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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.

Show MeSH
Pups suckled by Src knockout mice have dramatically reduced weight gain. Graphs demonstrating the weight gain of pups foster nursed by Src-/- mice compared to the weight gain of pups nursed by control dams. Litters were normalized to eight pups. N = 3 for the control mice and error bars depict the s.e.m. in the growth of these control litters. The start of each graph represents day 1 of lactation (first day post-partum) for the nursing mother. The weights of pups nursed by Src-/- mice were recorded for 9 days. A) Weight gain of a single litter of eight newborn FVB pups fostered by Src-/- mouse in comparison to three litters of FVB pups nursed by FVB dams. B) Weight gain of a single litter of eight C57Bl6 pups that were 8 days old when they were cross-fostered onto a Src-/- mouse that was one day post-partum. The growth rate is compared to three litters of C57Bl6 pups nursed by control C57Bl6 dams. C) Weight gain by a single litter of 6-day old Src+/- pups cross-fostered onto a Src-/- mouse that was one day post-partum in comparison to three litters nursed by Src+/- dams.
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Figure 3: Pups suckled by Src knockout mice have dramatically reduced weight gain. Graphs demonstrating the weight gain of pups foster nursed by Src-/- mice compared to the weight gain of pups nursed by control dams. Litters were normalized to eight pups. N = 3 for the control mice and error bars depict the s.e.m. in the growth of these control litters. The start of each graph represents day 1 of lactation (first day post-partum) for the nursing mother. The weights of pups nursed by Src-/- mice were recorded for 9 days. A) Weight gain of a single litter of eight newborn FVB pups fostered by Src-/- mouse in comparison to three litters of FVB pups nursed by FVB dams. B) Weight gain of a single litter of eight C57Bl6 pups that were 8 days old when they were cross-fostered onto a Src-/- mouse that was one day post-partum. The growth rate is compared to three litters of C57Bl6 pups nursed by control C57Bl6 dams. C) Weight gain by a single litter of 6-day old Src+/- pups cross-fostered onto a Src-/- mouse that was one day post-partum in comparison to three litters nursed by Src+/- dams.

Mentions: Secretory activation occurs following parturition and leads to the production of copious amounts of milk required to nourish pups. The first indication that Src is an important mediator of mammary gland development was observed when the Src knockout mice were unable to support growth and survival of their pups. The litters born to Src-/- dams were small and over 90% of the pups died within three days. In order to assess the apparent lactation failure of the Src-/- mice, average pup weights during the first 9 days of lactation were measured. The pups from the Src-/- mice were replaced with eight newborn pups from wildtype FVB, C57Bl6, or Src+/- mothers (Figure 3, panels A, B and C, respectively). Due to the poor growth and recurrent death of these foster pups, resulting in the death of over 80% of the fostered litters, we also tried replacing the pups of the Src-/- dams with older pups that were "experienced" at nursing. In the course of the experiments to characterize the lactation potential of the Src-/- dams only three fostered litters nursed by the Src-/- dams survived. The pups in each of these three litters had reduced weight gain over the nine days of lactation analyzed compared to litters nursed by control mothers (Figure 3). In Figure 3A eight newborn FVB pups were nursed by a Src-/- mouse. For the first few days the FVB pups showed very little weight gain then after day 4 their weight steadily increased and after nine days of lactation the average pup weight was 2.8 grams, which is much reduced compared to the 4.7 grams for FVB pups nursed by an FVB mother (Figure 3A). When eight day old C57Bl6 pups were cross-fostered onto a Src-/- mouse, in Figure 3B, the growth of these pups was severely diminished compared to those nursed by control dams (Figure 3B). Indeed on day 16, when the cross-fostered pups had been nursed by the Src-/- mouse for 9 days, the average pup weight was 3.7 grams compared to 5.8 grams for the control C57Bl6 pups nursed by a C57Bl6 dam. The 6 day old pups from a Src+/- litter that were fostered by the Src-/- mouse in Figure 3C demonstrated the least detrimental effect on their growth. When they were 9 days old (day 4 of lactation for the Src-/- mouse) the average pup weight was 3.7 grams compared to 4.5 grams for those nursed by Src+/- dams. For each of these studies the growth of pups nursed by Src-/- dams was compared to 3 litters of pups nursed by control dams (FVB, C57Bl6 and Src+/-). Due to the high mortality of pups nursed by Src-/- dams there were no additional litters to include in this analysis. These results demonstrate that Src knockout mice have diminished lactation capacity regardless of the strain or genotype of the pups nursed and therefore support the conclusion that Src is essential for secretory activation and/or lactation.


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)

Pups suckled by Src knockout mice have dramatically reduced weight gain. Graphs demonstrating the weight gain of pups foster nursed by Src-/- mice compared to the weight gain of pups nursed by control dams. Litters were normalized to eight pups. N = 3 for the control mice and error bars depict the s.e.m. in the growth of these control litters. The start of each graph represents day 1 of lactation (first day post-partum) for the nursing mother. The weights of pups nursed by Src-/- mice were recorded for 9 days. A) Weight gain of a single litter of eight newborn FVB pups fostered by Src-/- mouse in comparison to three litters of FVB pups nursed by FVB dams. B) Weight gain of a single litter of eight C57Bl6 pups that were 8 days old when they were cross-fostered onto a Src-/- mouse that was one day post-partum. The growth rate is compared to three litters of C57Bl6 pups nursed by control C57Bl6 dams. C) Weight gain by a single litter of 6-day old Src+/- pups cross-fostered onto a Src-/- mouse that was one day post-partum in comparison to three litters nursed by Src+/- dams.
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Figure 3: Pups suckled by Src knockout mice have dramatically reduced weight gain. Graphs demonstrating the weight gain of pups foster nursed by Src-/- mice compared to the weight gain of pups nursed by control dams. Litters were normalized to eight pups. N = 3 for the control mice and error bars depict the s.e.m. in the growth of these control litters. The start of each graph represents day 1 of lactation (first day post-partum) for the nursing mother. The weights of pups nursed by Src-/- mice were recorded for 9 days. A) Weight gain of a single litter of eight newborn FVB pups fostered by Src-/- mouse in comparison to three litters of FVB pups nursed by FVB dams. B) Weight gain of a single litter of eight C57Bl6 pups that were 8 days old when they were cross-fostered onto a Src-/- mouse that was one day post-partum. The growth rate is compared to three litters of C57Bl6 pups nursed by control C57Bl6 dams. C) Weight gain by a single litter of 6-day old Src+/- pups cross-fostered onto a Src-/- mouse that was one day post-partum in comparison to three litters nursed by Src+/- dams.
Mentions: Secretory activation occurs following parturition and leads to the production of copious amounts of milk required to nourish pups. The first indication that Src is an important mediator of mammary gland development was observed when the Src knockout mice were unable to support growth and survival of their pups. The litters born to Src-/- dams were small and over 90% of the pups died within three days. In order to assess the apparent lactation failure of the Src-/- mice, average pup weights during the first 9 days of lactation were measured. The pups from the Src-/- mice were replaced with eight newborn pups from wildtype FVB, C57Bl6, or Src+/- mothers (Figure 3, panels A, B and C, respectively). Due to the poor growth and recurrent death of these foster pups, resulting in the death of over 80% of the fostered litters, we also tried replacing the pups of the Src-/- dams with older pups that were "experienced" at nursing. In the course of the experiments to characterize the lactation potential of the Src-/- dams only three fostered litters nursed by the Src-/- dams survived. The pups in each of these three litters had reduced weight gain over the nine days of lactation analyzed compared to litters nursed by control mothers (Figure 3). In Figure 3A eight newborn FVB pups were nursed by a Src-/- mouse. For the first few days the FVB pups showed very little weight gain then after day 4 their weight steadily increased and after nine days of lactation the average pup weight was 2.8 grams, which is much reduced compared to the 4.7 grams for FVB pups nursed by an FVB mother (Figure 3A). When eight day old C57Bl6 pups were cross-fostered onto a Src-/- mouse, in Figure 3B, the growth of these pups was severely diminished compared to those nursed by control dams (Figure 3B). Indeed on day 16, when the cross-fostered pups had been nursed by the Src-/- mouse for 9 days, the average pup weight was 3.7 grams compared to 5.8 grams for the control C57Bl6 pups nursed by a C57Bl6 dam. The 6 day old pups from a Src+/- litter that were fostered by the Src-/- mouse in Figure 3C demonstrated the least detrimental effect on their growth. When they were 9 days old (day 4 of lactation for the Src-/- mouse) the average pup weight was 3.7 grams compared to 4.5 grams for those nursed by Src+/- dams. For each of these studies the growth of pups nursed by Src-/- dams was compared to 3 litters of pups nursed by control dams (FVB, C57Bl6 and Src+/-). Due to the high mortality of pups nursed by Src-/- dams there were no additional litters to include in this analysis. These results demonstrate that Src knockout mice have diminished lactation capacity regardless of the strain or genotype of the pups nursed and therefore support the conclusion that Src is essential for secretory activation and/or lactation.

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