In Vivo Inhibition Followed by Exogenous Supplementation Demonstrates Galactopoietic Effects of Prolactin on Mammary Tissue and Milk Production in Dairy Cows

ABSTRACT: It has been previously shown that the long-term inhibition of milking-induced prolactin (PRL) release by quinagolide (QN), a dopamine agonist, reduces milk yield in dairy cows. To further demonstrate that PRL is galactopoietic in cows, we performed a short-term experiment that used PRL injections to restore the release of PRL at milking in QN-treated cows. Nine Holstein cows were assigned to treatments during three 5-d periods in a 3 × 3 Latin square design: 1) QN: twice-daily i.m. injections of 1 mg of QN; 2) QN-PRL: twice-daily i.m. injections of 1 mg of QN and twice-daily (at milking time) i.v. injections of PRL (2 μg/kg body weight); and 3) control: twice-daily injections of the vehicles. Mammary epithelial cells (MEC) were purified from milk so that their viability could be assessed, and mammary biopsies were harvested for immunohistological analyses of cell proliferation using PCNA and STAT5 staining. In both milk-purified MEC and mammary tissue, the mRNA levels of milk proteins and BAX were determined using real-time reverse-transcription PCR. Daily QN injections reduced milking-induced PRL release. The area under the PRL curve was similar in the control and PRL injection treatments, but the shape was different. The QN treatment decreased milk, lactose, protein, and casein production. Injections of PRL did not restore milk yield but tended to increase milk protein yield. In mammary tissue, the percentage of STAT5-positive cells was reduced during QN but not during QN-PRL in comparison with the control treatment. The percentage of PCNA-positive cells was greater during QN-PRL injections than during the control or QN treatment and tended to be lower during QN than during the control treatment. In milk-purified MEC, κ-casein and α-lactalbumin mRNA levels were lower during QN than during the control treatment, but during QN-PRL, they were not different from the control treatment. In mammary tissue, the BAX mRNA level was lower during QN-PRL than during QN. The number of MEC exfoliated into milk was increased by QN injections but tended to be decreased by PRL injections. Injections of PRL also increased the viability of MEC harvested from milk. Although PRL injections at milking could not reverse the effect of QN treatment on milk production, their effects on cell survival and exfoliation and on gene expression suggest that the effect of QN treatment on the mammary gland is due to QN’s inhibition of PRL secretion

Unilateral once daily milking locally induces differential gene expression in both mammary tissue and milk epithelial cells revealing mammary remodeling

Once daily milking reduces milk yield, but the underlying mechanisms are not yet fully understood. Local regulation due to milk stasis in the tissue may contribute to this effect, but such mechanisms have not yet been fully described. To challenge this hypothesis, one udder half of six Holstein dairy cows was milked once a day (ODM), and the other twice a day (TDM). On the 8th day of unilateral ODM, mammary epithelial cells (MEC) were purified from the milk using immunomagnetic separation. Mammary biopsies were harvested from both udder halves. The differences in transcript profiles between biopsies from ODM and TDM udder halves were analyzed by a 22k bovine oligonucleotide array, revealing 490 transcripts that were differentially expressed. The principal category of upregulated transcripts concerned mechanisms involved in cell proliferation and death. We further confirmed remodeling of the mammary tissue by immunohistochemistry, which showed less cell proliferation and more apoptosis in ODM udder halves. Gene expression analyzed by RT-qPCR in MEC purified from milk and mammary biopsies showed a common downregulation of six transcripts (ABCG2, FABP3, NUCB2, RNASE1 and 5, and SLC34A2) but also some discrepancies. First, none of the upregulated transcripts in biopsies varied in milk-purified MEC. Second, only milk-purified MEC showed significant LALBA downregulation, which suggests therefore that they correspond to a mammary epithelial cell subpopulation. Our results, obtained after unilateral milking, suggest that cell remodeling during ODM is due to a local effect, which may be triggered by milk accumulation

Chronic inhibition of prolactin discharge at milking affects mammary cell activity and turnover in dairy cows

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Methionine supplementation impacts casein gene expression and cell death in the mammary tissue of lactating dairy goats fed low and adequate net energy supplies

International audienceMethionine supplementation is known to modulate milk and protein yields in dairy ruminants. However, the mechanism involved in this phenomenon is still unknown. The cellular and molecular responses in the mammary tissue to methionine (Met) supplementation through HMBi (MetaSmart, Adisseo) at low (LE) and adequate levels (AE) of NEL were investigated according to a 2 × 2 factorial arrangement on 48 multiparous Alpine goats at mid lactation, assigned to a randomized block design in 4 groups during 5 weeks. Each goat was genotyped for CSN1S1 gene to balance each group according to the high or low genotypes for CSN1S1 gene. A fixed amount of hay was distributed per group (1.17 Mcal/kg DM) and a fixed amount of concentrates was distributed individually (1.72 vs. 1.83 Mcal/kg DM in LE and AE, respectively). Metabolizable methionine were 1.95 vs. 2.45% of metabolizable protein in unbalanced (LE and AE) vs. balanced (LEMET and AEMET) diets, respectively. Goats (23) were slaughtered after 5 weeks of treatment. Mammary tissues were collected for the analyses of milk protein mRNA levels by real time RT-PCR and apoptosis by immunohistochemistry using TUNEL assay. A variance analysis was performed with genotype, E, Met and E x Met, E x genotype, Met x genotype interactions as fixed effects and for milk, pre-treatment data yields was used as a covariate. Met supplementation increased milk protein yield (+6.5 g/d; P = 0.03). The treatments did no modify LALBA whereas an interaction E x Met (P = 0.001) were observed for both CSN3 and CSN1S1 mRNA levels showing 2-fold increases with Met supplementation in goats fed AE diet. The supplementation of Met reduced by 51% apoptosis rates in the mammary tissue (P < 0.01). Apoptosis rate was correlated with milk yield (−0.48; P < 0.05). These results suggest that Met increased casein gene expression at AE level and reduced cell death which could partly explain the positive effect of Met supplementation on milk synthesis in the goat mammary tissue

Cell junction disruption after 36h milk accumulation was associated with changes in mammary secretory tissue activity and dynamics in lactating dairy goats

International audienceMilk stasis in the mammary gland is a situation that induces a reduction in milk yield and a change in its composition. Theses changes could be related to a decrease in the number and/or activity of secretory cells. Previous observations showed that a disruption of cell junctions is one of the early phenomena accompanying milk accumulation in the mammary gland. The aim of the present study was to investigate the local effect of 36 h milk accumulation on mammary cells activity, apoptosis and proliferation. We also studied the expression of cell junction proteins after 36 h of milk stasis. We observed a decrease in the quantity of milk produced after 36 h of milk stasis in goat. Lower milk lactose and protein yields were also observed, which was associated with a decrease in the transcripts of genes involved in synthesis of these constituents, such as alpha-lactalbumin and kappa-casein. Mammary gland apoptosis was more intensive than mammary cell proliferation after 36 h of milk accumulation. All these changes were associated with an up-regulation of E-cadherin protein and increase of its transcripts levels. It could suggest that these adjustments are made in order to limit losses of secretory cell number and activity

Tight junction opening after 36h milk accumulation was associated to a change in mammary secretory tissue activity and dynamics in lactating dairy goats

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Caractérisation du processus d’exfoliation des cellules épithéliales mammaires au cours de la traite chez la vache laitière

Caractérisation du processus d’exfoliation des cellules épithéliales mammaires au cours de la traite chez la vache laitière. 23. Rencontres autour des Recherches sur les Ruminant

Changes in mammary secretory tissue during lactation in ovariectomized dairy cows

In dairy animals, the milk yield (MY) changes during a lactation and is influenced by several physiological, livestock management and environmental factors. The MY produced by a mammary gland depends on synthetic activity of mammary epithelial cells (MECs) as well as MEC number and mammary secretory tissue organization. It has been suggested that ovarian steroids (estradiol and progesterone) have a negative effect on MY in lactating cows. In a previous study, we showed that the suppression of ovarian secretions by an ovariectomy improved lactation persistency in dairy cows. Here we were interested in the effects of ovariectomy on plasma estradiol and progesterone concentrations and on changes that occur in mammary secretory tissue during lactation. We demonstrated that the ovariectomy of lactating cows at the time of the lactation peak induced a rapid and dramatic drop in plasma progesterone and a smaller reduction in plasma estradiol. Interestingly, the study of the changes in mammary secretory tissue over time revealed that the improvement of MY measured in the ovariectomized cows was associated with a limited increase in estradiol receptivity in MECs, a reduced mammary tissue remodeling and reduced blood protein concentration in milk, in late lactation. These results suggest that ovarian secretions, particularly estradiol and progesterone, act to enhance processes for mammary gland involution in late-lactating dairy cows. Copyright 2013 Elsevier Ltd. All rights reserved

Inhibiting prolactin by cabergoline accelerates mammary gland remodeling during the early dry period in dairy cows

The inhibition of prolactin release using cabergoline, a dopamine agonist, is an effective strategy to accelerate the changes in mammary secretion composition after drying-off. The objective of this study was to determine how cabergoline may affect mammary tissue remodeling during early involution. Holstein dairy cows were treated with either a single i.m. administration of 5.6 mg of cabergoline (Velactis, Ceva Santé Animale, Libourne, France, n = 7) or placebo (n = 7) at the time of drying-off. Mammary biopsy samples were collected 1 wk before drying-off (d -6), after 30 h of milk accumulation (d 1), and again 8 d following drying-off (d 8) to determine changes in gene expression, lactoferrin content, and cell turnover. Blood and mammary secretion samples were collected at d -6 and again at d 1, 2, 3, 4, 8, and 14 following the abrupt cessation of lactation to evaluate indicators of blood-milk barrier integrity and other markers of mammary tissue remodeling. Cabergoline induced less SLC2A1, BAX, CAPN2, and IGFBP5 mRNA expression. In contrast, cabergoline did not modify changes in cell proliferation and apoptosis. Following the cessation of lactation, changes in mammary secretion composition (Na(+) and K(+)) and blood lactose concentrations were indicative of a loss in the blood-milk barrier function in both treatment groups. Cabergoline treatment affected only Na(+) and K(+) concentrations at d 1, suggesting a moderate increase in tight junction permeability. The increase in the activity of MMP9 and in mammary epithelial cell concentration in mammary secretions was greater in cabergoline-treated cows than in control cows, suggesting more mammary tissue remodeling. The increase in lactoferrin immunostaining in the mammary tissue occurred earlier for cabergoline-treated cows than for control cows, and was essentially localized in the stroma. Changes in some key markers of mammary involution suggest that cabergoline accelerates mammary gland remodeling. Thus, a single injection of cabergoline after the last milking would facilitate drying-off by enhancing mammary gland involution