Pegbovigrastim use in periparturient embryo-recipient cows: Effects on health and reproduction.

The objective of this randomized clinical study was to evaluate the effects of periparturient administration of pegbovigrastim (recombinant bovine granulocyte colony stimulating factor; PEG) on postpartum health and reproductive outcomes in dairy cows carrying transferred embryos. A total of 60 multiparous Jersey cows (second parity n = 21, >second parity n = 39) that were in vitro-fertilized embryo recipients were enrolled. Animals were selected from a commercial herd and assigned to 1 of 2 treatments: (1) PEG (n = 31), which received a subcutaneous injection of 15 mg of PEG 7 d before expected calving and a second dose within 1 h of parturition; or (2) CON, untreated control (n = 29). Parturition was induced pharmacologically in all cows at 280 d of pregnancy with 500 µg of cloprostenol and 25 mg of dexamethasone intramuscularly. Cow health was monitored daily during the periparturient period and included assessment for retained fetal membranes within 24 h after calving and metritis until 20 d in milk. Descriptive statistics were calculated, to determine the incidence of postpartum diseases, causes of culling, and early-lactation reproductive outcomes, including number of inseminations, days to first heat, days to first service, and interval from calving to conception. The Wilcoxon rank sum test was used to assess treatment effects on early-lactation reproductive performance outcomes. The overall incidence of metritis in this cohort of cows was 17.14%. Animals treated with PEG tended to have more cases of metritis (25%) compared with CON (8.8%). Mastitis incidence in PEG was 19.35% compared with CON at 27.59%. These data suggest that in this dairy, with embryo-recipient cows induced to calve at 280 d of pregnancy, the administration of 2 doses of PEG tended to increase postpartum metritis but had no effect on mastitis incidence, culling rate, or early-lactation reproductive outcomes.

Oxylipids are associated with higher disease risk in postpartum cows.

Postpartum diseases are a major animal welfare and economic concern for dairy producers. Dysregulated inflammation, which may begin as soon as the cessation of lactation, contributes to the development of postpartum diseases. The ability to regulate inflammation and mitigate postpartum health diseases relies, in part, on the production of inflammatory mediators known as oxylipids. The objective of this study was to examine associations between oxylipids and postpartum diseases. Plasma samples were collected from 16 cattle via coccygeal venipuncture at the following time points: 6 d before dry-off; dry-off (d 0); 1, 2, 6, and 12 d after dry-off; 14 ± 3 d before the expected calving date; and 7 ± 2 d after calving. After calving, cows were grouped according to if clinical disease was undetected throughout the sampling period (n = 7) or if they developed a disease postpartum (n = 9). Liquid chromatography-tandem mass spectrometry was used to analyze plasma concentrations of 63 oxylipid species. Of the 32 oxylipids detected, concentrations of 7 differed between cows with no detected disease and diseased cows throughout the sampling period. Thus, a variable oxylipid profile was demonstrated through 2 major physiological transitions of a lactation cycle. Further, the information gained from this pilot study using a small number of animals with diverse diseases from a single herd suggests that it may be possible to use oxylipids at early mammary involution to alert dairy producers of cows at risk for disease after calving. Future studies should be performed in larger populations of animals, including cows from diverse geographies and dairying styles, and focus on specific diseases to evaluate the utility of oxylipids as biomarkers. Furthermore, it is important to determine the clinical implications of variable oxylipid concentrations throughout the lactation cycle and if the oxylipid profile can be modulated to improve inflammatory outcomes.

Dynamics of lipid droplets in the endometrium and fatty acids and oxylipins in the uterine lumen, blood, and milk of lactating cows during diestrus.

Our objective was to investigate the lipid content of uterus, blood plasma, and milk at early, mid, and late diestrus. Lactating cows (n = 30) had the estrous cycle and ovulation synchronized by administration of exogenous hormones. Cows were blocked by parity and assigned randomly to receive transcervical uterine flushing and biopsy on d 5 (early diestrus), 10 (mid diestrus) or 15 (late diestrus) of the estrous cycle. Flushing and endometrial biopsy were performed in the uterine horn ipsilateral to the corpus luteum. The recovered flushing was used for analyses of lipid composition by liquid chromatography-tandem mass spectrometry and the biopsy was used for investigation of lipid droplet abundance in endometrial cryosections using a neutral lipid fluorescent dye. In addition, blood and milk samples were collected from all cows on d 5, 10, and 15. All blood samples were used to measure the concentration of progesterone in plasma, and all milk samples were used to determine milk composition. Subsamples of blood plasma and milk were also used to evaluate the composition of fatty acids and oxylipins using the same methodology used for uterine flushing samples. The abundance of lipid droplets in the endometrium increased 1.9-fold from d 5 to 10, and 2-fold from d 10 to 15. Concentration of long-chain fatty acids and oxylipins in uterine flushing were, on average, 2.2 and 2.5 times greater in samples collected on d 15 compared with those collected on d 5 and 10. These differences were not observed in blood and milk, suggesting that accumulation of fatty acids and oxylipins in the uterus is regulated locally. In addition to concentration, the profile of individual fatty acids and oxylipins in uterine lumen changed substantially during diestrus. The main categories with increased abundance at late diestrus were mono- and polyunsaturated fatty acids, and oxylipins derived from arachidonic acid, dihomo-γ-linolenic acid, and docosahexaenoic acid. In conclusion, fatty acids and oxylipins accumulate in the uterine lumen during diestrus and might work as a mechanism to supply these lipids to the developing conceptus at late diestrus, when the onset of elongation occurs and substantial synthesis of biomass and cell signaling by lipid mediators are required.

Diet starch concentration and starch fermentability affect markers of inflammatory response and oxidant status in dairy cows during the early postpartum period.

Our objective was to evaluate the effects of diet starch concentration and starch fermentability on inflammatory response markers and oxidant status during the early postpartum (PP) period and its carryover effects. Fifty-two multiparous Holstein cows were used in a completely randomized block design experiment with a 2 × 2 factorial arrangement of treatments. Treatments were starch concentration and starch fermentability of diets; diets were formulated to 22% (low starch, LS) or 28% (high starch, HS) starch with dry-ground corn (DGC) or high-moisture corn (HMC) as the primary starch source. Treatments were fed from 1 to 23 d PP and then switched to a common diet until 72 d PP to measure carryover (CO) effects. Treatment period (TP) diets were formulated to 22% forage neutral detergent fiber and 17% crude protein. The diet for the CO period was formulated to 20% forage neutral detergent fiber, 17% crude protein, and 29% starch. Coccygeal blood was collected once a week during the TP and every second week during the CO period. Liver and adipose tissue biopsies were performed within 2 d PP and at 20 ± 3 d PP. Blood plasma was analyzed for concentrations of albumin, haptoglobin, reactive oxygen and nitrogen species (RONS), and antioxidant potential (AOP), with lipopolysaccharide-binding protein (LBP) and TNFα evaluated during the TP only. Oxidative stress index (OSi) was calculated as RONS/AOP. Abundance of mRNA from genes involved in inflammation and glucose metabolism in liver and genes involved in lipogenesis in adipose tissue were determined. Data were analyzed separately for the TP and CO periods. During the TP, treatments interacted to affect concentrations of TNFα, haptoglobin, and LBP, with HMC increasing their concentrations for HS (9.38 vs. 7.45 pg/mL, 0.45 vs. 0.37 mg/mL, and 5.94 vs. 4.48 µg/mL, respectively) and decreasing their concentrations for LS (4.76 vs. 12.9 pg/mL, 0.27 vs. 0.41 mg/mL, and 4.30 vs. 5.87 µg/mL, respectively) compared with DGC. Effects of treatments diminished over time for LBP and haptoglobin with no differences by the end of the TP and no main CO effects of treatment for haptoglobin. The opposite treatment interaction was observed for albumin, with HMC tending to decrease its concentration for HS (3.24 vs. 3.34 g/dL) and increase its concentration for LS (3.35 vs. 3.29 g/dL) compared with DGC, with no carryover effect. Feeding DGC increased the OSi during the first week of the TP compared with HMC, with this effect diminishing over time; during the CO period HMC increased OSi for HS and decreased it for LS compared with DGC, with this effect diminishing toward the end of CO. Feeding HMC increased the abundance of genes associated with inflammation and gluconeogenesis in liver for HS and decreased it for LS compared with DGC. Feeding HS increased the mRNA abundance of genes associated with adipose tissue lipogenesis compared with LS. Results during the TP suggest that feeding LS-DGC and HS-HMC elicited a more pronounced inflammatory response and induced an upregulation of genes associated with inflammation and gluconeogenesis in liver, without effects on OSi, but effects on plasma markers of inflammation diminished during the CO period.

Effects of timing of palmitic acid supplementation during early lactation on nutrient digestibility, energy balance, and metabolism of dairy cows.

The objective of our study was to evaluate the effects of timing of palmitic acid (C16:0) supplementation during early lactation on nutrient digestibility, energy intake and balance, and metabolic responses of dairy cows. Fifty-two multiparous cows were used in a randomized complete block design experiment. During the fresh (FR) period (1-24 d in milk) cows were assigned to either a control diet containing no supplemental fat (CON) or a C16:0-supplemented diet [PA; 1.5% of diet dry matter (DM)]. During the peak (PK) period (25-67 d in milk) cows were assigned to either a CON diet or a PA diet (1.5% of diet DM) in a 2 × 2 factorial arrangement of treatments considering the diet that they received during the FR period. During the FR period, compared with CON, PA increased DM digestibility by 3.0 percentage units and neutral detergent fiber (NDF) digestibility by 4.4 percentage units, and the increase in these variables was consistent over time. Although PA did not affect 18-carbon fatty acid (FA) digestibility, it decreased 16-carbon FA digestibility by 10.8 percentage units and total FA digestibility by 4.7 percentage units compared with CON. We observed a tendency for an interaction between treatment and time for total FA digestibility and 16-carbon FA digestibility due to the difference in FA digestibility between PA and CON reducing over time. Compared with CON, PA increased digestible energy intake by 3.9 Mcal/d, metabolizable energy intake by 3.5 Mcal/d, and net energy for lactation intake by 2.5 Mcal/d. The PA diet also increased milk energy output, negative energy balance, and plasma nonesterified fatty acid concentration and reduced plasma insulin concentration. We also observed a tendency for an interaction between treatment and time for energy balance due to cows receiving the PA treatment being in a greater negative energy balance over time. During the PK period, PA increased DM digestibility by 2.9 percentage units and NDF digestibility by 3.5 percentage units compared with CON. Although PA decreased 16-carbon FA digestibility by 7.0 percentage units, PA did not affect 18-carbon FA digestibility or total FA digestibility. Feeding PA during the PK period increased energy intake and milk energy output and did not affect energy balance. In conclusion, feeding a C16:0 supplement to early-lactation cows consistently increased DM and NDF digestibilities and energy intake compared with a control diet containing no supplemental fat. Feeding C16:0 markedly increased milk energy output in both the FR and PK periods but increased negative energy balance only in the FR period.

Influence of adipocyte size and adipose depot on the number of adipose tissue macrophages and the expression of adipokines in dairy cows at the end of pregnancy.

The aim of this study was to determine the number of adipose tissue macrophages (ATM) and the mRNA expression of adipokines [adiponectin (ADIPOQ), leptin (LEP), interleukin 6 (IL6), tumor necrosis factor (TNF), and interleukin 10 (IL10)] in different adipose depots from cows with a variable body condition score (BCS) at the end of the dry period. We hypothesized that the number of ATM and the expression of these adipokines depend on adipocyte size and the anatomical location of the adipose depot. Subcutaneous, omental, mesenteric, perirenal, and intrapelvic adipose tissue samples were taken immediately after euthanasia of 10 Holstein Friesian dairy cows (upcoming parity 2 to 5, age 3.9 ± 1.4 yr; mean ± standard deviation) at the end of pregnancy (actual days of pregnancy at the moment of euthanasia: 269 ± 5 d). During the dry period, all animals received similar diets to meet but not exceed requirements. Five animals were considered to have a normal BCS (2.5-3.5) and 5 animals were considered to be over-conditioned (BCS = 3.75-5). Body weight of the animals at the moment of euthanasia was 717 ± 77 kg. Expression of the different genes was determined by reverse transcription quantitative real-time PCR. Adipocyte size was determined by measuring the area of 100 adipocytes on histological sections. Average adipocyte area was 10,475 ± 1,019, 8,500 ± 780, 10,383 ± 1,227, 11,466 ± 1,039, and 11,087 ± 1,632 µm2 for the subcutaneous, mesenteric, omental, intrapelvic, and perirenal adipose depot, respectively. Immunohistochemistry using anti-bovine CD172a antibodies was performed to determine the proportion of ATM (the number of CD172a-positive cells per 100 adipocytes, given as a percentage). Expression of LEP, IL6, and TNF was positively associated with adipocyte size, whereas no association could be detected between ADIPOQ and IL10 with the size of the adipocytes. The omental adipose depot was especially infiltrated with ATM (1.92 ± 0.55, 1.10 ± 0.33, and 8.28 ± 2.24% for the subcutaneous, mesenteric, and omental adipose depot, respectively). The proportion of ATM was positively associated with the size of the adipocytes in the omental and mesenteric adipose depot. Expression of ADIPOQ, LEP, IL6, TNF, and IL10 differed among depots, which suggests differences in inflammatory characteristics depending on the anatomical location of depots. In conclusion, the results of the present study confirm the adipose tissue as a potential source of inflammatory mediators and demonstrate ATM infiltration, especially in the omental adipose depot.

Antimicrobial treatment of clinical mastitis in the eastern United States: The influence of dairy farmers' mastitis management and treatment behavior and attitudes.

To assess both the behaviors and social variables related to antimicrobial therapy for clinical mastitis, we sent a survey to 1,700 dairy farms in Michigan, Pennsylvania, and Florida in January and February 2013. The survey included questions related to 7 major areas: sociodemographic and farm characteristics, milking proficiency, milking systems, cow environment, infected cow monitoring and treatment, farm labor, and attitudes toward mastitis and related antimicrobial use. The overall response rate was 41% (21% in Florida, 39% in Michigan, and 45% in Pennsylvania). Herd size ranged from 9 to 5,800 cows. Only a small proportion of herds frequently or always cultured milk samples for bacteriology from cows with a high somatic cell count (17%), cows with clinical mastitis (18%), or bulk tank milk (13%). Likewise, only 56% of herds frequently or always maintained records of all treated cows and 49% reviewed records before administering mastitis treatments. Multivariate analysis determined that use of treatment records was associated with increased likelihood of frequent use for both intramammary (IMA) and systemic (SYA) administration of antimicrobial drugs for therapy of clinical mastitis. As would be expected, use of natural (organic) therapies was associated with decreased use of IMA, as was the respondent being a member of an Amish community. Lower levels of education and the use of bacterins to control Staphylococcus aureus mastitis were also associated with decreased IMA, whereas increased use of IMA at dry off and the belief that "bad luck" plays a role in mastitis problems were associated with increased IMA. Use of an internal teat sealant, the respondent being the sole proprietor, being from Michigan, use of conductivity to measure subclinical mastitis, the respondent placing increasing importance on decreasing antibiotic residues in cull cows, and having financial incentives for employees linked to somatic cell count were associated with increased use of SYA for the treatment of clinical mastitis. Use of sand or mattresses for bedding were associated with decreased SYA. These findings highlight the need to improve the acceptance of practices that are consistent with prudent antimicrobial use for the treatment of clinical mastitis on dairy farms. Additionally, the willingness of dairy farmers to administer antimicrobial drugs for the treatment of clinical mastitis is associated with other mastitis-related practices and attitudes.

Cultural lag: A new challenge for mastitis control on dairy farms in the United States.

Recent changes in the US dairy industry include increases in herd size and the proportion of milk that is produced by large herds. These changes have been accompanied by an increased reliance on hired employees and an increasing role of immigrant labor to perform critical tasks such as milking cows. Thus, there is a growing need for training and education programs for dairy employees because many employees lack previous dairy experience and employee turnover rates are problematic on many farms. Although extension programs have played an important role in the education and support of dairy producers and allied professionals in attaining improved milk quality, dairy employees have limited access to educational programs. Additionally, metrics to assess employee learning are not validated and the ability to sustain work-related behavioral change has not been well described. In this article, we propose a model that may further our understanding of communication and cultural barriers between dairy managers and employees, based on a demonstration project in 12 Michigan dairy herds. As part of this demonstration, a pilot survey was tested to assess the management culture on dairy farms. Results from this survey found that only 23% of employees across all herds were able to meet with farm management on a regular basis, 36% of employees did not know somatic cell count goals for the farm for which they worked, and 71% of employees stated they primarily received training on milking protocols by other employees or that they learned on their own. Latino employees were more likely to not know farm goals or receive primary training on milking protocols from other employees or on their own compared with their English-speaking counterparts. The survey information, along with input from focus group discussions with participating dairy producers, veterinarians, and employees, suggests that extension needs to build capacity for on-farm training and education for employees to support their engagement within dairy operations.

Herd management and social variables associated with bulk tank somatic cell count in dairy herds in the eastern United States.

The ability to reduce somatic cell counts (SCC) and improve milk quality depends on the effective and consistent application of established mastitis control practices. The US dairy industry continues to rely more on nonfamily labor to perform critical tasks to maintain milk quality. Thus, it is important to understand dairy producer attitudes and beliefs relative to management practices, as well as employee performance, to advance milk quality within the changing structure of the dairy industry. To assess the adoption rate of mastitis control practices in United States dairy herds, as well as assess social variables, including attitudes toward employees relative to mastitis control, a survey was sent to 1,700 dairy farms in Michigan, Pennsylvania, and Florida in January and February of 2013. The survey included questions related to 7 major areas: sociodemographics and farm characteristics, milking proficiency, milking systems, cow environment, infected cow monitoring and treatment, farm labor, and attitudes toward mastitis and related antimicrobial use. The overall response rate was 41% (21% in Florida, 39% in Michigan, and 45% in Pennsylvania). Herd size ranged from 9 to 5,800 cows. Self-reported 3-mo geometric mean bulk tank SCC (BTSCC) for all states was 194,000 cells/mL. Multivariate analysis determined that proven mastitis control practices such as the use of internal teat sealants and blanket dry cow therapy, and not using water during udder preparation before milking, were associated with lower BTSCC. Additionally, farmer and manager beliefs and attitudes, including the perception of mastitis problems and the threshold of concern if BTSCC is above 300,000 cells/mL, were associated with BTSCC. Ensuring strict compliance with milking protocols, giving employees a financial or other penalty if BTSCC increased, and a perceived importance of reducing labor costs were negatively associated with BTSCC in farms with nonfamily employees. These findings highlight the need for a comprehensive approach to managing mastitis, one that includes the human dimensions of management to maintain the practice of scientifically validated mastitis control practices.

Forkhead box protein 3(+) regulatory T cells and Helios(+) subset in perinatally acquired HIV.

Forkhead box protein 3 (FoxP3)(+) regulatory T cells (Tregs ) are important not only in regulating the development of autoimmune conditions, but also in chronic infectious diseases. Given their cardinal function in suppressing immune activation, research has focused upon whether they play a detrimental role in chronic infections, particularly HIV. While the role of Tregs in HIV has been investigated intensively, it remains an unresolved topic. However, it is generally accepted that Tregs are susceptible to HIV infection and are preferentially preserved over conventional CD4(+) T cells. It is unknown whether the peripheral-induced or the thymic-derived Tregs are more susceptible to HIV cytotoxicity. It has been recognized that Tregs can be segregated into two subsets based on Helios expression, with the vast majority being Helios(+) . This study examines the impact of HIV infection on total Tregs and their Helios subsets in a perinatal-acquired HIV-infected paediatric population. The finding indicates a selective expansion or survival of Tregs in association with CD4 depletion and increased viraemia. The Helios(+) and Helios(-) subsets within Tregs appear to be equally affected. However, the Helios(+) Tregs seem to be more preserved in patients with low CD4(+) ≤ 25% and detectable plasma HIV RNA >20 copies/ml. In this group, the frequencies of Tregs are increased, but their numbers appear insufficient to restrain immune activation. In conclusion, our findings suggest that both Helios subsets of Tregs are susceptible to HIV infection and are preferentially preserved compared to conventional CD4(+) T cells.

Association between polyunsaturated fatty acid-derived oxylipid biosynthesis and leukocyte inflammatory marker expression in periparturient dairy cows.

Peripheral blood mononuclear leukocytes from periparturient cows can have exacerbated inflammatory responses that contribute to disease incidence and severity. Oxylipids derived from the oxygenation of polyunsaturated fatty acids (PUFA) can regulate the magnitude and duration of inflammation. Although PUFA substrate for oxylipid biosynthesis in leukocytes is known to change across the periparturient period, the plasma oxylipid profile and how this profile relates to leukocyte inflammatory phenotype is not clear. The objective of this study was to determine if a relationship exists between the profile of pro- and antiinflammatory plasma oxylipids and the inflammatory phenotype of peripheral blood leukocytes during the periparturient period. Seven multiparous Holsteins were sampled from the prepartum period through peak lactation. Plasma oxylipids were measured by liquid chromatography-mass spectrometry, peripheral leukocyte mRNA expression was measured by quantitative PCR, and PUFA content of peripheral blood mononuclear cells was measured by gas chromatography-mass spectrometry. Concentrations of several hydroxyl products of linoleic and arachidonic acid changed over time. Linoleic acid and arachidonic acid concentrations in leukocytes increased during early lactation, suggesting that substrate availability for hydroxyoctadecadienoic and hydroxyeicosatetraenoic acid biosynthesis may influence the oxylipid profile. Leukocyte mRNA expressions of IL-12B, IL-1B, inducible nitric oxide synthase 2, and cyclooxygenase 2 were correlated with several plasma oxylipids. These are the first observations linking leukocyte inflammatory gene responses to shifts in oxylipid biosynthesis in periparturient dairy cows.

Mononuclear leukocyte fatty acid composition and inflammatory phenotype in periparturient and lactating sows.

Increased plasma NEFA concentrations and compromised immune responses are associated with increased disease susceptibility during farrowing and lactation. Increased plasma NEFA concentrations cause changes in the fatty acid (FA) content of plasma lipid fractions and peripheral blood mononuclear cells (PBMC) that could modify inflammatory responses. The goals of this study were to describe changes in plasma lipid composition and to characterize the FA content and proinflammatory phenotype of PBMC in periparturient and lactating sows. Blood samples from 10 sows were collected at 2 wk prefarrow, at 2 d after farrowing (hereafter referred to as farrowing), and at 18 d of lactation (hereafter referred to as lactation). Total lipids and lipid fractions were extracted from plasma and PBMC. Isolated PBMC also were assessed for gene expression of proinflammatory cytokines and enzymes involved in lipid mediator biosynthesis using quantitative PCR. The FA profile of plasma NEFA, phospholipids, neutral lipids, and PBMC phospholipids differed from the composition of total lipids in plasma. At farrowing and lactation, the proportion of palmitic and stearic acids increased (P<0.05) in the plasma NEFA and phospholipid fractions in comparison with prefarrowing concentrations. At the same time, the concentration of palmitic and linoleic acids increased (P<0.05) in the PBMC phospholipid fraction. Omega-3 FA, including docosapentaenoic and docosahexaenoic, increased (P<0.05) at farrowing in plasma and PBMC phospholipids compared with prefarrowing and lactation. Gene expression of IL-1ß, IL-6, IL-8, and tumor necrosis factor-α (TNFα) decreased (P<0.05) after farrowing and in lactation. Similarly, cyclooxygenase expression was reduced during lactation when compared with farrowing (P<0.05). This study demonstrated changes in FA composition of serum lipid fractions and PBMC cellular membranes. Furthermore, it provided an initial assessment of inflammatory responses in mononuclear cells as a function of plasma and PBMC content of saturated and omega-3 FA. Future studies need to address the effect of increased NEFA concentrations, the main hallmark of lipid mobilization, and changes in plasma and cellular lipid profiles on immune function.

Enhanced n-3 phospholipid content reduces inflammatory responses in bovine endothelial cells.

Uncontrolled inflammation contributes to the increased incidence and severity of infectious diseases in periparturient dairy cattle. The objective of this study was to determine if increasing n-3 fatty acid (FA) content and altering the profile of vasoactive eicosanoids could attenuate endothelial cell inflammatory responses. Bovine aortic endothelial cells (BAEC) were cultured with free FA mixtures that mimic the plasma NEFA composition during the first week of lactation of dairy cows or with a free FA mixture supplemented with a higher proportion of n-3 FA, including eicosapentaenoic and docosahexaenoic acids. The effects of increasing the docosahexaenoic and eicosapentaenoic acid content of BAEC on the expression of proinflammatory mediators and eicosanoid biosynthesis was assessed. Culturing BAEC with enriched concentrations of n-3 FA decreased the expression of proinflammatory cytokines, adhesion molecules, and reactive oxygen species with a concomitant increase in the biosynthesis of proresolving eicosanoids, including resolvins, protectins, and lipoxins. This study showed for the first time that increasing the n-3 FA content of endothelial cell phospholipids could alter the expression of eicosanoids and control the magnitude of inflammatory responses. Future studies are necessary to elucidate the mechanisms by which resolvins, protectins, and lipoxins may modify endothelial inflammatory pathways necessary to reduce the severity and duration of disease in periparturient cows.

Nonesterified fatty acids modify inflammatory response and eicosanoid biosynthesis in bovine endothelial cells.

Intense lipid mobilization during the transition period in dairy cows is associated with increased disease susceptibility. The potential impact of altered plasma nonesterified fatty acids (NEFA) concentrations and composition on host inflammatory responses that may contribute to disease incidence and severity are not known. The objective of this study was to evaluate if increased NEFA concentrations could modify vascular inflammatory responses in vitro by changing the expression of important inflammatory mediators that are important in the pathogenesis of infectious diseases of transition cows such as mastitis and metritis. Bovine aortic endothelial cells (BAEC) were cultured with different concentrations of a NEFA mixture that reflected the plasma NEFA composition during different stages of lactation. The expression of cytokines, adhesion molecules, and eicosanoids were measured to assess changes in BAEC inflammatory phenotype. Addition of NEFA mixtures altered the fatty acid profile of BAEC by increasing the concentration of stearic acid (C18:0) and decreasing the content of arachidonic acid (C20:4n6c) and other long-chain polyunsaturated fatty acids in the phospholipid fraction. A significant increase also occurred in mRNA expression of cytokine and adhesion molecules that are associated with increased inflammatory responses during the transition period. Expression of cyclooxygenase 2, an important enzyme associated with eicosanoid biosynthesis, was increased in a NEFA concentration-dependent manner. The production of linoleic acid-derived eicosanoids 9- and 13-hydroxyoctadecadienoic acids also was increased significantly after treatment with NEFA mixtures. This research described for the first time specific changes in vascular inflammatory response during in vitro exposure to NEFA mixtures that mimic the composition and concentration found in cows during the transition period. These findings could explain, in part, alterations in inflammatory responses observed during intense lipid mobilization stages such as in the transition period of dairy cows. Future studies should analyze specific mechanisms by which high NEFA concentrations induce a vascular proinflammatory phenotype including the effect of 9 and 13-hydroxyoctadecadienoic acids and other lipid mediators.

Changes in glucose transporter expression in monocytes of periparturient dairy cows.

The transition period of dairy cows is characterized by dramatic changes in metabolism and immune cell function that contributes to increased susceptibility to several economically important diseases. Monocyte and macrophage populations increase in blood and tissues of cows during the transition period and have enhanced inflammatory responses that may contribute to increased severity of disease. Glucose is a major energy source for activated monocytes and glucose uptake is facilitated by glucose transporters (GLUT). The objective of this study was to determine how bovine monocyte GLUT expression changes during lactogenesis and in response to proinflammatory stimulation. Blood samples were collected from 10 dairy cows approximately 5 wk before calving and during the first week of lactation. Monocytes were isolated from total peripheral blood mononuclear cells, and expression of GLUT1, GLUT3, and GLUT4 isoforms was assessed in resting cells and following endotoxin stimulation. In general, the onset of lactation served to decrease overall GLUT expression. Gene and protein expression of GLUT1 was significantly decreased after parturition, and GLUT3 and GLUT4 cell surface expression was also significantly decreased postcalving. Endotoxin stimulation, however, increased gene expression of GLUT3 and GLUT4, and gene expression for all GLUT isoforms was positively correlated to production of tumor necrosis factor-α. This study identified, for the first time, the presence of GLUT isoforms in bovine monocytes. Alterations in monocyte GLUT expression at the onset of lactation warrant further investigation to ascertain how changes in glucose uptake may contribute to periparturient inflammatory dysfunction.

Ethyl pyruvate diminishes the endotoxin-induced inflammatory response of bovine mammary endothelial cells.

The endotoxin-induced inflammatory response during coliform mastitis is difficult to control with the currently available therapeutics. Endothelial cells are among the first cell type to be engaged in the inflammatory response and can modulate the severity of inflammation by producing proinflammatory mediators upon endotoxin exposure. Ethyl pyruvate, an ethyl ester of pyruvic acid, can ameliorate endotoxin-induced inflammatory responses by inhibiting the production of proinflammatory mediators in several in vitro and in vivo endotoxemia models. The objective of this study was to determine the effect of ethyl pyruvate on the production of vascular proinflammatory mediators that are associated with the pathogenesis of coliform mastitis. The ability of ethyl pyruvate to reduce the expression of proinflammatory mediators was evaluated in cultured bovine mammary endothelial cells (BMEC) stimulated with endotoxin. Treatment of endotoxin-stimulated BMEC with ethyl pyruvate significantly reduced gene expression of IL-6, IL-8, and intercellular adhesion molecule 1 as well as expression of eicosanoid-producing enzymes, including cyclooxygenase 2 and 15-lipoxygenase 1. This is the first time that the effect of ethyl pyruvate was evaluated in an in vitro BMEC model of coliform mastitis. The ability of ethyl pyruvate to effectively inhibit gene and protein expression of potent vascular proinflammatory mediators in vitro warrants further investigations to assess in vivo efficacy. Ethyl pyruvate is safe for human consumption, and it may be an attractive candidate as a therapeutic in ameliorating the severe pathogenesis associated with coliform mastitis.

Management of multidrug-resistant enterococcal infections.

Enterococci are organisms with a remarkable ability to adapt to the environment and acquire antibiotic resistance determinants. The evolution of antimicrobial resistance in these organisms poses enormous challenges for clinicians when faced with patients affected with severe infections. The increased prevalence and dissemination of multidrug-resistant Enterococcus faecium worldwide has resulted in a major decrease in therapeutic options because the majority of E. faecium isolates are now resistant to ampicillin and vancomycin, and exhibit high-level resistance to aminoglycosides, which are three of the traditionally most useful anti-enterococcal antibiotics. Newer antibiotics such as linezolid, daptomycin and tigecycline have good in vitro activity against enterococcal isolates, although their clinical use may be limited in certain clinical scenarios as a result of reduced rates of success, possible underdosing for enterococci and low serum levels, respectively, and also by the emergence of resistance. The experimental agent oritavancin may offer some hope for the treatment of vancomycin-resistant enterococci but clinical data are still lacking. Thus, optimal therapies for the treatment of multidrug-resistant enterococcal infections continue to be based on empirical observations and extrapolations from in vitro and animal data. Clinical studies evaluating new strategies, including combination therapies, to treat severe vancomycin-resistant E. faecium infections are urgently needed.

Lipomobilization in periparturient dairy cows influences the composition of plasma nonesterified fatty acids and leukocyte phospholipid fatty acids.

The periparturient period is characterized by sudden changes in metabolic and immune cell functions that predispose dairy cows to increased incidence of disease. Metabolic changes include alterations in the energy balance that lead to increased lipomobilization with consequent elevation of plasma nonesterified fatty acids (NEFA) concentrations. The objective of this study was to establish the influence of lipomobilization on fatty acid profiles within plasma lipid fractions and leukocyte phospholipid composition. Blood samples from 10 dairy cows were collected at 14 and 7 d before due date, at calving, and at 7, 14, and 30 d after calving. Total lipids and lipid fractions were extracted from plasma and peripheral blood mononuclear cells. The degree of lipomobilization was characterized by measurement of plasma NEFA concentrations. The fatty acid profile of plasma NEFA, plasma phospholipids, and leukocyte phospholipids differed from the composition of total lipids in plasma, where linoleic acid was the most common fatty acid. Around parturition and during early lactation, the proportion of palmitic acid significantly increased in the plasma NEFA and phospholipid fractions with a concomitant increase in the phospholipid fatty acid profile of leukocytes. In contrast, the phospholipid fraction of long-chain polyunsaturated fatty acids in leukocytes was diminished during the periparturient period, especially during the first 2 wk following parturition. This study showed that the composition of total plasma lipids does not necessarily reflect the NEFA and phospholipid fractions in periparturient dairy cows. These findings are significant because it is the plasma phospholipid fraction that contributes to fatty acid composition of membrane phospholipids. Increased availability of certain saturated fatty acids in the NEFA phospholipid fractions may contribute to altered leukocyte functions during the periparturient period.

Lipoxygenase metabolites modulate vascular-derived platelet activating factor production following endotoxin challenge.

Endotoxin promotes the production of potent pro-inflammatory lipid mediators, such as platelet activating factor (PAF) and eicosanoids, which contribute to the pathophysiology of endotoxic shock. Endothelial cells are both a target for and producers of these lipid mediators so it is vital to understand the pathways leading to their production in these cells. Previous research suggested a positive feedback loop between eicosanoids and PAF during endotoxemia. This study sought to determine if eicosanoids derived from the 15-lipoxygenase (15-LOX1) pathway can modulate the biosynthesis of PAF in cultured bovine aortic endothelial cells (BAEC) following endotoxin stimulation. Endotoxin stimulation increased the production of 15-LOX1-derived eicosanoids prior to PAF in primary BAEC. Exogenous addition of specific 15-LOX1 eicosanoids, as well as overexpression of 15-LOX1 in endotoxin-stimulated BAEC, further increased the endotoxin-induced production of PAF. Whereas increased expression of 15-LOX1 activity can further exacerbate endotoxin-induced PAF biosynthesis, inhibition of 15-LOX1 activity is not capable of abrogating the initial onset of endotoxin-induced PAF production. The results indicate that 15-LOX1 activity is not necessary for the initial induction of PAF following endotoxin stimulation. There may exist, however, a role for elevated 15-LOX1 activity in further escalating the extent of PAF biosynthesis in BAEC during endotoxic shock. Determining factors that can potentiate endotoxin-induced vascular dysfunction may lead to the development of novel therapeutic targets to diminish the pathophysiological effects of endotoxic shock.

Metabolic factors affecting the inflammatory response of periparturient dairy cows.

Dairy cattle are susceptible to increased incidence and severity of disease during the periparturient period. Increased health disorders have been associated with alterations in bovine immune mechanisms. Many different aspects of the bovine immune system change during the periparturient period, but uncontrolled inflammation is a dominant factor in several economically important disorders such as metritis and mastitis. In human medicine, the metabolic syndrome is known to trigger several key events that can initiate and promote uncontrolled systemic inflammation. Altered lipid metabolism, increased circulating concentrations of non-esterified fatty acids and oxidative stress are significant contributing factors to systemic inflammation and the development of inflammatory-based diseases in humans. Dairy cows undergo similar metabolic adaptations during the onset of lactation, and it was postulated that some of these physiological events may negatively impact the magnitude and duration of inflammation. This review will discuss how certain types of fatty acids may promote uncontrolled inflammation either directly or through metabolism into potent lipid mediators. The relationship of increased lipid metabolism and oxidative stress to inflammatory dysfunction will be reviewed as well. Understanding more about the underlying cause of periparturient health disorders may facilitate the design of nutritional regimens that will meet the energy requirements of cows during early lactation and reduce the susceptibility to disease as a function of compromised inflammatory responses.