Does bone mobilization interfere with energy metabolism in transition cows?

The onset of lactation represents a challenge for both mineral homeostasis and energy metabolism in high-performing dairy cows. It has been shown that subclinical and clinical hypocalcemia increases the risk of ketosis and recent studies suggest that bone-derived endocrine factors could play a role in intermediary metabolism. Therefore, we analyzed serum samples from calculated d -7, calculated d -3, d +1, d +3, and d +7 relative to calving from 15 multiparous cows for total Ca, the bone resorption marker CrossLaps, the bone formation marker intact osteocalcin, undercarboxylated osteocalcin (ucOC), insulin, glucose, nonesterified fatty acids, ß-hydroxybutyrate, and insulin-like growth factor 1. Serum concentrations of Ca on d -3 and d +1 were associated with parameters of energy metabolism on d +3 and d +7. As we found large variations for serum concentrations of ucOC already on d -7, we allocated the cows retrospectively to 3 groups: low ucOC, medium ucOC, and high ucOC. These groups differed not only in their ucOC dynamics, but also in insulin sensitivity estimated using the revised quantitative insulin sensitivity index (RQUICKI). High ucOC cows presented with the highest RQUICKI throughout the entire observation period. Our data further support the hypothesis that low serum Ca precedes disturbances of energy metabolism. Furthermore, from our preliminary results it can be assumed that the potential link between mineral homeostasis, bone turnover, and intermediary metabolism should be further investigated.

Gain and loss of subcutaneous and abdominal adipose tissue depot mass of German Holstein dairy cows with different body conditions during the transition period.

Subcutaneous adipose tissue (SCAT) and abdominal adipose tissue (AAT) depots are mobilized during the fresh cow period (FCP) and early lactation period (ELP) to counteract the negative energy balance (NEB). Earlier studies suggested that fat depots contribute differently to lipomobilization and may vary in functionality. Differences between the adipose depots might influence the development of metabolic disorders. Thus, the gain and loss of subcutaneous and abdominal adipose depot masses in Holstein cows with lower and higher body condition (mean body condition scores: 3.48 and 3.87, respectively) were compared in the period from d -42 to d 70 relative to parturition in this study. Animals of the 2 experimental groups represented adequately conditioned and overconditioned cows. Estimated depot mass (eDM) of SCAT, AAT, retroperitoneal, omental, and mesenteric adipose depots of 31 pluriparous German Holstein cows were determined via ultrasonography at d -42, 7, 28, and 70 relative to parturition. The cows were grouped according to the eDM of SCAT on d -42 [low body condition (LBC) group: n = 16, mean eDM 8.6 kg; high body condition (HBC) group: n = 15, mean eDM 15.6 kg]. Average daily change (prepartum gain and postpartum loss) in depot masses during dry period (DP; from d -42 to d 7), FCP (d 7 to d 28), and ELP (d 28 to d 70) were calculated and daily dry matter intake and lactation performance recorded. Cows of this study stored about 2 to 3 times more fat in AAT than in SCAT depots. After parturition, on average more adipose tissue mass was lost from the AAT than the SCAT depot (0.23 kg/d vs. 0.14 kg/d). Cows with high compared with low body condition had similar gains in AAT (0.33 kg/d) and SCAT (0.14 kg/d) masses during the DP but mobilized significantly more adipose tissue mass from both depots after calving (AAT, HBC vs. LBC: 0.30 vs. 0.17 kg/d; SCAT, HBC vs. LBC: 0.19 vs. 0.10 kg/d). Correlation analysis indicated a functional disparity between AAT and SCAT. In the case of AAT (R2 = 0.36), the higher the gain in adipose mass during DP, the higher the loss in FCP, but this was not the case for SCAT. During FCP, a greater NEB resulted in greater loss of mass from SCAT (R2 = 0.18). In turn, greater mobilization of SCAT mass led to a higher calculated feed efficiency (R2 = 0.18). However, AAT showed no such correlations. On the other hand, during ELP, loss of both SCAT and AAT mass correlated positively with feed efficiency (R2 = 0.35 and 0.33, respectively). The results indicate that feed efficiency may not be an adequate criterion for performance evaluation in cows during NEB. Greater knowledge of functional disparities between AAT and SCAT depots may improve our understanding of excessive lipomobilization and its consequences for metabolic health and performance of dairy cows during the transition period.

Modeling reticular and ventral ruminal pH of lactating dairy cows using ingestion and rumination behavior.

The prevention and control of metabolic and digestive diseases is an enormous challenge in dairy farming. Subacute ruminal acidosis (SARA) is assumed to be the most severe feed-related disorder and it impairs both animal health and economic efficiency. Currently, ruminal pH as well as variables derived from the daily pH curve are the main indicators for SARA. The objective of this study was to explain the daily pH course in the ventral rumen and reticulum of dairy cows using ingestion pattern and rumination behavior data gathered by automated data recording systems. The data of 13 ruminally fistulated lactating cows were collected at the experimental station of the Friedrich-Loeffler-Institut (Brunswick, Germany). The data included continuous pH measurements, which were recorded simultaneously in the reticulum by pH-measuring boluses and in the ventral rumen by a separate data logger. In addition, rumination behavior was measured using jaw movement sensors, and feed and water intakes were recorded by transponder-assisted systems. Milk yield and body weight were determined during and after each milking, respectively. For statistical evaluation, the data were analyzed using time-series modeling with multiple linear mixed regressions. Before applying the developed mathematical statistical modeling, we performed a plausibility assessment to ensure data quality. The major part of the mathematical statistical modeling consisted of data preparation, where all variables were transformed into a uniform 1-min resolution. Signal transformations were used to model individual feed and water intakes as well as rumination behavior events over time. Our results indicated that diurnal pH curves of both the reticulum and ventral rumen could be predicted by the transformed feed and water intake rates. Rumination events were associated with a marginal temporal increase in pH. We observed that the pH of the ventral rumen was delayed by approximately 37 min compared with that of the reticulum, which was therefore considered in the modeling. With the models developed in this study, 67.0% of the variance of the reticular pH curves and 37.8% of the variance of the ruminal pH curves could be explained by fixed effects. We deduced that the diurnal pH course is, to a large extent, associated with the animal's individual feed intake and rumination behavior.

Proteasome activity and expression of mammalian target of rapamycin signaling factors in skeletal muscle of dairy cows supplemented with conjugated linoleic acids during early lactation.

The mammalian target of rapamycin (mTOR) is a major regulator of protein synthesis via its main downstream effectors, ribosomal protein S6 kinase (S6K1) and eukaryotic initiation factor 4E binding protein (4EBP1). The ubiquitin-proteasome system (UPS) is the main proteolytic pathway in muscle, and the muscle-specific ligases tripartite motif containing 63 (TRIM63; also called muscle-specific ring-finger protein 1, MuRF-1) and F-box only protein 32 (FBXO32; also called atrogin-1) are important components of the UPS. We investigated 20S proteasome activity and mRNA expression of key components of mTOR signaling and UPS in skeletal muscle of dairy cows during late gestation and early lactation and tested the effects of dietary supplementation (from d 1 in milk) with conjugated linoleic acids (sCLA; 100 g/d; n = 11) compared with control fat-supplemented cows (CTR; n = 10). Blood and muscle tissue (semitendinosus) samples were collected on d -21, 1, 21, and 70 relative to parturition. Dry matter intake increased with time of lactation in both groups. It was lower in sCLA than in CTR on d 21, which resulted in a reduced calculated metabolizable protein balance. Most serum and muscle concentrations of AA followed time-related changes but were unaffected by CLA supplementation. In both groups, serum and muscle 3-methylhistidine (3-MH) concentrations and the ratio of 3-MH:creatinine increased from d -21 to d 1, followed by a decline on d 21. The mRNA abundance of MTOR on d 21 and 70 was greater in sCLA than in CTR. The abundance of 4EBP1 mRNA did not differ between groups but was upregulated in both on d 1. The mRNA abundance of S6K1 on d 70 was greater in CTR than in sCLA, but remained unchanged over time in both groups. The mRNA abundance of FBXO32 (encoding atrogin-1) on d 21 was greater in sCLA than in CTR. The mRNA abundance of TRIM63 (also known as MuRF1) showed a similar pattern as FBXO32 in both groups: an increase from d -21 to d 1, followed by a decline. The mRNA for the α (BCKDHA) and ß (BCKDHB) polypeptide of branched-chain α-keto acid dehydrogenase was elevated in sCLA and CTR cows on d 21, respectively, suggesting a role of CLA in determining the metabolic fate of branched-chain AA. For the mTOR protein, no group differences were observed. The abundance of S6K1 protein was greater across all time points in sCLA versus CTR. The antepartum 20S proteasome activity in muscle was elevated in both groups compared with postpartum, probably reflecting the start of protein mobilization before parturition. Plasma insulin concentrations decreased in both groups postpartum but to a greater extent in CTR than in sCLA, resulting in greater insulin concentrations in sCLA than in CTR. Thus, the greater abundance of MTOR mRNA and S6K1 protein in sCLA compared with CTR might be mediated by the greater plasma insulin postpartum. The upregulation of MTOR mRNA in sCLA cows on d 21, despite greater FBXO32 mRNA abundance, may reflect a simultaneous activation of both anabolic and catabolic signaling pathways, likely resulting in greater protein turnover.

Changes in tissue abundance and activity of enzymes related to branched-chain amino acid catabolism in dairy cows during early lactation.

Branched-chain α-keto acid dehydrogenase (BCKDH) complex catalyzes the irreversible oxidative decarboxylation of branched-chain α-keto acids. This reaction is considered as the rate-limiting step in the overall branched-chain amino acid (BCAA) catabolic pathway in mammals. For characterizing the potential enzymatic involvement of liver, skeletal muscle, adipose tissue (AT), and mammary gland (MG) in BCAA metabolism during early lactation, tissue and blood samples were examined on d 1, 42, and 105 after parturition from 25 primiparous Holstein cows. Serum BCAA profiles were analyzed and the mRNA and protein abundance as well as the activity in the different tissues were assessed for the BCAA catabolic enzymes, partly for the branched-chain aminotransferase and completely for BCKDH. Total BCAA concentration in serum was lowest on d 1 after parturition and increased thereafter to a steady level for the duration of the experiment. Pronounced differences between the tissues were observed at all molecular levels. The mRNA abundance of the mitochondrial isoform of branched-chain aminotransferase (BCATm) was greatest in AT as compared with the other tissues studied, indicating that AT might be an important contributor in the initiation of BCAA catabolism in dairy cows. From the different subunits of the BCKDH E1 component, only the mRNA for the ß polypeptide (BCKDHB), not for the α polypeptide (BCKDHA), was elevated in liver. The BCKDHA mRNA abundance was similar across all tissues except muscle, which tended to lower values. Highest BCKDHA protein abundance was observed in both liver and MG, whereas BCKDHB protein was detectable in these tissues but could not be quantified. Adipose tissue and muscle only displayed abundance of the α subunit, with muscle having the lowest BCKDHA protein of all tissues. We found similarities in protein abundance for both BCKDH E1 subunits in liver and MG; however, the corresponding overall BCKDH enzyme activity was 7-fold greater in liver compared with MG, allowing for hepatic oxidation of BCAA transamination products. Reduced BCKDH activity in MG associated with no measurable activity in AT and muscle may favor sparing of BCAA for the synthesis of the different milk components, including nonessential AA. Deviating from previously published data on BCAA net fluxes and isotopic tracer studies in ruminants, our observed results might in part be due to complex counter-regulatory mechanisms during early lactation.

Impact of diet composition and temperature-humidity index on water and dry matter intake of high-yielding dairy cows.

The temperature-humidity index (THI) is widely used to characterize heat stress in dairy cattle. Diet composition is known to induce variation in metabolic-associated heat production. However, the relationships between THI and diet are poorly characterized with regard to performance and intake behaviour. Therefore, the objectives were to evaluate the impact of THI on water intake (WI), dry matter intake (DMI) and the frequency of drinking and feeding bouts in lactating dairy cows offered four dietary treatments: each contained 20% grass silage and additionally (i) 20% maize silage, 60% concentrate (M-HC); (ii) 60% maize silage, 20% concentrate (M-LC); (iii) 20% pressed beet pulp silage, 60% concentrate (BPS-HC); or (iv) 60% pressed beet pulp silage, 20% concentrate (BPS-LC) (DM basis). Individual WI and DMI were recorded from April to July 2013. Furthermore, dietary effects on milk production and reticular pH were estimated. Milk yield was lowest for M-LC, while energy-corrected milk was similar for all diets. Milk fat percentage was higher and milk protein amount lower for cows offered both LC diets. Reticular pH below 6.3, 6.0 and 5.8 lasted longest for BPS-LC. WI was higher for HC diets. However, the frequency of drinking bouts was not influenced by the ration. Lower DMI occurred for BPS-LC compared to M-LC. Frequency of feeding bouts was significantly higher for LC diets. THI was significantly related to WI, DMI as well as drinking and feeding bouts. Per increasing THI, WI increased slightly more for LC diets and DMI decreased more for HC diets. Frequency of drinking bouts increased slightly higher for BPS rations per rising THI, while the decrease in feeding bouts was highest for M-HC. In conclusion, TMR composition and moderate heat stress impacted WI and DMI of dairy cows, while both dietary energy density and ruminal filling might intensify the THI impact.

Influence of conjugated linoleic acids and vitamin E on milk fatty acid composition and concentrations of vitamin A and α-tocopherol in blood and milk of dairy cows.

The objective of this trial was to investigate the influences of conjugated linoleic acid (CLA) and vitamin E (Vit. E) and their interactions on fatty acid composition and vitamins in milk (α-tocopherol, retinol and ß-carotene) as well as on α-tocopherol in blood of pluriparous cows from week 6 ante partum until week 10 post-partum (p.p.). We assigned 59 pluriparous German Holstein cows to four treatment groups with the treatment factors CLA and Vit. E at two levels in a 2 × 2 factorial design. Milk fatty acid composition and milk vitamins were analysed on lactation days 7 and 28. α-tocopherol in blood serum was analysed on days -42, -7, 1, 7, 14, 28 and 70 relative to parturition. Milk concentration of α-tocopherol was influenced by Vit. E (p < .001) and CLA (p = .034). Percentage of cis-9, trans-11 CLA in total milk fat was influenced by treatment with CLA (p < .001), while for percentage of trans-10, cis-12 CLA an interaction between treatment and day (p = .019), driven by an increase in both CLA groups from day 7 to day 28, was found. Serum ratios of α-tocopherol to cholesterol were influenced by Vit. E (p < .001). Results suggest that treatment with CLA during late pregnancy and early lactation is suitable to enhance the proportion of trans-10, cis-12 CLA in milk and thereby influencing nutritional properties. As treatment with Vit. E did not have an impact on milk fatty acid composition, it might be possible to increase the antioxidative capacity of the dairy cow without affecting milk properties. Consequently, combined treatment with CLA and Vit. E might elicit synergistic effects on the cow and milk quality by increasing the proportion of CLA in milk fat as well as the excretion of Vit. E and the Vit. E levels in serum.

Influence of conjugated linoleic acids and vitamin E on biochemical, hematological, and immunological variables of dairy cows during the transition period.

The objective of this experiment was to determine the effects of conjugated linoleic acid (CLA) and vitamin E as well as their interaction on biochemical and hematological variables and on leukocyte populations and their functionality. We assigned 59 German Holstein cows between the 2nd and 9th lactation to 4 dietary groups in a 2 × 2 factorial design with the factors CLA and vitamin E. Six weeks before calving the cows had a BCS of 3.7 to provoke a higher risk of developing ketosis, which might impair their immune function. Blood samples for analyses were taken on d -42, -14, -7, -3, 1, 3, 7, 10, 14, 21, 28, 35, 42, 56, and 70 relative to parturition. Furthermore, peripheral blood mononuclear cells were cultured on d -42, -7, 1, 7, 14, 28, and 70 relative to calving. Most variables were characterized by a high variation between d 7 antepartum and d 7 postpartum. Treatments did not elicit any effect, with the exception of vitamin E, which increased serum urea concentrations and decreased monocyte percentages. Haptoglobin, aspartate-aminotransferase, red blood cell count, leukocyte percentage and populations, as well as peripheral blood mononuclear cells were influenced by parity. In conclusion, the impairment of immune function caused by calving was more severe in cows in ≥3rd parity than in younger cows. However, neither vitamin E nor CLA supplementation was successful to stabilize parity or parturition related variance in hematological and immunological traits.

Effects of body condition, monensin, and essential oils on ruminal lipopolysaccharide concentration, inflammatory markers, and endoplasmatic reticulum stress of transition dairy cows.

Evidence exists that dairy cows experience inflammatory-like phenomena in the transition period. Rumen health and alterations in metabolic processes and gene networks in the liver as the central metabolic organ might be key factors for cows' health and productivity in early lactation. This study made use of an animal model to generate experimental groups with different manifestations of postpartal fat mobilization and ketogenesis. In total, 60 German Holstein cows were allocated 6 wk antepartum to 3 high-body condition score (BCS) groups (BCS 3.95) and 1 low-BCS group (LC; BCS 2.77). High-BCS cows were fed an antepartal forage-to-concentrate ratio of 40:60 on dry matter basis, in contrast to 80:20 in the LC group, and received a monensin controlled-release capsule (HC/MO), a blend of essential oils (HC/EO), or formed a control group (HC). We evaluated serum haptoglobin, kynurenine, tryptophan, ruminal lipopolysaccharide concentration and mRNA abundance of nuclear factor kappa B (NF-κB), nuclear factor E2-related factor 2 (Nrf2), and endoplasmatic reticulum stress-induced unfolded protein response (UPR) target genes in liver biopsy samples from d -42 until +56 relative to calving. Nearly all parameters were highly dependent on time, with greatest variation near calving. The ruminal lipopolysaccharide concentration and evaluated target genes were not generally influenced by antepartal BCS and feeding management. The kynurenine-to-tryptophan ratio was higher in LC than in HC/MO treatment on d 7. Ruminal lipopolysaccharide concentration was higher in HC/MO than in the HC group, but not increased in HC/EO group. Abundance of UPR target gene X-box binding protein 1 was higher in HC/MO than in HC/EO group on d 7. Hepatic mRNA abundance of Nrf2 target gene glutathione peroxidase 3 was higher, whereas expression of NF-κB target gene haptoglobin tended to be higher in LC than in HC/EO cows. The HC/MO cows showed the most prominent increase in the abundance of glutathione peroxidase 3 and haptoglobin after calving in comparison to antepartal values. Results indicate the presence of inflammatory-like phenomena near calving. Simultaneously, alterations in UPR and Nrf2 target genes with antioxidative properties and haptoglobin occurred, being most prominent in LC and HC/MO group.

Influence of conjugated linoleic acid and vitamin E on performance, energy metabolism, and change of fat depot mass in transitional dairy cows.

The objective of this experiment was to determine the effects of conjugated linoleic acid (CLA) and vitamin E as well as their interaction on performance variables and lipomobilization during late pregnancy and early lactation (wk 6 antepartum until wk 10 postpartum). For this purpose, 59 pluriparous German Holstein cows were assigned to 4 dietary groups in a 2 × 2 design with the factors CLA and vitamin E at 2 levels. For this trial, we selected cows with a high body condition score because they are more likely to mobilize fat and consequently are at a higher risk of developing ketosis. Furthermore, concentrate proportions were adjusted to provoke ketosis. Lactation performance variables were analyzed in 3 periods (d 42 antepartum until calving, 1 to 21 d in milk, 22 to 70 d in milk). Dry matter intake and net energy intake were reduced in animals receiving CLA. Milk fat content was reduced in the CLA group compared with the control group (4.83 vs. 5.46% in period 2; 3.36 vs. 4.57% in period 3). In the vitamin E and the CLA + vitamin E groups, reduction of milk fat content was observed in period 3 (3.76 vs. 4.57% compared with the control group). Milk yield was not affected by treatment. ß-Hydroxybutyrate concentrations and liver lipid contents were not influenced by CLA or vitamin E. Moreover, longitudinal changes of adipose tissue depot mass were not affected by dietary treatments. Results suggest that the effects CLA had on milk composition were compensated by an increased milk yield and a decreased dry matter intake. Reduced milk energy output in CLA-treated animals was compensated by a reduced dry matter intake. Therefore, the net energy balance was not affected by either treatment. Consequently, we found no group effect on the mobilization of adipose tissue.

Effects of monensin and essential oils on immunological, haematological and biochemical parameters of cows during the transition period.

Using a model to generate experimental groups with different manifestations of post-partum (p.p.) fat mobilization and ketogenesis, the effects of a dietary and a medical intervention on biochemical and haematological parameters, antibody titre, leucocytes subsets and function of transition cows were examined. In total, 60 German Holstein cows were allocated 6 weeks antepartum (a.p.) to 3 high-body condition score (BCS) groups (BCS 3.95) and 1 low-BCS group (LC, BCS 2.77). High-BCS cows received a monensin controlled-release capsule (HC/MO) or a blend of essential oils (HC/EO) or formed a control group (HC). Parameters were evaluated in 3 periods (day (d) -42 until calving, 1 until 14 days in milk (DIM), 15 until 56 DIM). Over the course of trial, various parameters were influenced by period with greatest variability next to calving. White blood cell count was higher in the HC (8.42 × 103 /µl) and HC/EO (8.38 × 103 /µl) groups than in the HC/MO group (6.81 × 103 /µl) considering the whole trial. Supplementation of monensin decreased aspartate aminotransferase in comparison with the HC group similar to LC treatment. Bilirubin concentration was nearly doubled in all high-BCS cows in period 2. In period 3, essential oils increased γ-glutamyltransferase (80.4 Units/l) in comparison with all other groups and glutamine dehydrogenase (61 Units/l) in comparison with the LC (19 Units/l) and the HC/MO group (18 Units/l). Results suggest that parameters were generally characterized by a high variability around calving. Based on biochemical characteristics, it appeared that the HC cows seemed to have compromised hepatocyte integrity when compared to the LC cows. From the immune parameters investigated, the BVDV antibody response was more pronounced in HC/MO compared to HC/EO.

Excretion pathways and ruminal disappearance of glyphosate and its degradation product aminomethylphosphonic acid in dairy cows.

From 6 balance experiments with total collection of feces and urine, samples were obtained to investigate the excretion pathways of glyphosate (GLY) in lactating dairy cows. Each experiment lasted for 26d. The first 21d served for adaptation to the diet, and during the remaining 5d collection of total feces and urine was conducted. Dry matter intake and milk yield were recorded daily and milk and feed samples were taken during the sampling periods. In 2 of the 6 experiments, at the sampling period for feces and urine, duodenal contents were collected for 5d. Cows were equipped with cannulas at the dorsal sac of the rumen and the proximal duodenum. Duodenal contents were collected every 2h over 5 consecutive days. The daily duodenal dry matter flow was measured by using chromium oxide as a volume marker. All samples (feed, feces, urine, milk and duodenal contents were analyzed for GLY and aminomethylphosphonic acid (AMPA). Overall, across the 6 experiments (n=32) the range of GLY intake was 0.08 to 6.67mg/d. The main proportion (61±11%; ±SD) of consumed GLY was excreted with feces; whereas excretion by urine was 8±3% of GLY intake. Elimination via milk was negligible. The GLY concentrations above the limit of quantification were not detected in any of the milk samples. A potential ruminal degradation of GLY to AMPA was derived from daily duodenal GLY flow. The apparent ruminal disappearance of GLY intake was 36 and 6%. In conclusion, the results of the present study indicate that the gastrointestinal absorption of GLY is of minor importance and fecal excretion represents the major excretion pathway. A degradation of GLY to AMPA by rumen microbes or a possible retention in the body has to be taken into account.

The effects of a ration change from a total mixed ration to pasture on rumen fermentation, volatile fatty acid absorption characteristics, and morphology of dairy cows.

To investigate the effect of the change from a concentrate and silage-based ration (total mixed ration, TMR) to a pasture-based ration, a 10-wk trial (wk 1-10) was performed, including 10 rumen- and duodenum-fistulated German Holstein dairy cows (182±24 d in milk, 23.5±3.5kg of milk/d; mean ± standard deviation). The cows were divided in either a pasture group (PG, n=5) or a confinement group (CG, n=5). The CG stayed on a TMR-based ration (35% corn silage, 35% grass silage, 30% concentrate; dry matter basis), whereas the PG was gradually transitioned from a TMR to a pasture-based ration (wk 1: TMR only; wk 2: 3 h/d on pasture wk 3 and 4: 12 h/d on pasture wk 5-10: pasture only). Ruminal pH, volatile fatty acids (VFA), NH3-N, and lipopolysaccharide (LPS) concentrations were measured in rumen fluid samples collected medially and ventrally on a weekly basis. Ruminal pH was continuously recorded during 1 to 4 consecutive days each week using ruminal pH measuring devices. In wk 1, 5, and 10, rumen contents were evacuated and weighed, papillae were collected from 3 locations in the rumen, and subsequently a VFA absorption test was performed. In the PG, mean rumen pH and molar acetate proportions decreased, and molar butyrate proportions increased continuously over the course of the trial, which can most likely be ascribed to an increased intake of rapidly fermentable carbohydrates. During the first weeks on a full grazing ration (wk 5-7), variation of rumen pH decreased, and in wk 5 a lower rumen content, papillae surface area, and potential for VFA absorption were observed. In wk 8 to 10, variation of rumen pH and total VFA concentrations increased again, and acetate/propionate ratio decreased. In wk-10 rumen content, papillae area and VFA absorption characteristics similar to initial levels were observed. Although continuous rumen pH assessments and LPS concentrations did not reveal an increased risk for subacute rumen acidosis (SARA) during the adaption period, histopathology of rumen papillae and potential for VFA absorption indicated a possible risk for rumen health. An increased risk for SARA was observed in wk 9 and 10 in the PG, but rumen LPS concentrations and histopathology were not adversely affected. Results of the present study suggest that after behavioral and metabolic adaptation to the transition from a TMR to a pasture-based ration, no adverse effects on rumen morphology and absorption capacity occurred, although rumen pH after adaptation to pasture indicated increased risk of SARA.

Effect of monensin and essential oils on performance and energy metabolism of transition dairy cows.

This work examined preventive effects of a dietary and a medical intervention measure on postpartum (p.p.) ketogenesis in dairy cows overconditioned in late pregnancy. Sixty German Holstein cows were allocated 6 weeks antepartum (a.p.) to three high body condition score (BCS) groups (BCS 3.95 ± 0.08) and one low BCS group (LC, BCS 2.77 ± 0.14). Concentrate proportion in diet a.p. was higher (60% vs. 20%) and increase in proportion p.p. from 30% up to 50% decelerated (3 vs. 2 weeks) in high BCS groups. High BCS cows received a monensin controlled-release capsule (CRC) (HC/MO), a blend of essential oils (HC/EO) or formed a control group (HC). Performance parameters and energy status were evaluated in three periods [day (d) -42 until calving, one until 14 days in milk (DIM), 15 until 56 DIM]. Feed efficiency was 65% and 53% higher in HC/MO than in LC (p < 0.001) and HC groups (p = 0.002) in the second period. Milk fat content was higher in HC/EO (5.60 vs. 4.82%; p = 0.012) and milk urea higher in HC/MO (135 mg/kg) than in LC cows (107 mg/kg; p < 0.001). Increased p.p. levels of non-esterified fatty acids in serum were found in HC (p = 0.003), HC/MO (p = 0.068) and HC/EO (p = 0.002) in comparison with LC cows. Prevalence of subclinical and clinical ketosis was 54% and 46%, respectively, in HC group. Monensin decreased the prevalence to 50% and 7% respectively. Ruminal fermentation pattern showed higher proportions of propionate (23.43 mol % and 17.75 mol %, respectively; p < 0.008) and lower acetate:propionate ratio (2.66 vs. 3.76; p < 0.001) in HC/MO than HC group. Results suggest that a monensin CRC improved energy status and feed efficiency of transition dairy cows while essential oils failed to elicit any effect.

Short communication: aquaporin-7 mRNA in adipose depots of primiparous and pluriparous dairy cows: long-term physiological and conjugated linoleic acid-induced changes.

Aquaglyceroporins act as channel proteins and regulate water and glycerol exchange through cell membranes. The aquaglyceroporin aquaporin-7 (AQP7) is abundantly expressed in adipose tissue (AT) and regulates the release of glycerol produced by lipolysis. We aimed to investigate the expression of AQP7 mRNA during lactation in subcutaneous (s.c.) and visceral (v.c.) adipose depots of primiparous and pluriparous dairy cows. In 2 independent experiments, Holstein cows were supplemented with conjugated linoleic acids (CLA) or a control (CON) fat supplement at 100g/d. Pluriparous cows were supplemented starting with the first day in milk (DIM) up to 182 DIM and biopsies from s.c. AT were collected at d -21, 1, 21, 70, 105 182 196, 224, and 252 relative to calving (CLA=11; CON=10). Samples from 3s.c. and v.c. adipose depots were investigated in primiparous cows (n=25) receiving the supplements from 1 DIM until slaughter at 1, 42, or 105 DIM. The AQP7 mRNA abundance decreased from d -21 to 1 in s.c. AT of pluriparous cows without further increase to d 252 of lactation. In primiparous cows of the CON group, the AQP7 mRNA abundance increased from 1 to 105 DIM in s.c. AT from the tail head and in mesenteric AT. In retroperitoneal AT, the only depot for which a significant decrease in mass was observed with DIM, AQP7 mRNA abundance was greater at 42 and 105 than 1 DIM. Comparing the different fat depots, retroperitoneal AT had the highest and mesenterial AT had the lowest AQP7 mRNA abundance, but no general difference was observed between v.c. and s.c. fat depots. The values were not affected by CLA treatment with the exception of mesenteric AT, for which lower AQP7 mRNA abundance values were recorded in the CLA than in the CON group. The longitudinal characterization of the AQP7 mRNA expression profile throughout lactation revealed differences between primiparous and pluriparous cows, with an increase of AQP7 mRNA abundance up to 105 DIM only in the primiparous cows. Due to a lack of CLA effects in pluriparous cows and the limitation to just one fat depot in primiparous cows, a modulatory effect of CLA on AQP7 mRNA abundance in dairy cows is not supported by our study.

Immunohistochemical characterization of phagocytic immune cell infiltration into different adipose tissue depots of dairy cows during early lactation.

The present study aimed to investigate whether phagocytic immune cells infiltrate into bovine adipose tissue (AT) and to study the effects of lactation and conjugated linoleic acid (CLA) supplementation on the invasion of phagocytic immune cells into different s.c. and visceral (v.c.) fat depots of primiparous dairy cows during the first 105 d in milk (DIM). German Holstein-Friesian cows (HF; n = 25) with a mean body condition score of 3.0 were divided into a control (CON) and a CLA group. From 1 DIM until sample collection, CLA cows were fed 100 g of CLA supplement/d (about 6% of c9,t11 and t10,c12 isomers each), whereas the CON cows received 100 g/d of a fatty acid mixture instead of CLA. The CON cows (n = 5 each) were slaughtered at 1, 42, and 105 DIM, and the CLA cows (n = 5 each) were slaughtered at 42 and 105 DIM. Adipose tissues (n = 150) from 3s.c. (tailhead, withers, and sternum) and 3 v.c. (omental, mesenteric, and retroperitoneal) depots were sampled. In addition, s.c. tailhead biopsies were collected by repeated surgical biopsies (3 samplings within 7 wk; n = 36) from 12 nonpregnant, nonlactating Simmental heifers (SM; mean body condition score = 5.0) fed diets of varying energy density to compare the changes in phagocytic immune cell infiltration with early lactating cows. Immunohistochemical analyses of different fat depots revealed a low incidence of phagocytic immune cell infiltration in early lactating cows. The portion of infiltrating macrophages (CD68+) in a few positive AT samples of HF cows was slightly lower in s.c. than v.c. fat and was positively correlated with both empty body weight and adipocyte size. However, no differences with regard to DIM and CLA supplementation were observed in HF cows. Increased accumulation of phagocytic immune cells, albeit at low cell numbers, in nonpregnant, over-conditioned SM heifers might be related to larger adipocytes secreting higher amounts of chemoattractant adipokines compared with the early lactating cows. In conclusion, the extent of fatness in HF cows may not be high enough to stimulate significant infiltration of phagocytic cells in AT and, therefore, these immune cells might have no major role in the immunologic and metabolic adaptations of AT during early lactation.

Bovine haptoglobin as an adipokine: serum concentrations and tissue expression in dairy cows receiving a conjugated linoleic acids supplement throughout lactation.

The present study aimed to characterize serum haptoglobin (Hp) concentrations throughout an entire lactation period in both primi- and multiparous cows and to compare them to the Hp mRNA expression in liver and - in view of Hp being potentially an adipokine - also in different subcutaneous (s.c.) and visceral fat depots. In addition, potential anti-inflammatory effects of long-term supplementation with conjugated linoleic acids (CLA) were evaluated by assessing Hp. Trial 1 comprised 33 cows and 16 Holstein heifers from day 21 ante partum until day 252 postpartum. The animals received 100 or 50 g/day CLA or a control fat supplement. Blood samples and biopsy (tail head fat and liver) samples were collected. Trial 2 included 25 Holstein heifers, 5 animals were slaughtered on the day of parturition, the remaining animals were allocated to either CLA (100 g/day, n=10) or control fat supplement (n=10) and slaughtered on days 42 and 105 postpartum, respectively. At slaughter, fat samples were collected from 3 different visceral depots, 3 s.c. depots and from liver tissue. Results indicated no effects of CLA on serum Hp and liver Hp mRNA for both trials and on Hp mRNA in biopsies from s.c. tail head fat. In omental and s.c. withers fat from trial 2, CLA reduced Hp mRNA on both day 42 and day 105. Hp mRNA was detectable in fat tissues from both trials with abundance values being significantly lower than in liver. The Hp mRNA abundance in the s.c. fat depots was generally higher than in the visceral depots. Haptoglobin mRNA abundance in the different tissues from trial 2 was correlated whereby all s.c. depots were interrelated. The evidence of Hp mRNA expression in adipose tissues and the presence of Hp-immune staining in histological fat sections confirm that Hp can be classified as a bovine adipokine. The lack of an evident relationship between circulating Hp concentrations and normal body fat portions in dairy cattle demonstrates that varying degrees of adiposity are not confounding factors when using Hp as inflammatory marker. The physiological changes in serum Hp concentration seem to be limited to parity and parturition. In view of the lack of effects of CLA on serum Hp concentrations, the observed reaction in two out of six different fat depots seems of marginal importance for the organisms as an entity.

Effect of conjugated linoleic acid supplementation on body composition, body fat mobilization, protein accretion, and energy utilization in early lactation dairy cows.

The objective of the study was to investigate the effects of trans-10,cis-12 and cis-9,trans-11 conjugated linoleic acid (CLA) supplementation on body composition, mobilization or accretion of body fat and protein mass, as well as the energy metabolism of dairy cows during the first 105 d in milk (DIM). For this purpose, a comparative slaughter experiment was conducted with 25 primiparous German Holstein cows. The experiment started at 1 DIM with the slaughter of 5 animals of an initial group receiving no CLA supplement. The remaining animals were fed a CLA supplement (n=10) or a stearic acid-based control fat supplement (CON; n=10) from 1 DIM up to slaughter. After 42 DIM, 5 more cows from each treatment (42-CLA and 42-CON) were slaughtered. The remaining 5 cows in each treatment were slaughtered after 105 DIM (105-CLA and 105-CON). The animals of the CLA groups consumed 6.0 g/d of trans-10,cis-12 CLA and 5.7 g/d of cis-9,trans-11 CLA. During the slaughter process, the empty body mass was recorded and partitioned into 9 fractions (meat, bone, offal, hide, mammary gland, retroperitoneal fat, omental fat, mesenteric fat, and s.c. fat). The fractions were analyzed for dry matter, ether extract, crude protein, and ash to calculate the body composition of the empty body mass at the different slaughter times. The principle of the comparative slaughter technique was applied to estimate body fat or protein mobilization and accretion in the viewed periods from 1 DIM until 42 and 105 DIM. The heat production (HP) was calculated by subtracting the energy in milk and energy changes in body mass from the metabolizable energy intake. The body composition was not affected by CLA supplementation. However, the mobilization of body fat mass from 1 until 42 DIM was 24.1 kg in the 42-CON group and 14.3 kg in the 42-CLA group. This resulted in a trend to lower body mass (fat and protein) mobilization of 10.5 kg in the 42-CLA group. Energy mobilization from body mass was 21.2 MJ/d in the 42-CON and 11.5 MJ/d in the 42-CLA group. The HP was unchanged for the 42-CON and 42-CLA group with 123.0 and 116.9 MJ/d, respectively. From 1 until 105 DIM, the protein accretion was 4.3 kg and the daily energy retention in body protein was 1.0 MJ higher for CLA-supplemented cows. The HP was decreased in this period for the 105-CLA group (115.5 MJ/d) as compared with the 105-CON group (125.9 MJ/d). Overall, the trend for a decreased body mass mobilization suggested a protective effect of CLA supplementation against excessive use of body reserves within 42 DIM. Continuous CLA supplementation until 105 DIM increased protein accretion. The effects on body mass mobilization and protein accretion in combination with the decreased HP in the CLA-fed cows suggested a more efficient utilization of metabolizable energy in CLA-supplemented early lactation dairy cows.

Effect of trans-10, cis-12 conjugated linoleic acid on performance, adipose depot weights, and liver weight in early-lactation dairy cows.

In feeding practice, conjugated linoleic acid (CLA) supplements are used to decrease milk fat excretion in early-lactation dairy cows to save energy to counteract the physiological negative energy balance. The present study was conducted to examine the effects of CLA on energy metabolism, changes in liver weight, and the weight of different adipose depots during early lactation. Primiparous lactating German Holstein cows (n=25) were divided into 5 groups and each group contained 5 animals. The experiment started 21 d prepartum and continued until 105 d in milk (DIM). Cows were slaughtered at 1, 42, and 105 DIM. The experiment was divided into a prepartum period (21 d prepartum until calving), period 1 (1 until 42 DIM), and period 2 (>42 until 105 DIM). In the prepartum period, all animals were housed together and fed the same diet with no CLA supplementation. At 1 DIM, an initial group, with no CLA supplementation, was slaughtered. The 20 remaining cows were assigned to 2 diets. One group received 100g/d of a control fat supplement (CON; n=10) and the other group 100g/d of a CLA supplement (CLA; n=10) from 1 DIM until slaughter. Five cows of each feeding group were slaughtered after 42 DIM and the remaining animals after 105 DIM. The CLA supplement contained approximately 10% each of trans-10, cis-12 CLA and cis-9, trans-11 CLA. During the slaughter process the empty body weight was recorded and the omental, mesenteric, retroperitoneal, and s.c. adipose depots, as well as the liver, were dissected and weighed. The CLA treatment decreased milk fat content in period 1 (14.1%). In period 2, milk fat content (25.4%) and yield (17.1%) were lower in the CLA group. No effect of CLA on milk yield was observed. The net energy intake, milk energy output, and the calculated energy balance remained unchanged by CLA supplementation. No effect of CLA on the weights of liver, omental, mesenteric, or s.c. adipose depots was observed when related to empty body weight. Liver weight increased with DIM, whereas the retroperitoneal adipose depot weight decreased at the same time. Compared with the initial group, the retroperitoneal adipose depot weight for control animals slaughtered after 42 DIM was decreased (47.7%); however, for the CLA group slaughtered after 42 DIM, a trend to a lower retroperitoneal adipose depot weight (34.0%) was observed. This suggests a CLA-induced deceleration of mobilization of the retroperitoneal adipose depot during the first 42 DIM.

Physiological and conjugated linoleic acid-induced changes of adipocyte size in different fat depots of dairy cows during early lactation.

The aim of this study was to investigate the effects of lactation and conjugated linoleic acid (CLA) supplementation on adipocyte sizes of subcutaneous (s.c.) and visceral (VC) fat depots in primiparous dairy cows during the first 105 d in milk (DIM). German Holstein heifers (n=25) were divided into a control (CON) and a CLA group. From 1 DIM until sample collection, CLA cows were fed 100g of CLA supplement/d (about 6% of c9,t11 and t10,c12 isomers each), whereas the CON cows received 100g of fatty acid mixture/d instead of CLA. The CON cows (n=5 each) were slaughtered at 1, 42, and 105 DIM, and the CLA cows (n=5 each) were slaughtered at 42 and 105 DIM. Adipose tissues from 3s.c. depots (tailhead, withers, and sternum) and from 3 VC depots (omental, mesenteric, and retroperitoneal) were sampled. Hematoxylin-eosin staining was done to measure adipocyte area (µm(2)). Retroperitoneal adipocyte sizes were mostly larger than adipocytes from the other sites, independent of lactation time and treatment. Significant changes related to duration of lactation were limited to retroperitoneal fat: adipocyte sizes were significantly smaller at 105 DIM than at 1 DIM in CON cows. Adipocyte sizes were decreased in s.c. depots from the tailhead at 105 DIM and from the sternum at 42 DIM in CLA versus CON cows, whereas for VC depots, adipocyte sizes were decreased in mesenteric fat at 42 and 105 DIM, and in omental and retroperitoneal fat, at 105 DIM in CLA versus CON cows. Within the CLA group, adipocyte sizes were smaller in the s.c. depot from the tailhead at 105 DIM than at 42 DIM. Adipocyte sizes and depot weights were significantly correlated in s.c. depots (r=0.795) in the CLA group and in retroperitoneal fat both in the CON (r=0.698) and the CLA (r=0.723) group. In conclusion, CLA-induced decreases in adipocyte size indicate lipolytic or antilipogenic effects of CLA, or both effects, on adipose tissue in primiparous dairy cows.