Comparison between conjugated linoleic acid and essential fatty acids in preventing oxidative stress in bovine mammary epithelial cells

Some in vitro and in vivo studies have demonstrated protective effects of conjugated linoleic acid (CLA) isomers against oxidative stress and lipid peroxidation. However, only a few and conflicting studies have been conducted showing the antioxidant potential of essential fatty acids. The objectives of the study were to compare the effects of CLA to other essential fatty acids on the thiol redox status of bovine mammary epithelia cells (BME-UV1) and their protective role against oxidative damage on the mammary gland by an in vitro study. The BME-UV1 cells were treated with complete medium containing 50 μM of cis-9,trans-11 CLA, trans-10,cis-12 CLA, α-linolenic acid, γ-linolenic acid, and linoleic acid. To assess the cellular antioxidant response, glutathione, NADPH, and γ-glutamyl-cysteine ligase activity were measured 48 h after addition of fatty acids (FA). Intracellular reactive oxygen species and malondialdehyde production were also assessed in cells supplemented with FA. Reactive oxygen species production after 3 h of H2O2 exposure was assessed to evaluate and to compare the potential protection of different FA against H2O2-induced oxidative stress. All FA treatments induced an intracellular GSH increase, matched by high concentrations of NADPH and an increase of γ-glutamyl-cysteine ligase activity. Cells supplemented with FA showed a reduction in intracellular malondialdehyde levels. In particular, CLA isomers and linoleic acid supplementation showed a better antioxidant cellular response against oxidative damage induced by H2O2 compared with other FA

The effect of dietary rumen-protected trans-10,cis-12 conjugated linoleic acid or a milk fat-depressing diet on energy metabolism, inflammation, and oxidative stress of dairy cows in early lactation

The objective of this study was to determine the effects of milk fat depression induced by supplementing conjugated linoleic acid (CLA; trans-10,cis-12 and cis-9,trans-11 CLA) or feeding a higher starch and oil-containing diet (HSO) on metabolic changes in dairy cows after calving. The main hypothesis was that the 2 strategies to decrease milk fat yield could have different effects on performance, energy balance (EB), and inflammatory status in early lactation. Thirty-three Nordic Red dairy cows were used in a randomized block design from 1 to 112 d of lactation and fed one of the following treatments: control (CON), CLA-supplemented diet, or HSO diet. Dry matter intake and milk yield were measured daily whereas milk composition was measured weekly throughout the experiment. Nutrient digestibility, EB, and plasma hormones and metabolites were measured at 3, 7, 11, and 15 wk of lactation in respiration chambers. The HSO diet led to lower intakes of dry matter, neutral detergent fiber, and gross energy compared with CON and CLA diets. The CLA diet and especially the HSO diet resulted in lower energy-corrected milk yield during the first 7 wk of lactation than those fed CON. The EB was numerically higher for HSO and CLA diets compared with CON at wk 3 and 7. Plasma glucose concentration was higher by the CLA diet at wk 3 and by the HSO diet from wk 3 to 15 compared with CON. Plasma nonesterified fatty acids were higher at wk 3 in the CON group (indicating more lipid mobilization) but decreased thereafter to similar levels with the other groups. The HSO-fed cows had higher plasma ceruloplasmin, paraoxonase, and total bilirubin concentrations in the entire experiment and showed the highest levels of reactive oxygen metabolites. These results suggest an increased inflammatory and oxidative stress state in the HSO cows and probably different regulation of the innate immune system. This study provides evidence that milk fat depression induced by feeding HSO (as well as CLA) decreased milk fat secretion and improved EB compared with CON in early lactation. The increase in plasma glucose and paraoxonase levels with the HSO diet may imply a better ability of the liver to cope with the metabolic demand after parturition. However, the negative effect of HSO on feed intake, and the indication of increased inflammatory and oxidative stress warrant further studies before the HSO feeding strategy could be supported as an alternative to improve EB in early lactation

Changes in fatty acids in plasma and association with the inflammatory response in dairy cows abomasally infused with essential fatty acids and conjugated linoleic acid during late and early lactation.

Dairy cows are exposed to increased inflammatory processes in the transition period from late pregnancy to early lactation. Essential fatty acids (EFA) and conjugated linoleic acid (CLA) are thought to modulate the inflammatory response in dairy cows. The present study investigated the effects of a combined EFA and CLA infusion on the fatty acid (FA) status in plasma lipids, and whether changes in the FA pattern were associated with the acute phase and inflammatory response during late pregnancy and early lactation. Rumen-cannulated Holstein cows (n = 40) were assigned from wk 9 antepartum to wk 9 postpartum to 1 of 4 treatment groups. Cows were abomasally supplemented with coconut oil (CTRL, 76 g/d), linseed and safflower oil (EFA, 78 g/d of linseed oil and 4 g/d of safflower oil; ratio of oils = 19.5:1; n-6:n-3 FA ratio = 1:3), Lutalin (CLA, 38 g/d; isomers cis-9,trans-11 and trans-10,cis-12; each 10 g/d), or both (EFA+CLA). Blood samples were taken to measure changes in FA in blood plasma on d -63, -42, 1, 28, and 56, and in plasma lipid fractions (cholesterol esters, free fatty acids, phospholipids, and triglycerides) on d -42, 1, and 56 relative to calving, and in erythrocyte membrane (EM) on d 56 after calving. Traits related to the acute phase response and inflammation were measured in blood throughout the study. Liver samples were obtained for biopsy on d -63, -21, 1, 28, and 63 relative to calving to measure the mRNA abundance of genes related to the inflammatory response. The concentrations of α-linolenic acid and n-3 FA metabolites increased in lipid fractions (especially phospholipids) and EM due to EFA supplementation with higher α-linolenic acid but lower n-3 metabolite concentrations in EFA+CLA than in EFA treatment only. Concentration of linoleic acid decreased in plasma fat toward calving and increased during early lactation in all groups. Concentration of plasma arachidonic acid was lower in EFA- than in non-EFA-treated groups in lipid fractions and EM. The cis-9,trans-11 CLA increased in all lipid fractions and EM after both CLA treatments. Plasma haptoglobin was lowered by EFA treatment before calving. Plasma bilirubin was lower in EFA and CLA than in CTRL at calving. Plasma concentration of IL-1β was higher in EFA than in CTRL and EFA+CLA at certain time points before and after calving. Plasma fibrinogen dropped faster in CLA than in EFA and EFA+CLA on d 14 postpartum. Plasma paraoxonase tended to be elevated by EFA treatment, and was higher in EFA+CLA than in CTRL on d 49. Hepatic mRNA abundance revealed time changes but no treatment effects with respect to the inflammatory response. Our data confirmed the enrichment of n-3 FA in EM by EFA treatment and the inhibition of n-3 FA desaturation by CLA treatment. The elevated n-3 FA status and reduced n-6:n-3 ratio by EFA treatment indicated a more distinct effect on the inflammatory response during the transition period than the single CLA treatment, and the combined EFA+CLA treatment caused minor additional changes on the anti-inflammatory response

Conjugated linoleic acid isomers strongly improve the redox status of bovine mammary epithelial cells (BME-UV1)

Some studies have shown the protective effects of conjugated linoleic acid (CLA) isomers against oxidative stress and lipid peroxidation in animal models, but no information is available about CLA and changes in oxidative status of the bovine mammary gland. The objectives of the study were to assess in vitro the effect of CLA on the cellular antioxidant response of bovine mammary cells, to examine whether CLA isomers could play a role in cell protection against the oxidative stress, and to study the molecular mechanism involved. For the study, BME-UV1 cells, a bovine mammary epithelial cell line, were used as the experimental model. The BME-UV1 cells were treated with complete medium containing 50 μM cis-9,trans-11 CLA (c9,t11 CLA), trans-10,cis-12 CLA (t10,c12 CLA), and CLA mixture (1:1, cis-9,trans-11: trans-10,cis-12 CLA). To monitor cellular uptake of CLA isomers, cells and culture medium were collected at 0, 3, and 48 h from CLA addition for lipid extraction and fatty acid analyses. To assess the cellular antioxidant response, glutathione (GSH/GSSH), NADPH, and γ-glutamyl-cysteine ligase activity was measured after 48 h from addition of CLA. Cytoplasmic superoxide dismutase, glutathione peroxidase, glutathione S-transferase, and glutathione reductase activities and mRNA were also determined. Intracellular reactive oxygen species and thiobarbituric acid reactive substance production were assessed in cells supplemented with CLA isomers. Cell viability after 3 h to H2O2 exposure was assessed to evaluate and to compare the potential protection of different CLA isomers against H2O2-induced oxidative stress. Mammary cells readily picked up all CLA isomers, their accumulation was time dependent, and main metabolites at 48 h are two 18:3 isomers. The CLA treatment induced an intracellular GSH increase, matched by high concentration of NADPH, and an increase of γ-glutamyl-cysteine ligase activity mainly in cells treated with the t10,c12 CLA isomer. The CLA isomer treatment of bovine mammary cells increased superoxide dismutase, glutathione peroxidase, and glutathione S-transferase activity and decreased glutathione reductase activity, but no changes in gene expression of these antioxidant enzymes were observed. Cells supplemented with CLA isomers showed a reduction in intracellular reactive oxygen species and thiobarbituric acid reactive substance levels. All CLA isomers were able to enhance cell resistance against H2O2-induced oxidative stress. These suggest an antioxidant role of CLA, in particular of t10,c12 CLA, by developing a significantly high redox status in cells. © 2015 American Dairy Science Association

Glucose metabolism and the somatotropic axis in dairy cows after abomasal infusion of essential fatty acids together with conjugated linoleic acid during late gestation and early lactation.

Sufficient glucose availability is crucial for exploiting the genetic potential of milk production during early lactation, and endocrine changes are mainly related to repartitioning of nutrient supplies toward the mammary gland. Long-chain fatty acids, such as essential fatty acids (EFA) and conjugated linoleic acid (CLA), have the potential to improve negative energy balance and modify endocrine changes. In the present study, the hypothesis that combined CLA and EFA treatment supports glucose metabolism around the time of calving and stimulates insulin action and the somatotropic axis in cows in an additive manner was tested. Rumen-cannulated German Holstein cows (n = 40) were investigated from wk 9 antepartum (AP) until wk 9 postpartum (PP). The cows were abomasally supplemented with coconut oil (CTRL, 76 g/d); 78 g/d of linseed and 4 g/d of safflower oil (EFA); Lutalin (CLA, isomers cis-9,trans-11 and trans-10,cis-12 CLA, each 10 g/d); or the combination of EFA+CLA. Blood samples were collected several times AP and PP to determine the concentrations of plasma metabolites and hormones related to glucose metabolism and the somatotropic axis. Liver tissue samples were collected several days AP and PP to measure glycogen concentration and the mRNA abundance of genes related to gluconeogenesis and the somatotropic axis. On d 28 AP and 21 PP, endogenous glucose production (eGP) and glucose oxidation (GOx) were measured via tracer technique. The concentration of plasma glucose was higher in CLA than in non-CLA-treated cows, and the plasma β-hydroxybutyrate concentration was higher in EFA than in non-EFA cows on d 21 PP. The eGP increased from AP to PP with elevated eGP in EFA and decreased eGP in CLA-treated cows; GOx was lower in CLA than in CTRL on d 21 PP. The plasma insulin concentration decreased after calving in all groups and was higher in CLA than in non-CLA cows at several time points. Plasma glucagon and cortisol concentrations on d 21 PP were lower in CLA than non-CLA groups. The glucagon/insulin and glucose/insulin ratios were higher in CTRL than in CLA group during the transition period. Plasma IGF-I concentration was lower in EFA than non-EFA cows on d 42 AP and was higher during the dry period and early lactation in CLA than in non-CLA cows. The IGF binding protein (IGFBP)-3/-2 ratio in blood plasma was higher in CLA than in non-CLA cows. Hepatic glycogen concentration on d 28 PP was higher, but the mRNA abundance of PC and IGFBP2 was lower in CLA than non-CLA cows on d 1 PP. The EFA treatment decreased the mRNA abundance of IGFBP3 AP and PCK1, PCK2, G6PC, PCCA, HMGCS2, IGFBP2, and INSR at several time points PP. Results indicated elevated concentrations of plasma glucose and insulin along with the stimulation of the somatotropic axis in cows treated with CLA, whereas EFA treatment stimulated eGP but not mRNA abundance related to eGP PP. The systemic effects of the combined EFA+CLA treatment were very similar to those of CLA treatment, but the effects on hepatic gene expression partially corresponded to those of EFA treatment

Effects of conjugated linoleic acids isomers on oxidative mammary gland metabolism

The study aimed at investigating the effect of CLA isomers cis- 9,trans-11 (c9,t11) and trans-10,cis-12 (t10,c12), on oxidative status of bovine mammary gland. For the study bovine epithelial cell line BME-UV1 and the aflatoxin B1 (AFB1) as oxidizing agent as experimental model have been used. The uptake rates of CLA in cells at 3 and 48h were tested using DAD HPLC analy- sis. The cells were pre-incubated with complete medium contain- ing 50 mM c9,t11, 50 mM t10,c12 and 50 mM CLA Mix (50% c9,t11 and 50% t10,c12) and then AFB1 (20 mg/mL) was added. After 48h of incubation the cells treated in the presence or absence of AFB1 were collected for determining cell viability (XTT assay), oxidative markers such as nicotinamide adenine dinucleotide phosphate (NADP+/NADPH), glutathione (GSH/GSSG), thiobar- bituric acid reactive substances (TBARS), protein carbonyl groups (CP), superoxide dismutase (SOD), glutathione peroxi- dase (GPx1), glutathioneS-transferase (GST) and glutathione reductase (GR). The mRNAs quantification of bovine GSHPx-1, GSR, GST and SOD, and milk α-CN, β-CN and α-Lalb was per- formed by rt-PCR. Data were analysed by ANOVA and differences were declared significant at P<0.05. The results showed that the uptake rates of CLA in cells increased from 3 to 48h. Cells exposed to AFB1 showed a loss of cell viability after 48h. CLA have increased (P<0.05) the concentration of reduced GSH and NADPH and decreased (P<0.01) the levels of GSSG, mostly in cells treated with t10,c12. The activity of GR and GST was decreased (P<0.05) in cells treated with CLA. Higher levels of GSPx1 and SOD (P<0.01) activities were observed in cells treat- ed with CLA in presence of AFB1. Increase of TBARS levels was observed in cells treated with CLA in presence of AFB1, whereas CP content increased both in cells treated with CLA and with CLA in presence of AFB1. Regarding the mRNA’s expression of GPX1, GR, GST and SOD no differences were observed among all treat- ments. No treatments, CLA alone or CLA plus AFB1, had a sub- stantial effect on gene expression of milk proteins. Findings of the present study corroborate an antioxidant role of CLA by developing a significant improvement of redox status in cells, in particular t10,c12. CLA treatment might enhance the intracellu- lar redox homeostasis and could be of help in improving physio-logical oxidative stress situations as the periparturient period in dairy cow

Effect of maternal supplementation with essential fatty acids and conjugated linoleic acid on metabolic and endocrine development in neonatal calves.

We tested the hypothesis that the maternal supply of essential fatty acids (EFA), especially α-linolenic acid, and conjugated linoleic acid (CLA), affects glucose metabolism, the endocrine regulation of energy metabolism and growth, and the intestinal development of neonatal calves. We studied calves from dams that received an abomasal infusion of 76 g/d coconut oil (CTRL; n = 9), 78 g/d linseed oil and 4 g/d safflower oil (EFA; n = 9), 38 g/d Lutalin (BASF SE) containing 27% cis-9,trans-11 and trans-10,cis-12 CLA (CLA; n = 9), or a combination of EFA and CLA (EFA+CLA; n = 11) during the last 63 d of gestation and early lactation. Calves received colostrum and transition milk from their own dam for the first 5 d of life. Insulin-like growth factor (IGF)-I, leptin, and adiponectin concentrations were measured in milk. Blood samples were taken before first colostrum intake, 24 h after birth, and from d 3 to 5 of life before morning feeding to measure metabolic and endocrine traits in plasma. On d 3 of life, energy expenditure was evaluated by a bolus injection of NaH13CO3 and determination of CO2 appearance rate. On d 4, additional blood samples were taken to evaluate glucose first-pass uptake and 13CO2 enrichment after [13C6]-glucose feeding and intravenous [6,6-2H2]-glucose bolus injection, as well as postprandial changes in glucose, nonesterified fatty acids (NEFA), insulin, and glucagon. On d 5, calves were killed 2 h after feeding and samples of small intestinal mucosa were taken for histomorphometric measurements. The concentrations of IGF-I, adiponectin, and leptin in milk decreased during early lactation in all groups, and the concentrations of leptin in first colostrum was higher in EFA than in CTRL cows. Plasma glucose concentration before first colostrum intake was higher in EFA calves than in non-EFA calves and was lower in CLA calves than in non-CLA calves. Plasma IGF-I concentration was higher on d 1 before colostrum intake in EFA calves than in EFA+CLA calves and indicated an overall CLA effect, with lower plasma IGF-I in CLA than in non-CLA calves. Postprandial NEFA concentration was lowest in EFA and CLA calves. The postprandial rise in plasma insulin was higher in EFA than in non-EFA calves. Plasma adiponectin concentration increased from d 1 to d 2 in all groups and was higher on d 3 in CLA than in non-CLA calves. Plasma leptin concentration was higher on d 4 and 5 in EFA than in non-EFA calves. Maternal fatty acid treatment did not affect energy expenditure and first-pass glucose uptake, but glucose uptake on d 4 was faster in EFA than in non-EFA calves. Crypt depth was lower, and the ratio of villus height to crypt depth was higher in the ilea of CLA than non-CLA calves. Elevated plasma glucose and IGF-I in EFA calves immediately after birth may indicate an improved energetic status in calves when dams are supplemented with EFA. Maternal EFA and CLA supplementation influenced postprandial metabolic changes and affected factors related to the neonatal insulin response

Metabolic adaptation during early lactation: key to cow health, longevity and a sustainable dairy production chain

Enhancing longevity by reducing involuntary culling and consequently increasing productive life and lifetime production of dairy cows is not only a strategy to improve a farm’s profit, but is also related to improved animal welfare. High rates of involuntary culling in dairy cows are currently attributed to fertility problems, mastitis and locomotive disorders. Disease incidence is high in particular in the early-lactation period. The high disease incidence in early lactation has been attributed to metabolic stress related to the high metabolic priority for lactation and the inability of the cow to adapt effectively to the new lactation. Several biological mechanisms interact in the peripartum period of dairy cows and can result in this inability to adapt effectively to lactation. Biological mechanisms reviewed are metabolic adaptation, oxidative stress, immune function and inflammation, and feed intake capacity. Although relationships between these mechanisms become increasingly clear, these relationships are complex and not yet completely understood. Appropriate management of dairy cows in the peripartum period can facilitate cows to adapt to a new lactation. Nutritional and management strategies to ease adaptation are divided into strategies to restrict energy intake in the dry period, to improve energy intake in early lactation, alter repartitioning of energy between milk and body tissue, and strategies to support fat or carbohydrate metabolism. The success of various strategies, however, is often hampered by the complexity ofinteractions and high between-cow variation. We advocate for a multidisciplinary approach tounderstand and manage adaptation to a new lactation aiming at an improvement of cow welfareand longevity