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Lactation failure in Src knockout mice is due to impaired secretory activation.

Watkin H, Richert MM, Lewis A, Terrell K, McManaman JP, Anderson SM - BMC Dev. Biol. (2008)

Bottom Line: Failed secretory activation results in precocious involution in the mammary glands of Src-/- even when pups were suckling.Involution was accelerated following pup withdrawal perhaps as a result of incomplete secretory activation.Src appears to be required for increased expression of the prolactin receptor and successful downstream signaling, and alveolar cell organization.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathology, University of Colorado Health Sciences Center, Research Complex I, South Tower, Mail Stop 8104, 12801 East 17th Avenue, Aurora, CO 80045, USA. harriet.watkin@uchsc.edu

ABSTRACT

Background: Mammary gland development culminates in lactation and is orchestrated by numerous stimuli and signaling pathways. The Src family of nonreceptor tyrosine kinases plays a pivotal role in cell signaling. In order to determine if Src plays a role in mammary gland development we have examined mammary gland development and function during pregnancy and lactation in mice in which expression of Src has been eliminated.

Results: We have characterized a lactation defect in the Src-/- mice which results in the death of over 80% of the litters nursed by Src-/- dams. Mammary gland development during pregnancy appears normal in these mice; however secretory activation does not seem to occur. Serum prolactin levels are normal in Src-/- mice compared to wildtype controls. Expression of the prolactin receptor at both the RNA and protein level was decreased in Src-/- mice following the transition from pregnancy to lactation, as was phosphorylation of STAT5 and expression of milk protein genes. These results suggest that secretory activation, which occurs following parturition, does not occur completely in Src-/- mice. Failed secretory activation results in precocious involution in the mammary glands of Src-/- even when pups were suckling. Involution was accelerated following pup withdrawal perhaps as a result of incomplete secretory activation. In vitro differentiation of mammary epithelial cells from Src-/- mice resulted in diminished production of milk proteins compared to the amount of milk proteins produced by Src+/+ cells, indicating a direct role for Src in regulating the transcription/translation of milk protein genes in mammary epithelial cells.

Conclusion: Src is an essential signaling modulator in mammary gland development as Src-/- mice exhibit a block in secretory activation that results in lactation failure and precocious involution. Src appears to be required for increased expression of the prolactin receptor and successful downstream signaling, and alveolar cell organization.

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Secretory activation does not occur in Src-/- mice following parturition. Histological sections were prepared from the mammary glands of Src+/+ (A-C), Src+/- (D-F), and Src-/- (G-I) mice at P18 (A, D, and G) and L2 (B, C, E, F, H and I). The sections shown in A, B, D, E, G and H were stained anti-ADRP antibody to outline the cytoplasmic lipid droplets (red), Oregon-Green 488-conjugated WPA to outline the surface of secretory alveoli (green), and DAPI to stain the nuclei of mammary epithelial cells (blue). Hematoxylin and eosin-stained sections of mammary glands at L2 are shown in C, F, and I. The bar in panel H represents 10 μm and the bars in panels C, F and I represent 100 μm. Luminal space is indicated by the letter "L", the white arrowheads indicate cytoplasmic lipid droplets, and the white arrows indicates regions where staining with WGA represents atypical patterns not observed in wildtype mice. The black arrows in panel I indicate large cytoplasmic lipid droplets not observed in wildtype mice.
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Figure 5: Secretory activation does not occur in Src-/- mice following parturition. Histological sections were prepared from the mammary glands of Src+/+ (A-C), Src+/- (D-F), and Src-/- (G-I) mice at P18 (A, D, and G) and L2 (B, C, E, F, H and I). The sections shown in A, B, D, E, G and H were stained anti-ADRP antibody to outline the cytoplasmic lipid droplets (red), Oregon-Green 488-conjugated WPA to outline the surface of secretory alveoli (green), and DAPI to stain the nuclei of mammary epithelial cells (blue). Hematoxylin and eosin-stained sections of mammary glands at L2 are shown in C, F, and I. The bar in panel H represents 10 μm and the bars in panels C, F and I represent 100 μm. Luminal space is indicated by the letter "L", the white arrowheads indicate cytoplasmic lipid droplets, and the white arrows indicates regions where staining with WGA represents atypical patterns not observed in wildtype mice. The black arrows in panel I indicate large cytoplasmic lipid droplets not observed in wildtype mice.

Mentions: As described above, the mammary glands of Src-/- mice appeared similar to those of Src+/+ and Src+/- mice at P18 with prominent densely-staining secretory alveoli surrounded by adipocytes. By day 2 of lactation (L2) the luminal space of the alveoli from Src+/+ and Src-/- mice had expanded and are clearly evident due to the layer of densely staining mammary epithelial cells that surround the lumen (Figure 4, panels A and B). The mammary epithelial cells of the Src-/- mice have an abundance of large lipid droplets that are not present in mammary epithelial cells from Src+/+ or Src+/- mice, this observation was further investigated (See Figure 5 and below).


Lactation failure in Src knockout mice is due to impaired secretory activation.

Watkin H, Richert MM, Lewis A, Terrell K, McManaman JP, Anderson SM - BMC Dev. Biol. (2008)

Secretory activation does not occur in Src-/- mice following parturition. Histological sections were prepared from the mammary glands of Src+/+ (A-C), Src+/- (D-F), and Src-/- (G-I) mice at P18 (A, D, and G) and L2 (B, C, E, F, H and I). The sections shown in A, B, D, E, G and H were stained anti-ADRP antibody to outline the cytoplasmic lipid droplets (red), Oregon-Green 488-conjugated WPA to outline the surface of secretory alveoli (green), and DAPI to stain the nuclei of mammary epithelial cells (blue). Hematoxylin and eosin-stained sections of mammary glands at L2 are shown in C, F, and I. The bar in panel H represents 10 μm and the bars in panels C, F and I represent 100 μm. Luminal space is indicated by the letter "L", the white arrowheads indicate cytoplasmic lipid droplets, and the white arrows indicates regions where staining with WGA represents atypical patterns not observed in wildtype mice. The black arrows in panel I indicate large cytoplasmic lipid droplets not observed in wildtype mice.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 5: Secretory activation does not occur in Src-/- mice following parturition. Histological sections were prepared from the mammary glands of Src+/+ (A-C), Src+/- (D-F), and Src-/- (G-I) mice at P18 (A, D, and G) and L2 (B, C, E, F, H and I). The sections shown in A, B, D, E, G and H were stained anti-ADRP antibody to outline the cytoplasmic lipid droplets (red), Oregon-Green 488-conjugated WPA to outline the surface of secretory alveoli (green), and DAPI to stain the nuclei of mammary epithelial cells (blue). Hematoxylin and eosin-stained sections of mammary glands at L2 are shown in C, F, and I. The bar in panel H represents 10 μm and the bars in panels C, F and I represent 100 μm. Luminal space is indicated by the letter "L", the white arrowheads indicate cytoplasmic lipid droplets, and the white arrows indicates regions where staining with WGA represents atypical patterns not observed in wildtype mice. The black arrows in panel I indicate large cytoplasmic lipid droplets not observed in wildtype mice.
Mentions: As described above, the mammary glands of Src-/- mice appeared similar to those of Src+/+ and Src+/- mice at P18 with prominent densely-staining secretory alveoli surrounded by adipocytes. By day 2 of lactation (L2) the luminal space of the alveoli from Src+/+ and Src-/- mice had expanded and are clearly evident due to the layer of densely staining mammary epithelial cells that surround the lumen (Figure 4, panels A and B). The mammary epithelial cells of the Src-/- mice have an abundance of large lipid droplets that are not present in mammary epithelial cells from Src+/+ or Src+/- mice, this observation was further investigated (See Figure 5 and below).

Bottom Line: Failed secretory activation results in precocious involution in the mammary glands of Src-/- even when pups were suckling.Involution was accelerated following pup withdrawal perhaps as a result of incomplete secretory activation.Src appears to be required for increased expression of the prolactin receptor and successful downstream signaling, and alveolar cell organization.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathology, University of Colorado Health Sciences Center, Research Complex I, South Tower, Mail Stop 8104, 12801 East 17th Avenue, Aurora, CO 80045, USA. harriet.watkin@uchsc.edu

ABSTRACT

Background: Mammary gland development culminates in lactation and is orchestrated by numerous stimuli and signaling pathways. The Src family of nonreceptor tyrosine kinases plays a pivotal role in cell signaling. In order to determine if Src plays a role in mammary gland development we have examined mammary gland development and function during pregnancy and lactation in mice in which expression of Src has been eliminated.

Results: We have characterized a lactation defect in the Src-/- mice which results in the death of over 80% of the litters nursed by Src-/- dams. Mammary gland development during pregnancy appears normal in these mice; however secretory activation does not seem to occur. Serum prolactin levels are normal in Src-/- mice compared to wildtype controls. Expression of the prolactin receptor at both the RNA and protein level was decreased in Src-/- mice following the transition from pregnancy to lactation, as was phosphorylation of STAT5 and expression of milk protein genes. These results suggest that secretory activation, which occurs following parturition, does not occur completely in Src-/- mice. Failed secretory activation results in precocious involution in the mammary glands of Src-/- even when pups were suckling. Involution was accelerated following pup withdrawal perhaps as a result of incomplete secretory activation. In vitro differentiation of mammary epithelial cells from Src-/- mice resulted in diminished production of milk proteins compared to the amount of milk proteins produced by Src+/+ cells, indicating a direct role for Src in regulating the transcription/translation of milk protein genes in mammary epithelial cells.

Conclusion: Src is an essential signaling modulator in mammary gland development as Src-/- mice exhibit a block in secretory activation that results in lactation failure and precocious involution. Src appears to be required for increased expression of the prolactin receptor and successful downstream signaling, and alveolar cell organization.

Show MeSH