Evaluation of feed restriction and abomasal infusion of resistant starch as models to induce intestinal barrier dysfunction in healthy lactating cows.

Intestinal hyperpermeability and subsequent immune activation alters nutrient partitioning and thus, decreases productivity. Developing experimental models of intestinal barrier dysfunction in heathy cows is a prerequisite in identifying nutritional strategies to mitigate it. Six cannulated Holstein cows (mean ± standard deviation, 37 ± 10 kg/d milk yield; 219 ± 97 d in milk; 691 ± 70 kg body weight) were used in a replicated 3 × 3 Latin square design experiment with 21-d periods (16-d wash-out and 5-d challenge) to evaluate either feed restriction or hindgut acidosis as potential models for inducing intestinal hyperpermeability. Cows were randomly assigned to treatment sequence within square and treatment sequences were balanced for carryover effects. Treatments during the challenge were (1) control (CTR; ad libitum feeding); (2) feed restriction (FR; total mixed ration fed at 50% of ad libitum feed intake); and (3) resistant starch (RS; 500 g of resistant starch infused in abomasum once a day as a pulse-dose 30 min before morning feeding). The RS (ActiStar RT 75330, Cargill Inc.) was tapioca starch that was expected to be resistant to enzymatic digestion in the small intestine and highly fermentable in the hindgut. Blood samples were collected 4 h after feeding on d 13 and 14 of the wash-out periods (baseline data used as covariate), and on d 1, 3, and 5 of the challenge periods. Fecal samples were collected 4 and 8 h after the morning feeding on d 14 of the wash-out periods and d 5 of the challenge periods. By design, FR decreased dry matter intake (48%) relative to CTR and RS, and this resulted in marked reductions in milk and 3.5% FCM yields over time, with the most pronounced decrease occurring on d 5 of the challenge (34 and 27%, respectively). Further, FR increased somatic cell count by 115% on d 5 of the challenge relative to CTR and RS. Overall, FR increased nonesterified fatty acids (159 vs. 79 mEq/L) and decreased BHB (8.5 vs. 11.2 mg/dL), but did not change circulating glucose relative to CTR. However, RS had no effect on production or metabolism metrics. Resistant starch decreased fecal pH 8 h after the morning feeding (6.26 vs. 6.81) relative to CTR and FR. Further, RS increased circulating lipopolysaccharide binding protein (4.26 vs. 2.74 µg/mL) compared with FR only on d 1 of the challenge. Resistant starch also increased Hp (1.52 vs. 0.48 µg/mL) compared with CTR, but only on d 5 of the challenge. However, neither RS or FR affected concentrations of serum amyloid A, IL1ß, or circulating endotoxin compared with CTR. The lack of consistent responses in inflammatory biomarkers suggests that FR and RS did not meaningfully affect intestinal barrier function. Thus, future research evaluating the effects of hindgut acidosis and FR using more intense insults and direct metrics of intestinal barrier function is warranted.

Effects of hindgut acidosis on metabolism, inflammation, and production in dairy cows consuming a standard lactation diet.

Postruminal intestinal barrier dysfunction caused by excessive hindgut fermentation may be a source of peripheral inflammation in dairy cattle. Therefore, the study objectives were to evaluate the effects of isolated hindgut acidosis on metabolism, inflammation, and production in lactating dairy cows. Five rumen-cannulated lactating Holstein cows (32.6 ± 7.2 kg/d of milk yield, 242 ± 108 d in milk; 642 ± 99 kg of body weight; 1.8 ± 1.0 parity) were enrolled in a study with 2 experimental periods (P). During P1 (4 d), cows were fed ad libitum a standard lactating cow diet (26% starch dry matter) and baseline data were collected. During P2 (7 d), all cows were fed the same diet ad libitum and abomasally infused with 4 kg/d of pure corn starch (1 kg of corn starch + 1.25 L of H2O/infusion at 0600, 1200, 1800, and 0000 h). Effects of time (hour relative to the first infusion or day) relative to P1 were evaluated using PROC MIXED in SAS (version 9.4; SAS Institute Inc.). Infusing starch markedly reduced fecal pH (5.84 vs. 6.76) and increased fecal starch (2.2 to 9.6% of dry matter) relative to baseline. During P2, milk yield, milk components, energy-corrected milk yield, and voluntary dry matter intake remained unchanged. At 14 h, plasma insulin and ß-hydroxybutyrate increased (2.4-fold and 53%, respectively), whereas circulating glucose concentrations remained unaltered. Furthermore, blood urea nitrogen increased at 2 h (23%) before promptly decreasing below baseline at 14 h (13%). Nonesterified fatty acids tended to decrease from 2 to 26 h (40%). Circulating white blood cells and neutrophils increased on d 4 (36 and 73%, respectively) and somatic cell count increased on d 5 (4.8-fold). However, circulating serum amyloid A and lipopolysaccharide-binding protein concentrations were unaffected by starch infusions. Despite minor changes in postabsorptive energetics and leukocyte dynamics, abomasal starch infusions and the subsequent hindgut acidosis had little or no meaningful effects on biomarkers of immune activation or production variables.

The effects of feeding mixed tocopherol oil on whole-blood respiratory burst and neutrophil immunometabolic-related gene expression in lactating dairy cows.

The 4 major tocopherol isoforms differ in their biochemical reactivity and cellular effects due to basic chemical structural differences. Alpha-tocopherol has been well studied regarding effects on bovine polymorphonuclear leukocyte (PMN) function and its involvement in respiratory burst. However, no studies to date have identified the effects of supplementing a mixed tocopherol oil (Tmix) particularly enriched in non-α tocopherol isoforms (i.e., γ- and δ-isoforms) on fundamental immunometabolic changes in dairy cows. Therefore, the objectives of this study were to determine whether short-term feeding of vegetable oil-derived Tmix alters specific biomarkers of metabolism, whole-blood leukocyte populations, respiratory burst, immunometabolic-related gene expression of PMN, or gene expression of isolated PMN when challenged with lipopolysaccharides (LPS). Clinically healthy multiparous lactating Holstein cows (n = 12; 179 ± 17 d in milk, 40.65 ± 3.68 kg of milk yield) were fed Tmix (620 g/d) for 7 consecutive days. Jugular blood (EDTA anticoagulant) was collected from all cows on d 0 before treatment initiation and again on d 7 after Tmix feeding. Total stimulated respiratory burst activity (RBA) and leukocyte populations were assessed in whole blood, and tocopherol isoform concentrations, metabolites, and hormones were measured in plasma. For gene expression analysis, isolated PMN from cows before and after Tmix feeding were incubated with LPS at a final concentration of either 0.0 or 1.5 µg/mL. Feeding of Tmix for 7 d increased the concentrations of α- and γ-tocopherol. The Tmix did not alter plasma insulin but decreased cholesterol. The Tmix did not alter whole-blood RBA or the leukocyte populations. The LPS challenge increased the expression of proinflammatory genes TNFA and IL6. However, Tmix treatment did not alter the patterns of LPS-affected expression of genes (e.g., TNFA, ITGB2, PPARA, and RXRA) associated with the immune or metabolic response. In conclusion, short-term feeding of Tmix may have no negative effect on animal health as Tmix increased α- and γ-tocopherol concentrations in blood and did not impair whole-blood RBA or alter leukocyte populations. The data provide further support that the α- and γ-tocopherol isoforms do not interfere with normal immune or metabolic function.

Effects of energy supply on leucine utilization by growing steers at two body weights.

The effects of energy supplementation on Leu utilization in growing steers were evaluated in 2 experiments by using 6 ruminally cannulated Holstein steers. In Exp. 1, steers (initial BW = 150 +/- 7 kg) were limit-fed (2.3 kg of DM/d) a diet based on soybean hulls and received a basal ruminal infusion of 100 g of acetate/d, 75 g of propionate/d, and 75 g of butyrate/d, as well as abomasal infusions of 200 g of glucose/d and a mixture (215 g/d) containing all essential AA except Leu. Treatments were arranged as a 3 x 2 factorial, with 3 amounts of Leu infused abomasally (0, 4, and 8 g/d) and supplementation of diets with 2 amounts of energy (0 and 1.9 Mcal/d of GE). Supplemental energy was supplied by ruminal infusion of 100 g of acetate/ d, 75 g of propionate/d, and 75 g of butyrate/d, as well as abomasal infusion of 200 g of glucose/d to provide energy to the animal without affecting the microbial protein supply. When no supplemental energy was provided, Leu supplementation increased N balance, with no difference between 4 and 8 g/d of Leu (24.5, 27.0, and 27.3 g/d for 0, 4, and 8 g/d of Leu), but when additional energy was supplied, N retention increased linearly in response to Leu (25.6, 28.5, and 31.6 g/d for 0, 4, and 8 g/d of Leu; Leu x energy interaction, P = 0.06). The changes in N balance were the result of changes in urinary N excretion. The greater Leu retentions in response to energy supplementation when Leu was the most limiting nutrient indicate that energy supplementation improved the true efficiency of Leu utilization. In addition, supplemental energy increased the gross efficiency of Leu utilization when the Leu supply was not limiting by increasing the maximal rates of protein deposition. Experiment 2 was similar to Exp. 1, but steers had an initial BW of 275 +/- 12 kg and were limit-fed at 3.6 kg of DM/d. Retention of N was not affected (P = 0.22) by Leu supplementation, indicating that Leu did not limit protein deposition. Energy supply increased N retention (P < 0.01) independently of Leu supplementation (33.0 vs. 27.8 g/d). Overall, energy supplementation improved Leu utilization by modestly increasing N retention when Leu was limiting and by increasing the ability of steers to respond to the greatest amount of supplemental Leu. We conclude from these results that the assumption of a constant efficiency of AA utilization is unlikely to be appropriate for growing steers.

Silage chop length and hay supplementation on milk yield, chewing activity, and ruminal digestion by dairy cows.

The effects of whole-plant corn silage (CS) particle size and long unprocessed grass hay (LH) supplementation on milk yield, chewing activity, and ruminal digestion in dairy cows were evaluated in 2 experiments. In Experiment 1, corn silage harvested at fine (6 mm; FCS) or coarse (23 mm; CCS) theoretical cut length were fed to 22 lactating Holstein cows. Treatments were 2 total mixed rations containing 58% of dry matter (DM) as FCS or CCS. Diet DM intake tended to be higher in cows fed FCS than those fed CCS (23.4 vs. 22.1 kg/d). However, milk yield and composition, body condition score, and plasma metabolite concentrations were not affected by the dietary treatments. In the second experiment, 5 cannulated Holstein cows were used in a 5 x 5 Latin square design to evaluate the effects of the addition of LH to the diets evaluated in Experiment 1 on chewing activity and ruminal digestion. Treatments were 5 total mixed rations: FCS-based diet plus the addition of 0, 5, or 10% LH (DM basis) and CCS-based diet plus 0 or 5% LH. Long hay addition linearly decreased DM intake in cows fed FCS-based diets (25.0 to 21.7 kg/d), but increased DM intake in those fed CCS-based diets (22.7 to 27.1 kg/d). The intake of neutral detergent fiber (NDF) increased with LH addition in CCS-based diets (7.6 vs. 9.4 kg/d). Rumination time increased (16.8 to 21.0 min/kg of DM intake) when LH was added to FCS-based diets, but it decreased when included in CCS-based diets (18.8 vs. 12.9 min/kg of DM intake). Ruminal pH was higher (5.9 vs. 5.7) and lag-time for in situ NDF disappearance was shorter (3.5 vs. 8.7 h) for cows fed CCS compared with cows fed FCS. The rate of NDF disappearance tended to be higher for the CCS-based diet with 5% LH than for the diet with 0% LH (2.0 vs. 4.4 %/h), but solids passage rate was not affected by the treatments. These results suggest that addition of LH to FCS-based diets does not affect ruminal environment or digestion, but depressed DM intake. In contrast, addition of LH to CCS-based diets may improve ruminal NDF digestion, increasing DM intake by reducing filling effect and time needed for rumination.

Excess amino acid supply improves methionine and leucine utilization by growing steers.

In 2 experiments, 6 ruminally cannulated Holstein steers (205 +/- 23 and 161 +/- 14 kg initial BW in Exp. 1 and 2, respectively) housed in metabolism crates were used in 6 x 6 Latin squares to study the effects of excess AA supply on Met (Exp. 1) and Leu (Exp. 2) use. All steers received a diet based on soybean hulls (DMI = 2.66 and 2.45 kg/d in Exp. 1 and 2, respectively); ruminal infusions of 200 g of acetate/d, 200 g of propionate/d, and 50 g of butyrate/d, as well as abomasal infusion of 300 g of glucose/d to provide energy without increasing the microbial protein supply; and abomasal infusions of a mixture of all essential AA except Met (Exp. 1) or Leu (Exp. 2). Periods were 6 d, with 2-d adaptations and 4 d to collect N balance data. All treatments were abomasally infused. In Exp. 1, treatments were arranged as a 2 x 3 factorial, with 2 amounts of l-Met (0 or 4 g/d) and 3 AA supplements (no additional AA, control; 100 g/d of nonessential AA + 100 g/d of essential AA, NEAA + EAA; and 200 g/d of essential AA, EAA). Supplemental Met increased (P < 0.01) retained N and decreased (P < 0.01) urinary N and urinary urea N. Retained N increased (P < 0.01) with NEAA + EAA only when 4 g/d of Met was provided, but it increased (P < 0.01) with EAA with or without supplemental Met. Both AA treatments increased (P < 0.01) plasma urea and serum insulin. Plasma glucose decreased (P = 0.03) with supplemental Met. In Exp. 2, treatments were arranged as a 2 x 3 factorial with 2 amounts of L-Leu (0 or 4 g/d) and 3 AA supplements (control, NEAA + EAA, and EAA). Supplemental Leu increased (P < 0.01) retained N and decreased (P < 0.01) urinary N and urinary urea N. Both AA treatments increased (P < 0.01) retained N, and they also increased (P < 0.01) urinary N, urinary urea N, and plasma urea. Serum insulin increased (P = 0.06) with supplemental Leu and tended (P = 0.10) to increase with both AA treatments. Supplementation with excess AA improved Met and Leu use for protein deposition by growing cattle.

Effects of energy level on methionine utilization by growing steers.

We evaluated the effect of energy supplementation on Met use in growing steers. Six ruminally cannulated Holstein steers (228 +/- 8 kg of BW) were used in a 6 x 6 Latin square and fed 2.8 kg of DM/d of a diet based on soybean hulls. Treatments were abomasal infusion of 2 amounts of Met (0 or 3 g/d) and supplementation with 3 amounts of energy (0, 1.3, or 2.6 Mcal of GE/d) in a 2 x 3 factorial arrangement. The 1.3 Mcal/d treatment was supplied through ruminal infusion of 90 g/d of acetate, 90 g/d of propionate, and 30 g/d of butyrate, and abomasal infusion of 30 g/d of glucose and 30 g/d of fat. The 2.6 Mcal/d treatment supplied twice these amounts. All steers received basal infusions of 400 g/d of acetate into the rumen and a mixture (125 g/d) containing all essential AA except Met into the abomasum. No interactions between Met and energy levels were observed. Nitrogen balance was increased (P < 0.05) by Met supplementation from 23.6 to 27.8 g/d, indicating that protein deposition was limited by Met. Nitrogen retention increased linearly (P < 0.05) from 23.6 to 27.7 g/d with increased energy supply. Increased energy supply also linearly reduced (P < 0.05) urinary N excretion from 44.6 to 39.7 g/d and reduced plasma urea concentrations from 2.8 to 2.1 mM. Total tract apparent OM and NDF digestibilities were reduced linearly (P < 0.05) by energy supplementation, from 78.2 and 78.7% to 74.3 and 74.5%, respectively. Whole-body protein synthesis and degradation were not affected significantly by energy supplementation. Energy supplementation linearly increased (P < 0.05) serum IGF-I from 694 to 818 ng/mL and quadratically increased (P < 0.05) serum insulin (0.38, 0.47, and 0.42 ng/mL for 0, 1.3, and 2.6 Mcal/d, respectively). In growing steers, N retention was improved by energy supplementation, even when Met limited protein deposition, suggesting that energy supplementation affects the efficiency of AA use.

Effects of energy source on methionine utilization by growing steers.

We evaluated the effects of different supplemental energy sources on Met use in growing steers. Ruminally cannulated Holstein steers were used in two 6 x 6 Latin squares, and data were pooled for analyses. In Exp. 1, steers (148 kg) were fed 2.3 kg of DM/d of a diet based on soybean hulls. Treatments (2 x 3 factorial) were abomasal infusion of 0 or 3 g of l-Met/d, and supplementation with no energy or with glucose (360 g/d) or fat (150 g/d) continuously infused into the abomasum. In Exp. 2, steers (190 kg) received 2.6 kg of dietary DM/d and were provided (2 x 3 factorial) with 0 or 3 g of l-Met/d, and with no supplemental energy or with acetate (385 g/d) or propionate (270 g/ d) continuously infused into the rumen. In both experiments, the energy sources supplied 1.3 Mcal of GE/d, and all steers received basal infusions of 400 g of acetate/d into the rumen and a mixture (125 g/d) of all essential AA except Met into the abomasum. Nitrogen balance (18.8 vs. 23.5 g/d; P < 0.01) and whole-body protein synthesis (2.1 vs. 2.3 kg/d; P < 0.07) were increased by Met supplementation, indicating that protein deposition was limited by Met. Supplemental energy reduced (P < 0.01) urinary N excretion and increased (P < 0.01) N retention without differences among energy sources. Increases in N retention in response to Met were numerically greater when energy was supplemented. Efficiency of supplemental Met use was 11% when no energy was supplemented but averaged 21% when 1.3 Mcal of GE/d was provided. Whole-body protein synthesis and degradation were not affected by energy supplementation. Serum insulin concentrations were increased by glucose and propionate supplementation. Serum IGF-I concentrations were increased by supplementation with Met or glucogenic sources of energy. In growing steers, N retention was increased by energy supplementation even though protein deposition was limited by Met, suggesting that energy supplementation improves the efficiency of AA use. These responses were independent of the source of energy.

Milk fatty acid composition of cows fed a total mixed ration or pasture plus concentrates replacing corn with fat.

Thirty-one Holstein cows (six ruminally cannulated) were used to evaluate milk fatty acids (FA) composition and conjugated linoleic acid (CLA) content on three dietary treatments: 1) total mixed rations (TMR), 2) pasture (Avena sativa L.) plus 6.7 kg DM/d of corn-based concentrate (PCorn), and 3) pasture plus PCorn with 0.8 kg DM/d of Ca salts of unsaturated FA replacing 1.9 kg DM/d of corn (PFat). No differences were found in total (22.4 kg/d) or pasture (18.5 kg/d) dry matter intake, ruminal pH, or total volatile fatty acids concentrations. Fat supplementation did not affect pasture neutral detergent fiber digestion. Milk production did not differ among treatments (19.9 kg/d) but 4% fat-corrected milk was lower for cows fed the PFat compared to cows fed the TMR (16.1 vs. 19.5 kg/d) primarily because of the lower milk fat percentage (2.56 vs. 3.91%). Milk protein concentration was higher for cows fed the TMR than those on both pasture treatments (3.70 vs. 3.45%). Milk from the cows fed the PCorn had a lower content of short- (11.9 vs. 10.4 g/100 g) and medium-chain (56.5 vs. 47.6 g/100 g) FA, and a higher C18:3 percentage (0.07 vs. 0.57 g/100 g) compared with TMR-fed. Cows fed the PFat had the lowest content of short- (8.85 g/100 g) and medium-chain (41.0 g/100 g) FA, and the highest of long-chain FA (51.4 g/100 g). The CLA content was higher for cows in PCorn treatment (1.12 g/100 g FA) compared with cows fed the TMR (0.41 g/100 g FA), whereas the cows fed the PFat had the highest content (1.91 g/100 g FA). Pasture-based diets increased the concentrations of long-chain unsaturated FA and CLA in milk fat. The partial replacement of corn grain by Ca salts of unsaturated FA in grazing cows accentuated these changes. However, those changes in milk FA composition were related to a depression in milk fat.

Supplementation with partially hydrogenated oil in grazing dairy cows in early lactation.

Effects of partially hydrogenated oil on performance, loss of body weight and body condition score, and blood metabolite and hormone concentrations were evaluated in 37 multiparous Holstein cows in grazing conditions during the first 100 d of lactation. Six additional Holstein cows, each fitted with a ruminal cannula, were allocated to a replicated 3 x 3 Latin square to evaluate effects of supplemental fat on rumen environment and pasture digestion. All cows grazed mixed pastures based on alfalfa (Medicago sativa) and orchardgrass (Dactylis glomerata L.) and received 5.4 kg/d of a basal concentrate to which 0, 0.5, or 1 kg/cow per day of partially hydrogenated oil (melting point 58 to 60 degrees C) containing 30.3, 34.9, 21.8, and 3.3% of C16:0, C18:0, C18:1, and C182, respectively, was added. Feeding 1 kg/d of supplemental fat increased fat-corrected milk from 23.4 to 26.3 kg/d, milk fat content from 3.44 to 3.78%, and milk fat yield from 0.87 to 1.03 kg/d compared to control. Milk protein percentage and yield were not affected. Cows fed 1 kg/d of fat increased the content and yield of C16:0 and C18:0 in milk compared with cows fed no added oil. Dry matter intake (DMI) from pasture decreased from 17.8 kg/d for control cows to 13.6 kg/d for cows fed 1 kg of oil, whereas DMI from concentrate was higher for cows fed 1 kg/d of fat (6.0 kg/d) than for controls (5.2 kg/d). Supplemental fat did not affect total dry matter or estimated energy intake and did not change losses of body weight or body condition scores. Plasma concentrations of nonesterified fatty acids, insulin, somatotrophin, and insulin-like growth factor-I did not differ among treatments. Concentration of plasma triglycerides was lowered from 318.5 to 271.2 mg/dl, whereas plasma cholesterol was elevated from 185.0 to 235.8 mg/dl in cows receiving 1 kg/d of supplemental fat compared with controls. Responses to lipolytic or insulin challenges were not affected by feeding oil. Supplemental fat did not affect the digestion of pasture fiber. The addition of energy in the form of partially hydrogenated fat to early lactation dairy cows fed primarily on pasture increased the yield of fat-corrected milk and milk fat content when it represented about 11% of the total metabolizable energy requirement of cows, without affecting milk protein content. The partial hydrogenation of a byproduct of the oil industry apparently prevented detrimental effects of fat supplementation on ruminal digestion.

Fishmeal supplementation to grazing dairy cows in early lactation.

Our objectives were to determine if grazing dairy cows would respond to fishmeal supplementation and to determine if responses could be explained by stimulation of adipose tissue lipolysis. Thirty-four multiparous Holstein cows (25+/-11 DIM) were supplemented with isonitrogenous concentrates containing either fishmeal or pelleted sunflower meal. On a dry matter (DM) basis, concentrates contained fishmeal (14.5%) or sunflower meal (24.2%), corn grain (55.6% and 50.6%), wheat bran (26.7% and 22%), a mineral-vitamin complex (2.9%) and a flavoring agent (0.3%). Concentrates were consumed at a rate of 5 kg/cow per day. Herbage allowance averaged 49.8+/-6.1 kg of DM/cow per d. Milk (26.8 vs. 25.2 kg/d), fat-corrected milk (23.9 vs. 22.2 kg/d) and milk protein yields (0.90 vs. 0.81 kg/d) were increased by fishmeal. Milk protein percentage was similar among treatments. Milk fat yield and milk and plasma urea nitrogen tended to be higher in cows fed fishmeal. Plasma glucose and nonesterified fatty acids concentrations and differences in concentrations between jugular and mammary veins were increased by fishmeal. The in vivo lipolytic response to a beta-adrenergic agent or the antilipolytic + hypoglycemic action of insulin were not affected. The higher milk production observed with fishmeal can be explained by the quantity and quality of the absorbed protein, higher glucose availability to the mammary gland, and increased lipid mobilization without change in responsiveness of the adipose tissue to lipolytic stimuli.