Mastitis, an inflammatory reaction of the mammary gland that is usually caused by a microbial infection, is recognized as the most costly disease in dairy cattle. Decreased milk production accounts for approximately 70% of the total cost of mastitis. Mammary tissue damage reduces the number and activity of epithelial cells and consequently contributes to decreased milk production. Mammary tissue damage has been shown to be induced by either apoptosis or necrosis. These 2 distinct types of cell death can be distinguished by morphological, biochemical, and molecular changes in dying cells. Both bacterial factors and host immune reactions contribute to epithelial tissue damage. During infection of the mammary glands, the tissue damage can initially be caused by bacteria and their products. Certain bacteria produce toxins that destroy cell membranes and damage milk-producing tissue, whereas other bacteria are able to invade and multiply within the bovine mammary epithelial cells before causing cell death. In addition, mastitis is characterized by an influx of somatic cells, primarily polymorphonuclear neutrophils, into the mammary gland. With more immune cells migrating into the mammary gland and the breakdown of the blood-milk barrier, damage to the mammary epithelium worsens. It is well known that breakdown of the extracellular matrix can lead to death of the epithelial cells. Meanwhile, polymorphonuclear neutrophils can harm the mammary tissue by releasing reactive oxygen intermediates and proteolytic enzymes. In vitro and in vivo studies suggest that the use of antioxidants and other protective compounds in mastitis control programs is worth investigating, because they may aid in alleviating damage to secretory cells and thus reduce subsequent milk loss.
The periparturient period is marked by metabolic, hormonal, and immunological changes, which have an effect on the incidence of infectious and metabolic diseases. In a previous study, a slower increase in milk production was induced by milking cows once daily during the first week of lactation, leading to an improvement in levels of several metabolites, including nonesterified fatty acids (NEFA) and β-hydroxybutyrate (BHBA). The aim was to determine the influence of serum collected on d 2, 5, and 61 postpartum from cows milked once or twice daily on immune cell functions and to determine which of the constituents were responsible for these effects. Peripheral blood mononuclear cells (PBMC) and polymorphonuclear leukocytes were collected from healthy midlactation cows and their immune functions (i.e., proliferation and interferon-γ production and chemotaxis, phagocytosis, and oxidative burst, respectively), were evaluated in presence of serum, NEFA, and BHBA. Proliferation of PBMC was greater with d-61 (65.1±1.6%) serum than with d-2 (37.3±2.4%) or d-5 (48.4±1.6%) serum and greater with d-2 and -5 serum from cows milked once (42.2±3.7 and 54.0±2.5) compared with cows milked twice daily (32.4±3.0 and 42.9±2.1). Proliferation was inversely correlated with the concentration of NEFA and BHBA in the serum (r=-0.86). Adding NEFA to d-61 serum to reach the level present in d-5 serum decreased proliferation to the level observed with d-5 serum. No effect of BHBA addition was observed. The release of interferon-γ by PBMC was lower in d-5 serum (766±63 pg/mL) than in d-61 serum (1,187±90 pg/mL) and by NEFA. Milking frequency did not affect chemotaxis, phagocytosis, or oxidative burst of polymorphonuclear leukocytes. Phagocytosis decreased over time in serum from d 2 to 61. Similarly, oxidative burst was greater with d-5 serum (12.7×10(8) ± 1.6×10(8) relative light units) than with d-61 serum (9.0×10(8) ± 1.6×10(8) relative light units). The NEFA had a negative effect on oxidative burst, but BHBA did not. In conclusion, several immune cell functions appear affected by the NEFA concentration. Therefore, strategies that prevent increases in blood NEFA during the transition period may limit postpartum immunosuppression.
Sixteen Holstein cows in mid-lactation were used to determine whether alterations of mammary fatty acid metabolism are responsible for the milk fat depression associated with consumption of fish oil. Cows were given a total mixed ration with no added fish oil (control), unprotected fish oil (3.7% of dry matter), or glutaraldehyde-protected microcapsules of fish oil (1.5% or 3.0% of dry matter) for 4 weeks. Milk samples were taken once a week and a mammary biopsy was taken from a rear quarter at the end of the treatment period. Milk fat content was lower in cows given unprotected fish oil (26.0 g/kg), 1.5% protected fish oil (24.6 g/kg) and 3% protected fish oil (20.4 g/kg) than in cows fed the control diet (36.0 g/kg). This was mainly due to a decrease in the synthesis of short-chain fatty acids. Consumption of protected fish oil decreased the abundance of lipogenic enzymes mRNA in the mammary gland. Acetyl-CoA carboxylase, fatty acid synthase, and stearoyl-CoA desaturase mRNAs for cows given 3% protected fish oil averaged only 30%, 25% and 25% of control values, respectively. Dietary addition of unprotected fish oil slightly decreased mRNA abundance of these enzymes but markedly reduced the amount of lipoprotein lipase mRNA. Milk fat content was significantly correlated with gene expression of acetyl-CoA carboxylase, fatty acid synthase, and stearoyl-CoA desaturase but not lipoprotein lipase. These results suggest that fish oil reduces milk fat percentage by inhibiting gene expression of mammary lipogenic enzymes.
Milk production is a function of the number and activity of mammary epithelial cells, regardless of stage of lactation. Milk yield is generally higher in multiparous cows than in primiparous cows, but persistency is usually greater in the latter group. We compared several measures related to metabolic activity, apoptosis, and endocrine control of mammary cell growth in 8 primiparous and 9 multiparous cows throughout lactation. Mammary gland biopsies were taken in early [10 d in milk (DIM)], peak (50 DIM), and late (250 DIM) lactation to evaluate gene expression and determine DNA and fatty acid synthase (FAS) content. Milk samples taken the day before the biopsies were used to detect protease activities and to determine stanniocalcin-1 (STC) concentrations. Blood samples served to measure insulin-like growth factor-1, prolactin, and STC concentrations. Milk yield was higher in multiparous cows than in primiparous cows at the 10 DIM (32.8 +/- 1.3 and 25.2 +/- 0.8 kg/d) and 50 DIM (38.0 +/- 1.2 and 29.8 +/- 1.1 kg/d), but it was the same for both groups at 250 DIM (23.9 +/- 1.5 and 23.8 +/- 1.1 kg/d). Except for stearoyl-coenzyme A desaturase, expression of genes related to milk synthesis was not affected by stage of lactation. However, gene expression of acetyl-coenzyme A carboxylase, beta-casein, and FAS was lower in early lactation in primiparous cows. Expression of both proapoptotic bax and antiapoptotic bcl-2 genes was higher in primiparous cows, whereas the bax-to-bcl-2 ratio was not changed. Mammary DNA concentration was higher in multiparous cows, as was the amount of FAS protein in early lactation. Two bands of protease activity were found in milk samples, and one of the bands had an apparent molecular weight similar to gelatinase A and was dependent on the stage of lactation. Serum insulin-like growth factor-1 increased with day of lactation and was higher in primiparous cows. Serum prolactin decreased in late lactation, but peak values were observed in early lactation for primiparous cows and peak lactation for multiparous cows. Milk STC content increased with advancing lactation. The results are consistent with a lower degree of differentiation and a greater capacity for cell renewal in the mammary gland of primiparous cows.
The transition from pregnancy to lactation is marked by metabolic, hormonal, and immunological changes that have an impact on the incidence of infectious and metabolic diseases. The aim of this study was to evaluate the effect on immune function and blood metabolite concentration of limiting milk production in early lactation to reduce negative energy balance. Twenty-two multiparous Holstein cows were milked either once a day (1x) or twice a day (2x) for the first week postpartum. All cows were milked twice daily for the rest of lactation. Blood concentrations of nonesterified fatty acids (NEFA), beta-hydroxybutyric acid (BHBA), calcium, bilirubin, urea, phosphorus, glucose, leptin, stanniocalcin-1, and 17beta-estradiol were determined in samples collected from 5 wk before scheduled calving to 5 wk after calving. Polymorphonuclear leukocytes (PMNL) were isolated from blood to conduct assays for chemotaxis, phagocytosis, and respiratory burst. Peripheral blood mononuclear cells (PBMC) were isolated to evaluate lymphocyte proliferation and cytokine production (tumor necrosis factor-alpha, IL-4, and interferon-gamma). Cows milked 1x produced 31% less milk than cows milked 2x during the first week of lactation. Over the following 13 wk of lactation, the milk production of cows milked 1x during the first week was 8.1% lower than for cows milked 2x. However, because the percentages of fat and protein were greater in the milk from 1x cows, the yields of milk components and energy-corrected milk were similar. Calving induced an increase in the concentrations of NEFA, BHBA, urea, and bilirubin. The increases in levels of NEFA and BHBA were greater in cows milked 2x than in cows milked 1x. During the same period, the serum glucose concentration decreased but remained greater in cows milked 1x. Serum calcium on d 4 and serum phosphorus on d 4 and 5 were greater in cows milked 1x. The differences between the 2 groups persisted beyond treatment until postpartum d 24 for NEFA and glucose and until postpartum d 14 for BHBA. After calving, the concentrations of leptin and stanniocalcin-1 decreased. During the first week postpartum, the decrease of leptin was less marked in cows milked 1x. The immune functions of PBMC and PMNL isolated from experimental cows and incubated using a standard medium did not show clear-cut peripartum immunosuppression. These variables were not significantly affected by the treatments, with the exception of interferon-gamma secretion, which was greater on d 5 and 14 in cows milked 1x. In conclusion, limiting milk production in early lactation had positive effects on metabolite concentration, but larger studies are necessary to establish if this could reduce disease incidence.
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