Cell survival signaling in the bovine mammary gland during the transition from lactation to involution.
In dairy cows, mammary gland involution, and thus a decline in milk production, occurs following peak lactation. To examine the cell signaling pathways regulating involution of the mammary gland, signal transducer and activator of transcription factors (STAT5 and 3), suppressors of cytokine signaling (SOCS1-3 and CIS), insulin-like growth factors (IGF1 and 2), and protein kinase B (Akt) were examined. Mammary involution was induced by termination of milking, and alveolar tissue was collected from 52 nonpregnant, primiparous, mid-lactation Holstein-Friesian cows killed at 0, 6, 12, 18, 24, 36, 72, and 192h postmilking. Qualitative immunohistochemistry showed that activated (phosphorylated) STAT5-P was localized in nuclei of mammary epithelial cells at the early time points, with detection levels decreasing by 24h postmilking. In contrast, STAT3-P was barely detectable at the early time points, with detection levels increasing following longer postmilking periods. This was supported by Western analysis, which showed a decline in STAT5 and STAT5-P protein levels by 24h postmilking, no change in STAT3 levels, and an increase in STAT3-P protein (barely detectable at the early time points) by 72h postmilking. Quantitative real-time reverse transcription PCR analysis showed SOCS1 and SOCS3 mRNA increased by 72h postmilking compared with 6h postmilking. The SOCS2 mRNA remained unchanged across the time series, whereas CIS decreased by 18h postmilking and remained lower compared with that at 6h postmilking until 72h postmilking. The IGF1 mRNA increased by 192h postmilking, whereas IGF2 mRNA decreased by 18h postmilking compared with 6h postmilking. The IGFBP5 mRNA and protein levels of Akt and Akt-P remained unchanged over the time series. These results show that reciprocal activation of STAT5 and STAT3 occurs at the onset of mammary gland involution in the bovine, albeit at a slower rate than in rodents. Mathematical modeling of the pathways indicated that activated STAT3 could block the STAT5 pathway by upregulating SOCS3. The regulation of IGF1-Akt signaling suggests that by 192h postmilking in dairy cows, the involution process is still in the reversible phase, with quiescent mammary epithelial cells not yet in the senescent phase.
AgResearch Ltd., Ruakura Research Centre, Private Bag 3123, Hamilton 3214, New Zealand. Electronic address: kuljeet.singhparhar@outlook.com.
K
K
Singh
AgResearch Ltd., Ruakura Research Centre, Private Bag 3123, Hamilton 3214, New Zealand. Electronic address: kumar.vetharaniam@agresearch.co.nz.
I
I
Vetharaniam
AgResearch Ltd., Ruakura Research Centre, Private Bag 3123, Hamilton 3214, New Zealand.
J M
JM
Dobson
AgResearch Ltd., Ruakura Research Centre, Private Bag 3123, Hamilton 3214, New Zealand.
M
M
Prewitz
AgResearch Ltd., Ruakura Research Centre, Private Bag 3123, Hamilton 3214, New Zealand.
K
K
Oden
AgResearch Ltd., Ruakura Research Centre, Private Bag 3123, Hamilton 3214, New Zealand.
R
R
Murney
AgResearch Ltd., Ruakura Research Centre, Private Bag 3123, Hamilton 3214, New Zealand.
K M
KM
Swanson
AgResearch Ltd., Ruakura Research Centre, Private Bag 3123, Hamilton 3214, New Zealand.
R
R
McDonald
AgResearch Ltd., Ruakura Research Centre, Private Bag 3123, Hamilton 3214, New Zealand.
H V
HV
Henderson
SciLactis Ltd., Waikato Innovation Park, Ruakura Road, Hamilton 3214, New Zealand.
K
K
Stelwagen
D000818:Animals; D002417:Cattle / Q000235:genetics / Q000502:physiology*; D002470:Cell Survival; D005260:Female; D007774:Lactation; D008321:Mammary Glands, Animal / Q000502:physiology*; D050796:STAT3 Transcription Factor / Q000235:genetics / Q000378:metabolism; D050799:STAT5 Transcription Factor / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D000071223:Suppressor of Cytokine Signaling 3 Protein / Q000235:genetics / Q000378:metabolism
D050796:STAT3 Transcription Factor; D050799:STAT5 Transcription Factor; D000071223:Suppressor of Cytokine Signaling 3 Protein
cell survival; dairy cow; mammary gland involution; signal transducer and activator of transcription factors