S.S. Donkin

Replacing corn with glycerol in diets for transition dairy cows.

Expansion of the biofuels industry has increased the availability of glycerol as an alternative feed for dairy cows. The objective of this study was to determine the effects of glycerol on feed intake, milk production, rumen volatile fatty acids, and metabolic parameters in transition dairy cows. Multiparous Holstein cows were fed diets containing either high-moisture corn (n=11) or glycerol (n=12) from -28 to +56 d relative to calving. Glycerol was included at 11.5 and 10.8% of the ration dry matter for the pre- and postpartum diets, respectively. Prepartum feed intake was not changed by glycerol feeding (14.9 vs. 14.6 kg/d, control vs. glycerol) nor did postpartum feed intake differ (19.8 vs. 20.7 kg/d, control vs. glycerol). Overall milk yield did not differ (35.8 vs. 37 kg/d, control vs. glycerol) and milk composition, milk urea nitrogen, somatic cells, and energy balance were not different with glycerol feeding. Blood glucose content was decreased in cows fed glycerol during the prepartum period (59.1 vs. 53.4 mg/dL), and β-hydroxybutyrate concentration was increased (0.58 vs. 0.82 mmol/L, control vs. glycerol). Concentrations of blood nonesterified fatty acids did not differ between the treatment groups, and no response to glycerol for blood metabolites during the postpartum period was observed. Total rumen volatile fatty acid concentrations (mmol/L) did not differ between treatments, but proportions of rumen propionate and butyrate were greater for cows fed glycerol (22.7 vs. 28.6% of propionate, control vs. glycerol; and 11.5 vs. 15.3% of butyrate, control vs. glycerol) at the expense of acetate (61.4 vs. 51.5%, control vs. glycerol). These data indicate that glycerol is a suitable replacement for corn grain in diets for transition dairy cows.

Periparturient effects of feeding a low dietary cation-anion difference diet on acid-base, calcium, and phosphorus homeostasis and on intravenous glucose tolerance test in high-producing dairy cows

Feeding rations with low dietary cation-anion difference (DCAD) to dairy cows during late gestation is a common strategy to prevent periparturient hypocalcemia. Although the efficacy of low-DCAD rations in reducing the incidence of clinical hypocalcemia is well documented, potentially deleterious effects have not been explored in detail. The objective of the study presented here was to determine the effect of fully compensated metabolic acidosis on calcium and phosphorus homeostasis, insulin responsiveness, and insulin sensitivity as well as on protein metabolism. Twenty multiparous Holstein-Friesian dairy cows were assigned to 1 of 2 treatment groups and fed a low-DCAD ration (DCAD = -9 mEq/100g, group L) or a control ration (DCAD = +11 mEq/100g, group C) for the last 3 wk before the expected calving date. Blood and urine samples were obtained periodically between 14 d before to 14 d after calving. Intravenous glucose tolerance tests and 24-h volumetric urine collection were conducted before calving as well as 7 and 14 d postpartum. Cows fed the low-DCAD ration had lower urine pH and higher net acid excretion, but unchanged blood pH and bicarbonate concentration before calving. Protein-corrected plasma Ca concentration 1 d postpartum was higher in cows on the low-DCAD diet when compared with control animals. Urinary Ca and P excretion was positively associated with urine net acid excretion and negatively associated with urine pH. Whereas metabolic acidosis resulted in a 6-fold increase in urinary Ca excretion, the effect on renal P excretion was negligible. A more pronounced decline of plasma protein and globulin concentration in the periparturient period was observed in cows on the low-DCAD diets resulting in significantly lower total protein and globulin concentrations after calving in cows on low-DCAD diets. Intravenous glucose tolerance tests conducted before and after calving did not reveal group differences in insulin response or insulin sensitivity. Our results indicate that fully compensated metabolic acidosis increased the Ca flux resulting in increased urinary calcium excretion before calving and increased plasma Ca concentration on the day after calving, whereas the effect on P homeostasis was unlikely to be clinically relevant. The clinical relevance of the effect of metabolic acidosis on the plasma protein and globulin concentration is unclear but warrants further investigation.

Characterization of bovine pyruvate carboxylase promoter 1 responsiveness to serum from control and feed-restricted cows1

Pyruvate carboxylase (PC; EC 6.4.1.1) is critical in gluconeogenesis from lactate and maintenance of tricarboxylic acid cycle intermediates. Whereas increases in PC mRNA have been observed during feed restriction, the mechanism of regulation is unknown; however, coinciding increases in circulating NEFA concentrations suggests that fatty acids may contribute to regulation of gene expression during feed restriction. The objective of this study was to examine the direct effect of exposure to serum from full-fed control cows with serum from cows that were restricted to 50% of ad libitum intake for 5 d on PC expression in vitro. Rat hepatoma (H4IIE) cells were transiently transfected with bovine promoter-luciferase constructs containing bovine PC promoter 1 and treated with serum from control cows, serum from feed-restricted cows, or modified serum. Modified serum pools were generated by supplemented serum from control cows with C14:0, C16:0, C18:0, C18:1n-9 cis, C18:2n-6 cis, and C18:3n-3 cis to match the total NEFA in serum from feed-restricted cows (1.3 mM) in the relative proportion found in serum from control or feed-restricted cows. Exposure of cells to serum from feed-restricted cows increased (P < 0.05) PC promoter 1 activity 2.2-fold compared with cells exposed to control cow serum. Exposure to serum from control cows with fatty acids added to a NEFA concentration of 1.3 mM to reflect the fatty acid profile of control and feed-restricted cows increased (P < 0.05) promoter 1 activity 2.1- and 2.5-fold, respectively, compared with cells incubated with control cow serum. There was no difference (P ≥ 0.05) in promoter 1 activity in cells treated with modified serum compared with serum from feed-restricted cows. These data indicate that promoter 1 is activated by fatty acids found in serum of feed-restricted cows. These data suggest a role of NEFA to regulate expression of bovine PC mRNA through specific activation of PC promoter 1.

Immune and production responses of dairy cows to postruminal supplementation with phytonutrients

This study investigated the effect of phytonutrients (PN) supplied postruminally on nutrient utilization, gut microbial ecology, immune response, and productivity of lactating dairy cows. Eight ruminally cannulated Holstein cows were used in a replicated 4×4 Latin square. Experimental periods lasted 23 d, including 14-d washout and 9-d treatment periods. Treatments were control (no PN) and daily doses of 2g/cow of either curcuma oleoresin (curcumin), garlic extract (garlic), or capsicum oleoresin (capsicum). Phytonutrients were pulse-dosed into the abomasum of the cows, through the rumen cannula, 2 h after feeding during the last 9 d of each experimental period. Dry matter intake was not affected by PN, although it tended to be lower for the garlic treatment compared with the control. Milk yield was decreased (2.2 kg/d) by capsicum treatment compared with the control. Feed efficiency, milk composition, milk fat and protein yields, milk N efficiency, and 4.0% fat-corrected milk yield were not affected by treatment. Rumen fermentation variables, apparent total-tract digestibility of nutrients, N excretion with feces and urine, and diversity of fecal bacteria were also not affected by treatment. Phytonutrients had no effect on blood chemistry, but the relative proportion of lymphocytes was increased by the capsicum treatment compared with the control. All PN increased the proportion of total CD4(+) cells and total CD4(+) cells that co-expressed the activation status signal and CD25 in blood. The percentage of peripheral blood mononuclear cells (PBMC) that proliferated in response to concanavalin A and viability of PBMC were not affected by treatment. Cytokine production by PBMC was not different between control and PN. Expression of mRNA in liver for key enzymes in gluconeogenesis, fatty acid oxidation, and response to reactive oxygen species were not affected by treatment. No difference was observed due to treatment in the oxygen radical absorbance capacity of blood plasma but, compared with the control, garlic treatment increased 8-isoprostane levels. Overall, the PN used in this study had subtle or no effects on blood cells and blood chemistry, nutrient digestibility, and fecal bacterial diversity, but appeared to have an immune-stimulatory effect by activating and inducing the expansion of CD4 cells in dairy cows. Capsicum treatment decreased milk yield, but this and other effects observed in this study should be interpreted with caution because of the short duration of treatment.

Glucagon increases hepatic mRNA concentrations of ureagenic and gluconeogenic enzymes in early-lactation dairy cows

Exogenous glucagon increases hepatic glucose synthesis in part by increasing hepatic extraction of amino acids from blood for conversion to glucose. To examine the role of glucagon in orchestrating gene expression of gluconeogenic and ureagenic enzymes, we determined the mRNA concentrations of key hepatic ureagenic and gluconeogenic enzymes at d 11, 15, and 22 postpartum in multiparous Holstein cows that received 0 or 5 mg of glucagon in 60 mL of saline by subcutaneous injection every 8 h for 14 d starting on d 8 postpartum. On d 11 postpartum, glucagon increased the hepatic mRNA concentrations for all measured ureagenic enzymes (carbamoylphosphate synthetase I, ornithine transcarbamylase, and argininosuccinate synthetase) and gluconeogenic enzymes (pyruvate carboxylase and cytosolic and mitochondrial forms of phosphoenolpyruvate carboxykinase) and increased or tended to increase mRNA concentrations of gluconeogenic enzymes on d 15 postpartum but not on d 22. The effect of glucagon to increase mRNA concentrations of ureagenic and gluconeogenic enzymes was limited to times when concentrations of plasma insulin were not increased. Our results suggest that hepatic gene expression of key ureagenic and gluconeogenic enzymes in early-lactation dairy cows is responsive to hormonal regulation by glucagon.

Short communication: Genetic differences between New Zealand and North American dairy cows alter milk production and gluconeogenic enzyme expression1

Continuous selection of dairy cows for production traits may alter the regulation of metabolic pathways. High-producing North American (NA) cows produce more milk and have a larger degree of somatotropic axis uncoupling than less intensively selected New Zealand (NZ) cows. The objective of this study was to determine if production-based selection priorities (i.e., NA cows) have altered the regulation of the gluconeogenic pathway relative to selection priorities based on production traits (i.e., NZ cows). In this study conducted in New Zealand, NZ (n=27) and NA cows (n=27) were monitored from 1 wk before calving to 12 wk post-calving. Cows were pasture-fed and supplemented with 0, 3, or 6 kg of concentrate DM/d. Liver biopsy samples were collected at 0, +1, and +4 wk relative to calving (WRTC) for mRNA analysis. Milk production of NA cows was greater during wk 5 to 11 postpartum and concentrate supplementation increased milk production for both NA and NZ cows. No genotype (NA vs. NZ) by diet interaction occurred for blood glucose, NEFA, or insulin. Expression of pyruvate carboxylase (PC) mRNA was increased at +1 and +4 WRTC compared with 0 WRTC (3.04 and 2.42 vs. 1.25±0.13 arbitrary units, respectively: mean ± standard error of the means) and expression of cytosolic phosphoenolpyruvate carboxykinase mRNA was increased at +4 compared with calving and +1 WRTC (4.78 vs. 2.18 and 2.48±1.41 arbitrary units, respectively). Expression of PC mRNA tended to be greater in NZ cows and tended to decrease with concentrate supplementation in both NZ and NA cows. The responses of NZ and NA cows to the transition to lactation and concentrate supplementation appeared to be similar; however, NZ cattle had a higher basal expression of PC.

Feeding behaviors of transition dairy cows fed glycerol as a replacement for corn.

Feed sorting is a natural behavior of dairy cows that can result in inconsistencies in the nutritive value of a total mixed ration (TMR). The objective of this study was to determine the effects of replacing high-moisture corn with glycerol on feed sorting and the feed intake pattern of transition dairy cows. Multiparous Holstein cows (n=26) were paired by expected calving date, housed in individual tie stalls, and fed diets containing either glycerol or high-moisture corn once daily from d -28 to +56 relative to calving. Glycerol was included at 11.5 and 10.8% of the ration dry matter for the pre- and postpartum diets, respectively. The feed consumption pattern was determined by measuring TMR disappearance during the intervals from 0 to 4 h, 4 to 8 h, 8 to 12 h, and 12 to 24 h relative to feed delivery. Feed sorting was determined on d -16, -9, 9, 16, and 51 relative to calving at 4, 8, 12 and 24 h after feeding. The TMR particle size profile was determined at feed delivery and at 4, 8, 12, and 24 after feed delivery by using the Penn State Particle Separator (Nasco, Fort Atkinson, WI) to yield long (>19 mm), medium (<19 mm, >8 mm), short (<8 mm, >1.18 mm), and fine (<1.18 mm) particles. Overall feed intake did not differ between diets and was 14.7±0.4 and 20.2±0.5 kg/d for the pre- and postpartum intervals, respectively. During the prepartum period, glycerol decreased the amount of feed consumed during the first 4h after feed delivery (7.22 vs. 5.59±0.35 kg; control vs. glycerol, respectively) but increased feed consumed from 12 through 24 h after feed delivery (2.22 vs. 3.82±0.35 kg; control vs. glycerol, respectively). Similar effects on the feed consumption pattern were observed after calving. During the prepartum period, cows fed the control diet sorted against long particles, whereas cows fed glycerol did not sort against long particles (77.2 vs. 101.5±3.50% of expected intake for control vs. glycerol; significant treatment effect). The data indicate that addition of glycerol to the TMR alters the feed consumption pattern to increase feed consumption late in the day at the expense of feed consumed immediately after feeding, and it reduces sorting behavior against long particles. Together, these may reduce diurnal variations in the rumen environment to promote greater rumen health in transition cows.

Effect of circulating exosomes from transition cows on Madin-Darby bovine kidney cell function

The greatest risk of metabolic and infectious disease in dairy cows is during the transition from pregnancy to lactating (i.e., the transition period). The objective of this experiment was to determine the effects of extracellular vesicles (microvesicles involved in cell-to-cell signaling) isolated from transition cows on target cell function. We previously identified differences in the protein profiles of exosomes isolated from cows divergent in metabolic health status. Therefore, we hypothesized that these exosomes would affect target tissues differently. To investigate this, 2 groups of cows (n = 5/group) were selected based on the concentration of β-hydroxybutyrate and fatty acids in plasma and triacylglycerol concentration in liver at wk 1 and 2 postcalving. Cows with high concentrations of β-hydroxybutyrate, fatty acids, and triacylglycerol were considered at increased risk of clinical disease during the transition period (high-risk group; n = 5) and were compared with cows that had low concentrations of the selected health indicators (low-risk group; n = 5). At 2 time points during the transition period (postcalving at wk 1 and 4), blood was sampled and plasma exosomes were isolated from the high-risk and low-risk cows. The exosomes were applied at concentrations of 10 and 1 µg/mL to 5 × 103 Madin-Darby bovine kidney cells grown to 50% confluence in 96-well plates. Results indicate a numerical increase in cell proliferation when exosomes from high-risk cows were applied compared with those from low-risk cows. Consistent with an effect on cell proliferation, quantitative reverse transcriptase PCR indicated a trend for upregulation of 3 proinflammatory genes (granulocyte colony-stimulating factor, ciliary neurotrophic factor, and CD27 ligand) with the application of high-risk exosomes, which are involved in cellular growth and survival. Proteomic analysis indicated 2 proteins in the low-risk group that were not identified in the high-risk group (endoplasmin and catalase), which may also be indicative of the metabolic state of origin. It is likely that the metabolic state of the transition cow affects cellular function through exosomal messaging; however, more in-depth research into cross-talk between exosomes and target cells is required to determine whether exosomes influence Madin-Darby bovine kidney cells in this manner.

Hepatic expression of aminoadipate semialdehyde synthase is unchanged by postruminal lysine supply in lactating dairy cows

Lysine supply is potentially limiting for milk production in dairy cows. The availability of Lys to the mammary gland and other tissues is a function of the quantity of metabolizable Lys supplied and Lys catabolism by the liver. Likewise, Lys catabolism may be influenced by Lys supply. This study evaluated the effect of increased postruminal Lys supply on the expression of aminoadipate semialdehyde synthase (AASS, a committing step in Lys catabolism in the liver) and ornithine transcarbamoylase and argininosuccinate synthase (key urea cycle enzymes that are responsive to protein supply). Eight multiparous peak Holstein cows were used in a replicated 4 × 4 Latin square. Cows were fed a Lys-limiting ration and infused postruminally with 0, 9, 27, or 63 g/d of Lys. The study consisted of 10 d of pretreatment followed by 10 d of Lys infusion. On the last day of each period, liver and milk samples were collected for mRNA analysis, and blood samples were collected for analysis of amino acids and Lys metabolites. Milk protein percent increased by 5.9%, plasma Lys increased by 74%, and α-aminoadipic acid increased by 51% with postruminal infusion of 63 g/d Lys compared with 0 g/d. Expression of AASS, ornithine transcarbamoylase, and argininosuccinate synthase mRNA in liver did not differ with postruminal infusion of Lys. Milk fat globule mRNA for major milk proteins and AASS were not affected by Lys infusion. Postruminal infusion of Lys resulted in an 86% greater increase in AASS mRNA in the liver compared with mammary mRNA. These changes suggest that hepatic Lys metabolism is not responsive to Lys supply at the transcription level, and that the availability of Lys to extrahepatic tissue may be determined by hepatic Lys metabolism.