown for day 3 and 5 of involution. Accumulating apoptotic cells in the lumen are indicated by a green arrow. (C) Mammary tissues of heterozygous and dab2 knockout at day 3 of forced involution were analyzed by transmission electron microscopy. An arrow indicates the presence of cells in the interior of the dab2 knockout mammary lumens.
Mentions: Despite the induction of Dab2 in wildtype mammary glands, the Dab2-deficient females progressed through pregnancy, lactation, and nursing without any obvious problems. However, we consistently observed that the kinetics of mammary regression were retarded in the Dab2-deficient mammary glands, in which cells with condensed nuclei persisted and cell clearance was delayed (Figure 5). Lactating female mice were separated from their pups 12 days after birth to initiate forced mammary involution, and mammary tissues were analyzed. Accumulation of cells and debris was evidenced in the alveolar lumens of Dab2-deficient mammary glands at day 2 of involution, compared to controls (Figure 5A). In the heterozygous control group, epithelial alveoli regressed greatly by day 3, while adipose cells repopulated the glands. In contrast, Dab2-deficient mammary tissues were still composed mostly of epithelial components at this stage, and few adipocytes were present. Images at higher magnification showed that the lumens harbored a large number of rounded cells with condensed nuclei at day 3 of involution in the Dab2-deficient mammary glands (Figure 5B, arrow). Such cells were present but scarce in control mammary glands. However, by day 5, the differences became minimal, and Dab2-deficiency seemed only to delay but not incapacitate epithelial regression in mammary involution (Figure 5A, B). We have repeatedly detected the delayed mammary involution in multiple (at least 5) independent experiments using groups (3–6 mice in each groups) of control and Dab2- mice over a period of 2 years. Thus, the impact of Dab2 in mammary involution, although transient, is robust and consistent.

Bottom Line: Loss of Dab2 had subtle effects on lactation, but Dab2-deficient mammary glands showed a strikingly delayed cell clearance during involution.However, Dab2 deletion did not affect Smad2 phosphorylation; rather TGF-beta-stimulated MAPK activation was enhanced in Dab2-deficient cells.We conclude that Dab2 expression is induced by hormones and Dab2 plays a role in modulating TGF-beta signaling to enhance apoptotic clearance of mammary epithelial cells during involution.

View Article: PubMed Central - PubMed

Affiliation: Sylvester Comprehensive Cancer Center, and Department of Cell Biology, Graduate Program in Molecular Cell and Developmental Biology, University of Miami Miller School of Medicine, Miami, Florida, United States of America.

ABSTRACT
Disabled-2 (Dab2) is a widely expressed endocytic adaptor that was first isolated as a 96 KDa phospho-protein, p96, involved in MAPK signal transduction. Dab2 expression is lost in several cancer types including breast cancer, and Dab2 is thought to have a tumor suppressor function. In mammary epithelia, Dab2 was induced upon pregnancy and further elevated during lactation. We constructed mutant mice with a mosaic Dab2 gene deletion to bypass early embryonic lethality and to investigate the roles of Dab2 in mammary physiology. Loss of Dab2 had subtle effects on lactation, but Dab2-deficient mammary glands showed a strikingly delayed cell clearance during involution. In primary cultures of mouse mammary epithelial cells, Dab2 proteins were also induced by estrogen, progesterone, and/or prolactin. Dab2 mammary epithelial cells were refractory to growth suppression induced by TGF-beta. However, Dab2 deletion did not affect Smad2 phosphorylation; rather TGF-beta-stimulated MAPK activation was enhanced in Dab2-deficient cells. We conclude that Dab2 expression is induced by hormones and Dab2 plays a role in modulating TGF-beta signaling to enhance apoptotic clearance of mammary epithelial cells during involution.

Show MeSH
Related in: MedlinePlus