HBCS Cows Research Articles

Bile acid profiles and messenger RNA expression of bile acid-related genes in the liver of dairy cows with high versus normal body condition

Bile acids (BA) play a crucial role not only in lipid digestion but also in the regulation of overall energy homeostasis, including glucose and lipid metabolism. The aim of this study was to investigate BA profiles and mRNA expression of BA-related genes in the liver of high versus normal body condition in dairy cows. We hypothesized that body condition and the transition from gestation to lactation affect hepatic BA concentrations as well as the mRNA abundance of BA-related receptors, regulatory enzymes, and transporters. Therefore, we analyzed BA in the liver as well as the mRNA abundance of BA-related synthesizing enzymes, transporters, and receptors in the liver during the transition period in cows with different body conditions around calving. In a previously established animal model, 38 German Holstein cows were divided into groups with high body condition score (BCS) (HBCS; n = 19) or normal BCS (NBCS; n = 19) based on BCS and backfat thickness (BFT). Cows were fed diets aimed at achieving the targeted differences in BCS and BFT (NBCS: BCS <3.5, BFT <1.2 cm; HBCS: BCS >3.75, BFT >1.4 cm) until they were dried off at wk 7 before parturition. Both groups were fed identical diets during the dry period and subsequent lactation. Liver biopsies were taken at wk -7, 1, 3, and 12 relative to parturition. For BA measurement, a targeted metabolomics approach with LC-ESI-MS/MS was used to analyze BA in the liver. The mRNA abundance of targeted genes related to BA-synthesizing enzymes, transporters, and receptors in the liver was analyzed using microfluidic quantitative PCR. In total, we could detect 14 BA in the liver: 6 primary and 8 secondary BA, with glycocholic acid (GCA) being the most abundant one. The increase of glycine-conjugated BA after parturition, in parallel to increasing serum glycine concentrations may originate from an enhanced mobilization of muscle protein to meet the high nutritional requirements in early lactating cows. Higher DMI in NBCS cows compared with HBCS cows was associated with higher liver BA concentrations such as GCA, deoxycholic acid (DCA), and cholic acid (CA). The mRNA abundance of BA-related enzymes measured herein suggests the dominance of the alternative signaling pathway in the liver of HBCS cows. Overall, BA profiles and BA metabolism in the liver depend on both, the body condition and lactation-induced effects in periparturient dairy cows.

Bile acid profiles and mRNA abundance of bile acid-related genes in adipose tissue of dairy cows with high versus normal body condition

Besides their lipid-digestive role, bile acids (BA) influence overall energy homeostasis, such as glucose and lipid metabolism. We hypothesized that BA along with their receptors, regulatory enzymes, and transporters are present in subcutaneous adipose tissue (scAT). In addition, we hypothesized that their mRNA abundance varies with the body condition of dairy cows around calving. Therefore, we analyzed BA in serum and scAT as well as the mRNA abundance of BA-related enzymes, transporters, and receptors in scAT during the transition period in cows with different body conditions around calving. In a previously established animal model, 38 German Holstein cows were divided into either a high (HBCS; n = 19) or normal BCS (NBCS; n = 19) group based on their BCS and back-fat thickness (BFT). Cows were fed different diets to achieve the targeted differences in BCS and BFT (NBCS: BCS <3.5, BFT <1.2 cm; HBCS: BCS >3.75, BFT >1.4 cm) until dry-off at 7 wk antepartum. During the dry period and subsequent lactation, both groups were fed the same diets according to their energy demands. Using a targeted metabolomics approach via liquid chromatography-electrospray ionization-MS /MS, BA were analyzed in serum and scAT at wk -7, 1, 3, and 12 relative to parturition. In serum, 15 BA were observed: cholic acid (CA), chenodeoxycholic acid (CDCA), glycocholic acid (GCA), taurocholic acid (TCA), glycochenodeoxycholic acid (GCDCA), taurochenodeoxycholic acid, deoxycholic acid (DCA), lithocholic acid, glycodeoxycholic acid (GDCA), glycolithocholic acid, taurodeoxycholic acid, taurolithocholic acid, β-muricholic acid, tauromuricholic acid (sum of α and β), and glycoursodeoxycholic acid, whereas in scAT 7 BA were detected: CA, GCA, TCA, GCDCA, taurochenodeoxycholic acid, GDCA, and taurodeoxycholic acid. In serum and scAT samples, the primary BA CA and its conjugate GCA were predominantly detected. Increasing serum concentrations of CA, CDCA, TCA, GCA, GCDCA, DCA, and β-muricholic acid with the onset of lactation might be related to the increasing DMI after parturition. Furthermore, serum concentrations of CA, CDCA, GCA, DCA, GCDCA, TCA, lithocholic acid, and GDCA were lower in HBCS cows compared with NBCS cows, concomitant with increased lipolysis in HBCS cows. The correlation between CA in serum and scAT may point to the transport of CA across cell membranes. Overall, the findings of the present study suggest a potential role of BA in lipid metabolism depending on the body condition of periparturient dairy cows.

Body condition prepartum and its association with term placentome nutrient transporters, one‑carbon metabolism pathway activity, and intermediate metabolites in Holstein cows

We investigated linkages among BCS prior to calving and placentome concentrations of metabolites, proteins in one‑carbon metabolism (OCM) and protein synthesis, and nutrient transport. Multiparous Holstein cows retrospectively divided by prepartal BCS at −4 weeks relative to parturition into high BCS (HBCS = 3.58 ± 0.23; n = 9) or normal BCS (NBCS = 3.02 ± 0.17; n = 13) were used. BCS was assessed using a 5-point scale (1 = thin, 5 = fat). Four placentomes per cow were collected at delivery and frozen in liquid N. Western blotting was used for protein abundance. Cystathionine-β-synthase (CBS) and betaine-homocysteine-S-methyltransferase (BHMT) activity were measured via 14C assays. Amino acids (AA) and metabolites in OCM were measured by liquid chromatography mass spectrometry (LC-MS). Compared with NBCS cows, the cellular stress sensor p-eIF2α was more than 2-fold greater (P = 0.04) in HBCS. Abundance of the AA-catabolism enzyme branched-chain α-ketoacid dehydrogenase complex was lower (P = 0.05) in HBCS cows. Although BHMT activity did not differ, greater concentration of betaine (P = 0.01) and lower (P = 0.05) concentration of dimethylglycine in HBCS cows suggested reduced flux through the methionine cycle. Despite a lack of difference in CBS activity, lower concentrations of cystathionine (P = 0.03) and hypotaurine (P = 0.04) along with lower cysteine and the tendency for lower total GSH (P = 0.10) in HBCS cows suggested a decrease in transsulfuration. Overall, associations between OCM in placentomes and BCS at calving exist. Identifying mechanisms responsible for these effects merits further research.

MicroRNA profiling of subcutaneous adipose tissue in periparturient dairy cows at high or moderate body condition

A growing body of evidence shows that microRNA (miRNA), play important roles in regulating adipose tissue (AT) physiology and function. The objective was to characterize the AT miRNA profile in over-conditioned (HBCS, n = 19) versus moderate-conditioned (MBCS, n = 19) periparturient dairy cows. Tail-head subcutaneous AT biopsied on d -49 and 21 relative to parturition were used for miRNA sequencing. The miR-486 was the most significant miRNA among the upregulated miRNA on d -49, which might be related to more pronounced changes in lipogenesis and altered insulin sensitivity in AT of HBCS cows at dry-off. Comparing HBCS to MBCS on d 21, 23 miRNA were downregulated and 20 were upregulated. The predicted targets of upregulated differentially expressed (DE)-miRNA on d 21 were enriched in different pathways, including pathways related to lysosomes and peroxisomes. The predicted targets of downregulated DE-miRNA on d 21 were enriched in various pathways, including epidermal growth factor receptor, insulin resistance, hypoxia-inducible factor 1 signaling pathway, and autophagy. The results showed that over-conditioning was associated with changes in SCAT miRNA profile mainly on d 21, of which most were downregulated. The enriched pathways may participate in over-conditioning-associated metabolic challenges during early lactation.

88 Impacts of Body Condition Score at Initiation of the Breeding Season on Reproductive Performance and Overall Productivity of Bos Taurus and B. Indicus Beef Cows

Abstract Responses of Bos taurus (BT) and B. indicus (BI) beef cows were compared according to their body condition score (BCS) at initiation of breeding season. Cows were assigned to an estrus synchronization + AI protocol and classified as BCS &amp;lt; 5.0 (LBCS, BT, n = 158; BI, n = 248) or BCS ≥ 5.0 (HBCS; BT, n = 173; BI, n = 122). Pregnancy status was verified via transrectal ultrasonography at 30 and 40 d after AI for BI and BT cows, respectively. All BT cows were exposed to natural service for 50 d (beginning 18 d after AI), and all non-pregnant BI cows were immediately exposed to natural service for 60 d. Pregnancy rates to AI did not differ (P ≥ 0.16) between HBCS or LBCS cows (60.9 vs. 53.5% for BT; 37.0 vs. 31.4% for BI; respectively). Pregnancy rates to natural service were greater (P = 0.03) in HBCS BT cows compared with LBCS (82.7 vs. 67.9%; respectively) but did not differ in BI cows (78.9 vs. 71.5%; respectively). Calving rate was greater (P &amp;lt; 0.03) in HBCS cows, which also calved earlier (P &amp;lt; 0.01) compared with LBCS (93.7 vs. 85.1% in BT, 83.6 vs. 73.3% in BI; respectively). HBCS Cows weaned more (P ≤ 0.05) calves compared with LBCS, which were older (P ≤ 0.04) (215 vs. 211 d in BT, 219 vs. 209 d in BI; respectively) and heavier (P ≤ 0.09) at weaning (258 vs. 252 kg in BT, 213 vs. 203 d in BI; respectively). Lastly, kg of calf weaned/cow was greater (P ≤ 0.03) in HBCS cows compared with LBCS (221 vs. 200 d in BT, 159 vs. 129 d in BI; respectively). Therefore, HBCS cows had greater reproductive efficiency and overall productivity, and this was observed in both BT and BI.

Metabolic Profiling of Plasma in Different Calving Body Condition Score Cows Using an Untargeted Liquid Chromatography-Mass Spectrometry Metabolomics Approach.

Simple SummaryOver-conditioning around calving predisposes cows to a higher risk of postpartum metabolic disorders and diseases. Using untargeted liquid chromatography-mass spectrometry, seven metabolites were found that were significantly correlated to high body condition, as well as to classic metabolic parameters indicating changes in lipid metabolism and inflammation. These seven metabolites include six distinct lysophosphatidylcholines and a phosphatidylethanolamine. By investigating the depth of metabolic changes in the plasma of over-conditioned fresh cows, this study may provide a metabolic basis for the diseases associated with over-conditioning.This study was undertaken to identify metabolite differences in plasma of dairy cows with a normal or high calving body condition score (CBCS), using untargeted liquid chromatography-mass spectrometry (LC-MS) metabolomics. Sixteen multiparous dairy cows were assigned to one of two groups based on CBCS (0 to 5 scale): Normal group (NBCS, 3.25 ≤ BCS ≤ 3.5, n = 8), and high BCS group (HBCS, BCS ≥ 4, n = 8). Plasma samples were collected for metabolomics analysis and evaluation of biomarkers of lipid metabolism (nonesterified fatty acid (NEFA) and β-hydroxybutyrate (BHB)), and cytokines (leptin, adiponectin, tumor necrosis factor–α (TNF-α) and interleukin 6 (IL-6)). A total of 23 differential metabolites were identified, and functional analyses were performed using the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. Among these metabolites, the concentrations of six lysophosphatidylcholines and one phosphatidylethanolamine, were lower in the HBCS group than in the NBCS group (p < 0.01). Furthermore, these metabolites were involved in these four pathways, among others: glycerophospholipid metabolism, retrograde endocannabinoid signaling, autophagy, and glycosylphosphatidylinositol (GPI)-anchor biosynthesis (p < 0.05). In addition, plasma concentrations of leptin (p = 0.06) and TNF-α (p = 0.08) tended to be greater while adiponectin (p = 0.09) lower in HBCS cows than in NBCS cows. The concentrations of NEFA, BHB, or IL-6 did not differ between NBCS and HBCS groups. More importantly, based on the results of the Spearman’s correlation analysis, the seven important metabolites were negatively correlated with indices of lipid metabolisms, proinflammatory cytokines, and leptin, but positively correlated with adiponectin. These results demonstrate that CBCS has a measurable impact on the plasma metabolic profile, even when NEFA and BHB are not different. In addition, the identified differential metabolites were significantly correlated to lipid metabolism and inflammation in the over-conditioned fresh cows, which are expected to render a metabolic basis for the diseases associated with over-conditioned dry cows.

Molecular networks of insulin signaling and amino acid metabolism in subcutaneous adipose tissue are altered by body condition in periparturient Holstein cows

Peripartal cows mobilize not only body fat but also body protein to satisfy their energy requirements. The objective of this study was to determine the effect of prepartum BCS on blood biomarkers related to energy and nitrogen metabolism, and mRNA and protein abundance associated with AA metabolism and insulin signaling in subcutaneous adipose tissue (SAT) in peripartal cows. Twenty-two multiparous Holstein cows were retrospectively classified into a high BCS (HBCS; n = 11, BCS ≥ 3.5) or normal BCS (NBCS; n = 11, BCS ≤ 3.17) group at d 28 before expected parturition. Cows were fed the same diet as a total mixed ration before parturition and were fed the same lactation diet postpartum. Blood samples collected at -10, 7, 15, and 30 d relative to parturition were used for analyses of biomarkers associated with energy and nitrogen metabolism. Biopsies of SAT harvested at -15, 7, and 30 d relative to parturition were used for mRNA (real time-PCR) and protein abundance (Western blotting) assays. Data were subjected to ANOVA using the MIXED procedure of SAS (v. 9.4; SAS Institute Inc., Cary, NC), with P ≤ 0.05 being the threshold for significance. Cows in HBCS had greater overall plasma nonesterified fatty acid concentrations, due to marked increases at 7 and 15 d postpartum. This response was similar (BCS × Day effect) to protein abundance of phosphorylated (p) protein kinase B (p-AKT), the insulin-induced glucose transporter (SLC2A4), and the sodium-coupled neutral AA transporter (SLC38A1). Abundance of these proteins was lower at -15 d compared with NBCS cows, and either increased (SLC2A4, SLC38A1) or did not change (p-AKT) at 7 d postpartum in HBCS. Unlike protein abundance, however, overall mRNA abundances of the high-affinity cationic (SLC7A1), proton-coupled (SLC36A1), and sodium-coupled amino acid transporters (SLC38A2) were greater in HBCS than NBCS cows, due to upregulation in the postpartum phase. Those responses were similar to protein abundance of p-mTOR, which increased (BCS × Day effect) at 7 d in HBCS compared with NBCS cows. mRNA abundance of argininosuccinate lyase (ASL) and arginase 1 (ARG1) also was greater overall in HBCS cows. Together, these responses suggested impaired insulin signaling, coupled with greater postpartum AA transport rate and urea cycle activity in SAT of HBCS cows. An in vitro study using adipocyte and macrophage cocultures stimulated with various concentrations of fatty acids could provide some insights into the role of immune cells in modulating adipose tissue immunometabolic status, including insulin resistance and AA metabolism.

Profiling of circulating microRNA and pathway analysis in normal- versus over-conditioned dairy cows during the dry period and early lactation

The objective of this study was to determine the circulating microRNA (miRNA) profile in over-conditioned (HBCS) versus normal-conditioned (NBCS) dairy cows in combination with pathway enrichment analyses during the transition period. Thirty-eight multiparous Holstein cows were selected 15 wk before anticipated calving date based on their current and previous body condition scores (BCS) for forming either a HBCS group (n = 19) or a NBCS group (n = 19). They were fed different diets during late lactation to reach the targeted differences in BCS and backfat thickness until dry-off. A subset of 15 animals per group was selected based on their circulating concentrations of nonesterified fatty acids (on d 14 postpartum) and β-hydroxybutyrate (on d 21 postpartum), representing the greater or the lower extreme values within their BCS group. Blood serum obtained at d -49 and 21 relative to parturition (3 pools with 5 cows per each group and time point) were used to identify miRNA that were differentially expressed (DE) between groups or time points using miRNA sequencing. No DE-miRNA were discovered between NBCS versus HBCS. Comparing pooled samples from d -49 and d 21 resulted in 7 DE-miRNA in the NBCS group, of which 5 miRNA were downregulated and 2 miRNA were overexpressed on d 21 versus -49. The abundance of 5 of these DE-miRNA was validated in all individual samples via quantitative PCR and extended to additional time points (d -7, 3, 84). Group differences were observed for miR-148a, miR-122 as well as miR-455-5p, and most DE-miRNA (miR-148a, miR-122, miR-30a, miR-450b, miR-455-5p) were downregulated directly after calving. Subsequently, the DE-miRNA was used for bioinformatics analysis to identify putative target genes and the most enriched biological pathways. The most significantly enriched pathways of DE-miRNA were associated with cell cycle and insulin signaling as well as glucose and lipid metabolism. Overall, we found little differences in circulating miRNA in HBCS versus NBCS cows around calving.

Body condition alters glutathione and nuclear factor erythroid 2-like 2 (NFE2L2)–related antioxidant network abundance in subcutaneous adipose tissue of periparturient Holstein cows

Dairy cows with high body condition score (BCS) in late prepartum are more susceptible to oxidative stress (OS). Nuclear factor erythroid 2-like 2 (NFE2L2) is a major antioxidant transcription factor. We investigated the effect of precalving BCS on blood biomarkers associated with OS, inflammation, and liver function, along with mRNA and protein abundance of targets related to NFE2L2 and glutathione (GSH) metabolism in s.c. adipose tissue (SAT) of periparturient dairy cows. Twenty-two multiparous Holstein cows were retrospectively classified into a high BCS (HBCS; n = 11, BCS ≥3.5) or normal BCS (NBCS; n = 11, BCS ≤3.17) on d 28 before parturition. Cows were fed a corn silage- and wheat straw-based total mixed ration during late prepartum, and a corn silage- and alfalfa hay-based total mixed ration postpartum. Blood samples obtained at -10, 7, 15, and 30 d relative to parturition were used for analyses of biomarkers associated with inflammation, including albumin, ceruloplasmin, haptoglobin, and myeloperoxidase, as well as OS, including ferric reducing ability of plasma (FRAP), reactive oxygen species (ROS), and β-carotene. Adipose biopsies harvested at -15, 7, and 30 d relative to parturition were analyzed for mRNA (real-time quantitative PCR) and protein abundance (Western blotting) of targets associated with the antioxidant transcription regulator nuclear factor, NFE2L2, and GSH metabolism pathway. In addition, concentrations of GSH, ROS and malondialdehyde were measured. High BCS cows had lower prepartum dry matter intake expressed as a percentage of body weight along with greater BCS loss between -4 and 4 wk relative to parturition. Plasma concentrations of ROS and FRAP increased after parturition regardless of treatment. Compared with NBCS, HBCS cows had greater concentrations of FRAP at d 7 postpartum, which coincided with peak values in those cows. In addition, NBCS cows experienced a marked decrease in plasma ROS after d 7 postpartum, while HBCS cows maintained a constant concentration by d 30 postpartum. Overall, ROS concentrations in SAT were greater in HBCS cows. However, overall mRNA abundance of NFE2L2 was lower and cullin 3 (CUL3), a negative regulator of NFE2L2, was greater in HBCS cows. Although HBCS cows had greater overall total protein abundance of NFE2L2 in SAT, ratio of phosphorylated NFE2L2 to total NFE2L2 was lower, suggesting a decrease in the activity of this antioxidant system. Overall, mRNA abundance of the GSH metabolism-related genes glutathione reductase (GSR), glutathione peroxidase 1 (GPX1), and transaldolase 1 (TALDO1), along with protein abundance of glutathione S-transferase mu 1 (GSTM1), were greater in HBCS cows. Data suggest that HBCS cows might experience greater systemic OS after parturition, while increased abundance of mRNA and protein components of the GSH metabolism pathway in SAT might help alleviate tissue oxidant status. Data underscored the importance of antioxidant mechanisms at the tissue level. Thus, targeting these pathways in SAT during the periparturient period via nutrition might help control tissue remodeling while allowing optimal performance.

Effects of body condition on the insulin resistance, lipid metabolism and oxidative stress of lactating dairy cows

BackgroundOverconditioned dairy cows are prone to greater insulin resistance in transition to successfully adapt to negative energy balance. The associations among body condition score (BCS), insulin resistance, lipid metabolism and oxidative stress in cows during late lactation with positive energy balance remain to be elucidated.MethodsThe objectives of this study were to investigate insulin sensitivity and oxidative status in late lactating dairy cows with different BCS but similar milk production, parity and days in milk. Forty-two multiparous Holstein cows were fed the same diet under the same management and divided into three groups based on BCS: low BCS (LBCS; BCS ≤ 2.75; n = 12), medium BCS (MBCS; 3.0 ≤ BCS ≤ 3.5; n = 15) or high BCS (HBCS; BCS ≥ 3.75; n = 15). Blood samples used for analysis of biochemical and hematological parameters were collected from the coccygeal vein at the end of experiment.ResultsThe concentrations of insulin and nonesterified fatty acid were higher and the revised quantitative insulin sensitivity check index (RQUICKI) was lower in HBCS cows than in LBCS and MBCS cows. These results suggest that insulin resistance exacerbates lipolysis in HBCS cows. Serum concentrations of very low-density lipoprotein, apolipoprotein A-I, and apolipoprotein B were lower in HBCS cows than in LBCS or MBCS cows. Although LBCS and MBCS cows had higher reactive oxygen species levels than did HBCS cows, the malondialdehyde concentration was not different among cows with different BCS. Ceruloplasmin activity was higher in MBCS and HBCS cows than in LBCS cows, but superoxide dismutase, glutathione peroxidase, and paraoxonase activities were not different among cows with different BCS. Despite the higher percentage of granulocytes in MBCS cows than in HBCS cows, no differences were found in leukocyte counts, red blood cell profiles and platelet profiles among the cows in the three groups.ConclusionsResults of this study showed that compared with MBCS and LBCS cows, HBCS cows at late lactation stage may have accumulated more hepatic triacylglycerol and lower antioxidant potential due to greater insulin resistance.

Metabolome profiling in skeletal muscle to characterize metabolic alterations in over-conditioned cows during the periparturient period

The transition from late gestation to early lactation is associated with extensive changes in metabolic, endocrine, and immune functions in dairy cows. Skeletal muscle plays an important role in maintaining the homeorhetic adaptation to the metabolic needs of lactation. The objective of this study was to characterize the skeletal muscle metabolome in the context of the metabolic changes that occur during the transition period in dairy cows with high (HBCS) versus normal body condition (NBCS). Fifteen weeks antepartum, 38 pregnant multiparous Holstein cows were assigned to 1 of 2 groups, which were fed differently to reach the targeted BCS and back fat thickness (BFT) until dry-off at -49 d before calving (HBCS: >3.75 and >1.4 cm; NBCS: <3.5 and <1.2 cm). During the dry period and the subsequent lactation, both groups were fed identical diets. The differences in both BCS and BFT were maintained throughout the study. The metabolome was characterized in skeletal muscle samples (semitendinosus muscle) collected on d -49, 3, 21, and 84 relative to calving using a targeted metabolomics approach (AbsoluteIDQ p180 kit; Biocrates Life Sciences AG, Innsbruck, Austria), which allowed for the quantification of up to 188 metabolites from 6 different compound classes (acylcarnitines, amino acids, biogenic amines, glycerophospholipids, sphingolipids, and hexoses). On d -49, the concentrations of citrulline and hydroxytetradecadienyl-l-carnitine in muscle were higher in HBCS cows than in NBCS cows, but those of carnosine were lower. Over-conditioning did not affect the muscle concentrations of any of the metabolites on d 3. On d 21, the concentrations of phenylethylamine and linoleylcarnitine in muscle were lower in HBCS cows than in NBCS cows, and the opposite was true for lysophosphatidylcholine acyl C20:4. On d 84, the significantly changed metabolites were mainly long-chain (>C32) acyl-alkyl phosphatidylcholine and di-acyl phosphatidylcholine, along with 3 long-chain (>C16) sphingomyelin that were all lower in HBCS cows than in NBCS cows. These data contribute to a better understanding of the metabolic adaptation in skeletal muscle of dairy cows during the transition period, although the physiological significance and underlying molecular mechanisms responsible for the regulation of citrulline, hydroxytetradecadienyl-l-carnitine, carnosine, and phenylethylamine associated with over-conditioning are still elusive and warrant further investigation. The changes observed in muscle lysophosphatidylcholine and phosphatidylcholine concentrations may point to an alteration in phosphatidylcholine metabolism, probably resulting in an increase in membrane stiffness, which may lead to abnormalities in insulin signaling in the muscle of over-conditioned cows.

Branched-chain amino acids: Abundance of their transporters and metabolizing enzymes in adipose tissue, skeletal muscle, and liver of dairy cows at high or normal body condition

Branched-chain amino acids (BCAA) are major components of milk protein and important precursors for nonessential AA. Thus, the BCAA transport and break-down play a key role in the metabolic adaptation to the high nutrient demands in lactation. However, in monogastrics, increased BCAA levels have been linked with obesity and certain metabolic disorders such as impaired insulin sensitivity. Our objective was to study the effect of over-conditioning at calving on plasma BCAA levels as well as the tissue abundance of the most relevant BCAA transporters and degrading enzymes in dairy cows during late pregnancy and early lactation. Thirty-eight Holstein cows were allocated 15 wk antepartum to either a normal- (NBCS) or over-conditioned (HBCS) group, receiving 6.8 or 7.2 MJ of NEL/kg of DM, respectively, during late lactation to reach the targeted differences in body condition score (BCS) and back fat thickness (BFT; NBCS: BCS <3.5, BFT <1.2 cm; HBCS: BCS >3.75, BFT >1.4 cm) until dry-off. During the dry period and next lactation, cows were fed the same diets, whereby differences in BCS and BFT were maintained: prepartum means were 3.16 ± 0.06 and 1.03 ± 0.07 cm (NBCS) vs. 3.77 ± 0.08 and 1.89 ± 0.11 cm (HBCS), postpartum means were 2.89 ± 0.06 and 0.81 ± 0.05 cm (NBCS) vs. 3.30 ± 0.06 and 1.38 ± 0.08 cm (HBCS). Blood and biopsies from liver, semitendinosus muscle, and subcutaneous adipose tissue (scAT) were sampled at d 49 antepartum, 3, 21, and 84 postpartum. Free BCAA were analyzed and the mRNA abundance of solute carrier family 1 member 5 (SLC1A5), SLC7A5, and SLC38A2 as well as branched-chain aminotransferase 2 (BCAT2), branched-chain α-keto acid dehydrogenase E1α (BCKDHA), and branched-chain α-keto acid dehydrogenase E1β (BCKDHB) as well as the protein abundance of BCKDHA were assessed. Concentrations of all BCAA changed with time, most markedly in HBCS cows, with a nadir around calving. Apart from Ile, neither individual nor total BCAA differed between groups. The HBCS group had greater BCKDHA mRNA as well as higher prepartum BCKDHA protein abundance in scAT than NBCS cows, pointing to a greater oxidative capacity for the irreversible degradation of BCAA transamination products in scAT of over-conditioned cows. Prepartum hepatic BCKDHA protein abundance was lower in HBCS than in NBCS cows. In both groups, SLC1A5, SLC7A5, and BCAT2 mRNA were most abundant in scAT, whereas SLC38A2 was higher in scAT and muscle compared with liver, and BCKDHA and BCKDHB mRNA were greatest in liver and muscle, respectively. Our results indicate that scAT may be a major site of BCAA uptake and initial catabolism, with the former, however, being independent of BCS and time relative to calving in dairy cows.

Mammalian target of rapamycin signaling and ubiquitin-proteasome–related gene expression in skeletal muscle of dairy cows with high or normal body condition score around calving

The objective of the current study was to investigate the effects of overconditioning around calving on gene expression of key components of the mammalian target of rapamycin (mTOR) pathway and ubiquitin-proteasome system (UPS) in skeletal muscle as well as the AA profiles in both serum and muscle of periparturient cows. Fifteen weeks antepartum, 38 multiparous Holstein cows were allocated to either a high body condition group (HBCS; n = 19) or a normal body condition group (NBCS; n = 19) and were fed different diets until dry-off (d -49 relative to calving) to amplify the difference. The groups were also stratified for comparable milk yields (NBCS: 10,361 ± 302 kg; HBCS: 10,315 ± 437 kg). At dry-off, the NBCS cows (parity: 2.42 ± 1.84; body weight: 665 ± 64 kg) had a body condition score (BCS) <3.5 and backfat thickness (BFT) <1.2 cm, whereas the HBCS cows (parity: 3.37 ± 1.67; body weight: 720 ± 57 kg) had a BCS >3.75 and BFT >1.4 cm. During the dry period and the subsequent lactation, both groups were fed identical diets but maintained the BCS and BFT differences. Blood samples and skeletal muscle biopsies (semitendinosus) were repeatedly (d -49, +3, +21, and +84 relative to calving) collected for assessing the concentrations of free AA and the mRNA abundance of various components of mTOR and UPS. The differences in BCS and BFT were maintained throughout the study. The circulating concentrations of most AA with the exception of Gly, Gln, Met, and Phe increased in early lactation in both groups. The serum concentrations of Ala (d +21 and +84) and Orn (d +84) were lower in HBCS cows than in NBCS cows, but those of Gly, His, Leu, Val, Lys, Met, and Orn on d -49 and Ile on d +21 were greater in HBCS cows than in NBCS cows. The serum concentrations of 3-methylhistidine, creatinine, and 3-methylhistidine:creatinine ratio increased after calving (d +3) but did not differ between the groups. The muscle concentrations of all AA (except for Cys) remained unchanged over time and did not differ between groups. The muscle concentrations of Cys were greater on d -49 but tended to be lower on d +21 in HBCS cows than in NBCS cows. On d +21, mTOR and eukaryotic translation initiation factor 4E binding protein 1 mRNA abundance was greater in HBCS cows than in NBCS cows, whereas ribosomal protein S6 kinase 1 was not different between the groups. The mRNA abundance of ubiquitin-activating enzyme 1 (d +21), ubiquitin-conjugating enzyme 1 (d +21), atrogin-1 (d +21), and ring finger protein-1 (d +3) enzymes was greater in HBCS cows than in NBCS cows, whereas ubiquitin-conjugating enzyme 2 was not different between the groups. The increased mRNA abundance of key components of mTOR signaling and of muscle-specific ligases of HBCS cows may indicate a simultaneous activation of anabolic and catabolic processes and thus increased muscle protein turnover, likely as a part of the adaptive response to prevent excessive loss of skeletal muscle mass during early lactation.

Metabolomics meets machine learning: Longitudinal metabolite profiling in serum of normal versus overconditioned cows and pathway analysis

This study aimed to investigate the differences in the metabolic profiles in serum of dairy cows that were normal or overconditioned when dried off for elucidating the pathophysiological reasons for the increased health disturbances commonly associated with overconditioning. Fifteen weeks antepartum, 38 multiparous Holstein cows were allocated to either a high body condition (HBCS; n = 19) group or a normal body condition (NBCS; n = 19) group and were fed different diets until dry-off to amplify the difference. The groups were also stratified for comparable milk yields (NBCS: 10,361 ± 302 kg; HBCS: 10,315 ± 437 kg; mean ± standard deviation). At dry-off, the cows in the NBCS group (parity: 2.42 ± 1.84; body weight: 665 ± 64 kg) had a body condition score (BCS) <3.5 and backfat thickness (BFT) <1.2 cm, whereas the HBCS cows (parity: 3.37 ± 1.67; body weight: 720 ± 57 kg) had BCS >3.75 and BFT >1.4 cm. During the dry period and the subsequent lactation, both groups were fed identical diets but maintained the BCS and BFT differences. A targeted metabolomics (AbsoluteIDQ p180 kit, Biocrates Life Sciences AG, Innsbruck, Austria) approach was performed in serum samples collected on d -49, +3, +21, and +84 relative to calving for identifying and quantifying up to 188 metabolites from 6 different compound classes (acylcarnitines, AA, biogenic amines, glycerophospholipids, sphingolipids, and hexoses). The concentrations of 170 metabolites were above the limit of detection and could thus be used in this study. We used various machine learning (ML) algorithms (e.g., sequential minimal optimization, random forest, alternating decision tree, and naïve Bayes-updatable) to analyze the metabolome data sets. The performance of each algorithm was evaluated by a leave-one-out cross-validation method. The accuracy of classification by the ML algorithms was lowest on d 3 compared with the other time points. Various ML methods (partial least squares discriminant analysis, random forest, information gain ranking) were then performed to identify those metabolites that were contributing most significantly to discriminating the groups. On d 21 after parturition, 12 metabolites (acetylcarnitine, hexadecanoyl-carnitine, hydroxyhexadecenoyl-carnitine, octadecanoyl-carnitine, octadecenoyl-carnitine, hydroxybutyryl-carnitine, glycine, leucine, phosphatidylcholine-diacyl-C40:3, trans-4-hydroxyproline, carnosine, and creatinine) were identified in this way. Pathway enrichment analysis showed that branched-chain AA degradation (before calving) and mitochondrial β-oxidation of long-chain fatty acids along with fatty acid metabolism, purine metabolism, and alanine metabolism (after calving) were significantly enriched in HBCS compared with NBCS cows. Our results deepen the insights into the phenotype related to overconditioning from the preceding lactation and the pathophysiological sequelae such as increased lipolysis and ketogenesis and decreased feed intake.

Comparison of performance and metabolism from late pregnancy to early lactation in dairy cows with elevated v. normal body condition at dry-off

Excessive mobilization of body reserves during the transition from pregnancy to lactation imposes a risk for metabolic diseases on dairy cows. We aimed to establish an experimental model for high v. normal mobilization and herein characterized performance, metabolic and endocrine changes from 7 weeks antepartum (a.p.) to 12 weeks postpartum (p.p.). Fifteen weeks a.p., 38 pregnant multiparous Holstein cows were allocated to two groups that were fed differently to reach either high or normal body condition scores (HBCS: 7.2 NEL MJ/kg dry matter (DM); NBCS: 6.8 NEL MJ/kg DM) at dry-off. Allocation was also based on differences in body condition score (BCS) in the previous and the ongoing lactation that was further promoted by feeding to reach the targeted BCS and back fat thickness (BFT) at dry-off (HBCS: >3.75 and >1.4 cm; NBCS: <3.5 and <1.2 cm). Thereafter, both groups were fed identical diets. Blood samples were drawn weekly from 7 weeks a.p. to 12 weeks p.p. to assess the serum concentrations of metabolites and hormones. The HBCS cows had greater BCS, BFT and BW than the NBCS cows throughout the study and lost more than twice as much BFT during the first 7 weeks p.p. compared with NCBS. Milk yield and composition were not different between groups, except that lactose concentrations were greater in NBSC than in HBCS. Feed intake was also greater in NBCS, and NBCS also reached a positive energy balance earlier than HBCS. The greater reduction in body mass in HBCS was accompanied by greater concentrations of non-esterified fatty acids, and β-hydroxybutyrate in serum after calving than in NBCS, indicating increased lipomobilization and ketogenesis. The mean concentrations of insulin across all time-points were greater in HBCS than in NBCS. In both groups, insulin and IGF-1 concentrations were lower p.p than in a.p. Greater free thyroxine (fT4) concentrations and a lower free 3-3′-5-triiodothyronine (fT3)/fT4 ratio were observed in HBCS than in NBCS a.p., whereas p.p. fT3/fT4 ratio followed a reverse pattern. The variables indicative for oxidative status had characteristic time courses; group differences were limited to greater plasma ferric reducing ability values in NBSC. The results demonstrate that the combination of pre-selection according to BCS and differential feeding before dry-off to promote the difference was successful in obtaining cows that differ in the intensity of mobilizing body reserves. The HBCS cows were metabolically challenged due to intense mobilization of body fat, associated with reduced early lactation dry matter intake and compromised antioxidative capacity.

Impacts of cow body condition score during gestation on weaning performance of the offspring

This experiment evaluated the impacts of cow body condition score (BCS) during gestation on productive parameters of the offspring. Three hundred multiparous, lactating, non-pregnant Angus × Hereford cows were assigned to a fixed-time artificial insemination (AI) protocol using semen from a single sire (d 0). Forty days after AI, cows were evaluated for pregnancy status via transrectal ultrasonography and BCS, and 100 pregnant cows (543±6kg of BW, 6.6±0.3yr of age, 4.83±0.06 of BCS, and 115±2 d postpartum) were selected for the experiment. Within these 100 cows, 20 cows had BCS ≥5.50 but ≤6.50 and were classified as adequate BCS (5.85±0.06; HBCS). The remaining cows had BCS ≤4.75 (4.52±0.03), and were divided into 4 groups (20 cows/group): LBCS (4.60±0.07), BCSG1 (4.43±0.07), BCSG2 (4.63±0.07), and BCSG3 (4.63±0.07). The HBCS and LBCS cows were managed to maintain their initial BCS throughout gestation. The BCSG1, BCSG2, and BCSG3 cows were managed to gain 1.50 BCS during the first, second, and third trimester of gestation, respectively, and maintain the resultant BCS until calving. Cow BCS was assessed again on d 102, 182, and 265. During the calving season (d 272-291), calf body weight (BW) was recorded within 3h after birth. Only cows that met the BCS maintenance (within 0.50 of BCS change) and change (≥1.25 and ≤1.75 of BCS increase within the trimester) criteria were maintained in the experiment (HBCS, n =14; LBCS, n =14; BCSG1, n =14; BCSG2, n =15, BCSG3, n =15). On d 344, cow milk production was estimated by the weigh-suckle-weigh method, and calves were weaned on d 475. No differences were detected (P≥0.42) for calving rate, calf birth BW, and cow milk production. Weaning rate and calf age at weaning were also similar among BCS groups (P≥0.15). However, calf weaning BW was greater (P≤0.05) for BCSG2 and BCSG3 cows (265 and 262kg, respectively; SEM =4) compared with HBCS and LBCS cows (248 and 249kg, respectively; SEM =4), and similar (P≥0.20) among all other comparisons. These results suggest that offspring weaning BW is directly influenced by BCS gain of beef cows during the second and third trimesters of gestation.

Late gestation supplementation of beef cows differing in body condition score: Effects on cow and calf performance1,2

A 2-yr study utilizing 120 mature, crossbred (Angus × Herford) cows/year, evaluated the influence of cow BCS and dried distillers grains with solubles (DDGS) supplementation during late gestation on cow performance and productivity of subsequent offspring. Treatments were arranged as a 2×2 factorial in a randomized complete block design with 2 BCS and with or without DDGS supplementation. Cows were nutritionally managed to enter the last trimester of gestation with a BCS of approximately 4 (LBCS) or 6 (HBCS) and were thereafter managed in a single herd (initial BCS were 4.4 and 5.7 for LBCS and HBCS treatments, respectively). During the last trimester, 12.7 kg/cow of low quality meadow hay (6.4% CP; DM basis) was provided each day. Supplemented cows were gathered and sorted into pens (12 pens; 5 cows/pen; 6 pens/BCS) every Monday, Wednesday, and Friday, and received the equivalent of 0.9 kg/cow daily of DDGS (31% CP; DM basis; supplement was consumed within 30 min on each supplementation day). Calf birth weight was greater for HBCS compared to LBCS (P=0.001) and for supplemented compared to nonsupplemented cows (P=0.04). Cow weight at weaning was greater for HBCS compared with LBCS (P<0.001); however, no differences were noted because of supplementation (P=0.16). Weaning weight was greater for the offspring of supplemented compared to nonsupplemented cows (P=0.02). There were no differences in postweaning calf performance (growing lot and feedlot) or carcass characteristics (P>0.05) due to treatments. Nevertheless, HBCS cows had approximately 10% more live calves at birth and at weaning (P≤0.01) compared to LBCS cows. Consequently, the total weaned calf weight per cow was 26 kg greater for HBCS compared with LBCS (P=0.004). Pregnancy rate was greater (P=0.05) for HBCS than LBCS cows (92% vs. 79%, respectively) but not affected by supplementation (P=0.94). This research demonstrates the potential consequences of not maintaining cows in adequate BCS at calving. Also, though it appears that supplementation of beef cows with DDGS during late gestation has a positive effect on weaning weight, there was no apparent developmental programming effect on feedlot performance and carcass characteristics of calves.

Short communication: Effects of serum obtained from dairy cows with low or high body condition score on in vitro embryo development

The objective of the study was to determine whether the serum obtained from animals differing in body condition score (BCS) affects in vitro embryo development. After in vitro fertilization, serum obtained from dairy cows of either low (L-BCS; 2.1±0.14 on a scale of 1 to 5) or high BCS (H-BCS; 4.0±0.0), or commercially available bovine serum (control) was added at 5% concentration to the in vitro culture medium. Use of serum obtained from H-BCS cows increased the cleavage rates compared with control serum at both 24 and 48h after in vitro fertilization (78.3 vs. 71.9% and 79.9 vs. 75.1%, respectively), whereas use of serum obtained from L-BCS cows increased the blastocyst rate compared with control serum at 7d (23.8 vs. 19.1%), but this difference was not evident at 8 or 9d after in vitro fertilization. As nonesterified fatty acid concentrations were highest in control serum, followed by serum from L-BCS and H-BCS cows (621, 559, and 272μEq/L, respectively), a high concentration of nonesterified fatty acids might adversely affect the very early stages of embryo development, and its negative effects might be greater immediately after fertilization compared with developmental stages after morula formation. Our findings also indicate that factors promoting early stage embryo development do not necessarily promote blastocyst development. Serum obtained from animals under different physiological conditions may be used for in vitro embryo culture to study the effects of nutritional management of dairy cattle on embryo development.

Levels of insulin, insulin-like growth factor-I and thyroid hormones in relation to the body condition score changes in periparturient dairy cows

The objective of this study was to determine the levels of insulin, insulin like growth factor I (IGF-I) and thyroid hormones in relation to the body condition score (BCS) of periparturient dairy cows. The study was carried out on twenty Holstein-Friesian dairy cows with average milk production of 7000 L/305 days in the previous lactation, parity ranging from 2-4. All cows were BCS scored during the early dry period, 7±3 days before and after parturition. Based on the BCS at the early dry period, cows were divided in two groups: cows with high BCS (3.75- 4.25, HBCS, n=10), and cows with moderate BCS (2.75-3.75, MBCS, n=10). Blood samples were taken at the time of BCS evaluation. Concentrations of insulin, IGF-I, triiodothyroinine (T3) and thyroxine (T4) were determined by radioimmunoassay (RIA, INEP-Zemun, Serbia). Statistical differences between mean values were determined using Student t-test (p<0.01). Mean BCS values at early dry period were significantly different compared to two other periods (p<0.01) within both groups, but BCS was similar in both groups at 7±3 days after calving (2.69±0.67:2.62± 0.27, p>0.05). IGF-I level in HBCS cows at days 7±3 before calving was significantly higher (16.28±3.07:11.76±2.28, p<0.01), with a reverse relationship after calving (3.77±1.64:8.46±2.37, p<0.01). Insulin level was significantly lower at 7±3 days before calving in HBCS cows (16.26±4.60:20.18±4.96mIU/L, p<0.05). Thyroid hormones levels were significantly lower in HBCS group et all examined periods. [Projekat Ministarstva nauke Republike Srbije, br. III 46002 i br. 31003]