Effects of Soybean and Linseed Oils Calcium Salts and Starter Protein Content on Growth Performance, Immune Response, and Nitrogen Utilization Efficiency in Holstein Dairy Calves.

This study aimed to investigate the interaction of fatty acid (FA) source [calcium salt of soybean oil (n-6 FA) vs. calcium salt of linseed oil (n-3 FA) both 3% DM basis] with protein content (18% vs. 22% CP, based on DM) on growth performance, blood metabolites, immune function, skeletal growth indices, urinary purine derivatives (PD), and microbial protein synthesis (MPS) in young dairy calves. Forty 3-day-old calves (20 females and 20 males) with a starting body weight (BW) of 40.2 kg were assigned in a completely randomized block design in a 2 × 2 factorial arrangement of treatments. Experimental diets were: (1) n-6 FA with 18% CP (n-6-18CP), (2) n-6 FA with 22% CP (n-6-22CP), (3) n-3 FA with 18% CP (n-3-18CP), and (4) n-3 FA with 22% CP (n-3-22CP). Starter feed intake and average daily gain (ADG) were not influenced by experimental diets (p > 0.05). However, before weaning and the entire period, feed efficiency (FE) was greater in calves fed n-3 FA compared to n-6 FA (p < 0.05). Heart girth (weaning, p < 0.05) and hip height (weaning, p < 0.05 and final, p < 0.01) were highest among experimental treatments in calves who received n-3-22CP diets. The greatest blood glucose (p < 0.05) and insulin (p < 0.01) concentrations in the pre-weaning period and the lowest serum concentration of tumor necrosis factor (before weaning, p < 0.05) were observed in calves fed the n-3-22CP diet. However, the greatest blood urea N (before weaning, p < 0.05; after weaning, p < 0.05) and urinary N excretion (p < 0.05) were found in calves fed n-6-22CP diets compared to other experimental arrangements. In conclusion, offering calves with Ca-salt of n-3 FA along with 22% CP content may be related to improved nitrogen efficiency and immune function.

Low-Protein Diets with Fixed Carbohydrate Content Promote Hyperphagia and Sympathetically Mediated Increase in Energy Expenditure.

SCOPE: Dietary protein restriction elicits hyperphagia and increases energy expenditure; however, less is known of whether these responses are a consequence of increasing carbohydrate content. The effects of protein-diluted diets with fixed carbohydrate content on energy balance, hormones, and key markers of protein sensing and thermogenesis in tissues are determined. METHODS AND RESULTS: Obesity-prone rats (n = 13-16 per group) are randomized to diets containing fixed carbohydrate (52% calories) and varying protein concentrations: 15% (control), 10% (mild protein restriction), 5% (moderate protein restriction) or 1% (severe protein restriction) protein calories, or protein-matched to 5% protein, for 21 days. Propranolol and ondansetron are administered to interrogate the roles of sympathetic and serotonergic systems, respectively, in diet-induced changes in energy expenditure. It is found that mild-to-moderate protein restriction promotes transient hyperphagia, whereas severe protein restriction induces hypophagia, with alterations in meal patterns. Protein restriction enhances energy expenditure that is partly attenuated by propranolol, but not ondansetron. Moderate to severe protein restriction decreases gains in body weight, lean and fat mass, decreased postprandial glucose and leptin, but increased fibroblast growth factor-21 concentrations. Protein-matching retains lean mass suggesting that intake of dietary protein, but not calories, is important for preserving lean mass. Notably, protein restriction increases the protein and/or transcript abundance of key amino acid sensing molecules in liver and intestine (PERK, eIF2α, ATF2, CHOP, 4EBP1, FGF21), and upregulated thermogenic markers (ß2AR, Klotho, HADH, UCP-1) in brown adipose tissue. CONCLUSION: Low-protein diets promote hyperphagia and sympathetically mediated increase in energy expenditure, prevent gains in tissue reserves, and concurrently upregulate hepatic and intestinal amino acid sensing intermediaries and thermogenic markers in brown adipose tissue.

Diets enriched in whey or casein improve energy balance and prevent morbidity and renal damage in salt-loaded and high-fat-fed spontaneously hypertensive stroke-prone rats.

High-fat diets induce obesity and increase risks of diabetes and cardiovascular and renal disorders. Whey- or casein-enriched diets decrease food intake and weight gain; however, their cardiovascular and renal benefits are unclear. We determined whether whey- and casein-enriched diets improve energy balance and are protective against renal damage and morbidity associated with stroke in an obesogenic and hypertensive experimental setting. We also assessed whether the hypophagic effects of these diets were due to reduced diet preference. In experiment 1, spontaneously hypertensive stroke-prone rats were randomized to (a) control (CON; 14% kcal protein, 33% fat), (b) whey (WHY; 40% protein, 33% fat), (c) casein (CAS; 40% protein, 33% fat) or (d) chow (CHW; 24% protein, 13% fat) for 12 weeks with 1% salt in drinking water for CON, WHY and CAS groups. Our results demonstrated that both WHY and CAS produced short-term hypophagia, moderately increased energy expenditure and decreased respiratory quotient, body weight and lean mass, with effects of WHY being more prolonged. Further, only WHY decreased fat mass and blood pressure. Importantly, both WHY and CAS prevented morbidity associated with stroke and decreased indices of renal inflammation (tumor necrosis factor-α, interleukin-6) and damage (osteopontin, renal lesions). In experiment 2, following four initial conditioning trials, the preference for CON, WHY or CAS diet was determined. Both WHY and CAS decreased food intake during conditioning and decreased preference. In conclusion, diets enriched in whey or casein improved energy balance, increased survival and prevented renal damage in salt-loaded and high-fat-fed spontaneously hypertensive stroke-prone rats.

Dietary Whey and Casein Differentially Affect Energy Balance, Gut Hormones, Glucose Metabolism, and Taste Preference in Diet-Induced Obese Rats.

BACKGROUND: Dietary whey and casein proteins decrease food intake and body weight and improve glycemic control; however, little is known about the underlying mechanisms. OBJECTIVE: We determined the effects of dietary whey, casein, and a combination of the 2 on energy balance, hormones, glucose metabolism, and taste preference in rats. METHODS: In Expt. 1, Obesity Prone CD (OP-CD) rats were fed a high-fat control diet (33% fat energy) for 8 wk, and then randomly assigned to 4 isocaloric dietary treatments (n = 12/group): the control treatment (CO; 14% protein energy from egg white), the whey treatment (WH; 26% whey + 14% egg white), the casein treatment (CA; 26% casein + 14% egg white), or the whey plus casein treatment (WHCA; 13% whey + 13% casein + 14% egg white) for 28 d. Measurements included food intake, energy expenditure, body composition, metabolic hormones, glucose tolerance and key tissue markers of glucose and energy metabolism. In Expt. 2, naïve OP-CD rats were randomly assigned to 3 groups (n = 8/group). During an 8 d conditioning period, each group received on alternate days either the CO or WH, CO or CA, or CO or WHCA. Subsequently, preferences for the test diets were assessed on 2 consecutive days with food intake measurements at regular intervals. RESULTS: In Expt. 1, food intake was decreased by 17-37% for the first 14 d in the WH and CA rats, and by 18-34% only for the first 4 d in the WHCA compared with the CO rats. Fat mass decreased by 21-28% for the WH rats and 17-33% for the CA rats from day 14 onward, but by 30% only on day 28 in WHCA rats, relative to CO rats. Thus, food intake, body weight, and fat mass decreased more rapidly in WH and CA rats than in WHCA rats. Energy expenditure in WH rats decreased for the first 4 d compared with CA and WHCA rats, and for the first 7 d compared with the CO rats. Circulating leptin, glucose-dependent insulinotropic polypeptide, interleukin 6, and glucose concentrations were lower in WH, CA, and WHCA rats than in CO rats. Plasma glucagon-like peptide 1 concentrations were greater in WH than in CA or WHCA rats. The improvements in glucose tolerance were greater in WH than in WHCA rats. The plasma membrane glucose transporter 4 (GLUT4)-to-total GLUT4 ratio in skeletal muscle was greater in CA and WHCA rats than in CO rats; other markers of glucose and energy metabolism in the adipose and cardiac tissues did not differ. In Expt. 2, during 4 conditioning trials, daily food intake was decreased in WH, CA, and WHCA rats by 26-37%, 30-43%, and 23-33%, respectively, compared with CO rats. Preferences for WH and CA rats were 45% and 31% lower, respectively, than those for CO rats, but that for WHCA rats did not differ. CONCLUSION: Together, these data demonstrate that in obese rats, whey, casein, and their combination improve energy balance through differential effects on food intake, taste preference, energy expenditure, glucose tolerance, and gut hormone secretion.

Effects of Roux-en-Y gastric bypass and ileal transposition surgeries on glucose and lipid metabolism in skeletal muscle and liver.

BACKGROUND: Roux-en Y gastric bypass (RYGB) and ileal transposition (IT) surgeries produce weight loss and improve diabetic control; however, the mechanisms of glycemic improvements are largely unknown. Because skeletal muscle and liver play a key role in glucose homeostasis, we compared the effects of RYGB and IT surgeries on key molecules of glucose and lipid metabolism in muscle and liver. METHODS: Sprague-Dawley rats were subjected to RYGB, IT, or sham surgeries; sham-animals were ad-lib fed or pair-fed to RYGB rats (n = 7-9/group). At 8 weeks postoperatively, blood samples were collected for glucagon-like peptide-1 (GLP-1) and insulin analyses by ELISA. Leg muscle and liver tissues were analyzed for mRNA (RT-qPCR) and/or protein abundance (immuno blotting) of important molecules of glucose and lipid metabolism [glucose transporter-4 (GLUT-4), hexokinase, phosphofructokinase (PFK), adenosine monophosphate activated protein kinase-α (AMPKα), cytochrome C oxidase-IV (COX-IV), citrate synthase, carnitine palmitoyl transferase-1 (CPT-1), medium-chain acyl-CoA dehydrogenase (MCAD), peroxisome proliferator-activated receptor gamma co-activator 1 α (PGC-1 α), PGC-1-related coactivator (PRC), uncoupling protein-3 (UCP-3)]. RESULTS: Plasma GLP-1 concentrations were increased comparably with RYGB and IT. RYGB and IT increased muscle GLUT-4 protein content, muscle hexokinase mRNA, and liver PFK mRNA. IT increased muscle AMPKα and COX-IV protein content and liver citrate synthase activity. IT increased muscle CPT-1, MCAD and PRC mRNA, whereas RYGB increased UCP-3 mRNA in muscle and liver, and PGC-1 α mRNA in liver. CONCLUSION: The data suggest that RYGB and IT surgeries lead to enhanced GLP-1 secretion and produce similar stimulatory effects on important molecules of glucose metabolism but differential effects on key molecules of lipid oxidation in muscle and liver.

Variation of inflammatory dynamics and mediators in primiparous cows after intramammary challenge with Escherichia coli.

The objective of the current study was to investigate (i) the outcome of experimentally induced Escherichia coli mastitis in primiparous cows during early lactation in relation with production of eicosanoids and inflammatory indicators, and (ii) the validity of thermography to evaluate temperature changes on udder skin surface after experimentally induced E. coli mastitis. Nine primiparous Holstein Friesian cows were inoculated 24 ± 6 days (d) after parturition in both left quarters with E. coli P4 serotype O32:H37. Blood and milk samples were collected before and after challenge with E. coli. The infrared images were taken from the caudal view of the udder following challenge with E. coli. No relationship was detected between severity of mastitis and changes of thromboxane B2 (TXB2), leukotriene B4 (LTB4) and lipoxin A4 (LXA4). However, prostaglandin E2 (PGE2) was related to systemic disease severity during E. coli mastitis. Moreover, reduced somatic cell count (SCC), fewer circulating basophils, increased concentration of tumor necrosis factor-α (TNF-α) and higher milk sodium and lower milk potassium concentrations were related to systemic disease severity. The thermal camera was capable of detecting 2-3 °C temperature changes on udder skin surface of cows inoculated with E. coli. Peak of udder skin temperature occurred after peak of rectal temperature and appearance of local signs of induced E. coli mastitis. Although infrared thermography was a successful method for detecting the changes in udder skin surface temperature following intramammary challenge with E. coli, it did not show to be a promising tool for early detection of mastitis.

Hydrophilic interaction LC of peptides: columns comparison and clustering.

A wide variety of hydrophilic interaction chromatography (HILIC) stationary phase surface chemistries are currently available. Although their selectivity can be considerably different, column comparison or clustering using peptides is limited. In this study, ten pharmaceutically relevant model peptides are analyzed on seven different HILIC columns (bare silica, amide, poly-hydroxyethyl aspartamide, diol and zwitterionic) for the evaluation of their performance and classification. The responses examined include single and multiple responses: plate number, asymmetry factor, LOD, geometric mean resolution, resolution product, time corrected resolution product, peak capacity and chromatographic response function. Column classification was performed using hierarchical clustering and principal component analysis. Moreover, the overall performance quality of the HILIC columns was compared using a linear desirability function. Hierarchical clustering and principal component analysis showed consistent clusters. The zwitterionic phase was clustered apart from the other HILIC columns and both poly-aspartamide columns were clustered together. In addition, the two bare silica phases represent two different clusters, and thus different selectivities. Overall, the responses showed the best performance for one of the bare silica columns (Alltima-Alltech), followed by the zwitterionic phase (ZIC)-HILIC. Thus, these columns, belonging to different clusters, were found to be the best performing systems in pharmaceutical peptide analysis for the selected peptide set.

Adsorption of peptides at the sample drying step: influence of solvent evaporation technique, vial material and solution additive.

Although the efficient and careful removal of solvent from samples by centrifugal evaporation or freeze-drying methods is an important step in peptidomics, the recovery of peptides has not yet been fully investigated with these sample drying methods. Moreover, the surface adsorption of the peptides by the container and efforts to reduce this adsorption by organic additives is only scarcely elaborated until now. In this experiment, the recovery of five model peptides, i.e. bovine insulin, mouse obestatin, goserelin, buserelin and leucine-enkephalin was investigated applying dimethylsulfoxide (DMSO), dimethylformamide (DMF), polyethylene glycol 400 (PEG 400), mannitol and n-nonyl-beta-d-glucopyranoside (C(9)-Glu) in function of the two applied solvent evaporation processes (freeze-drying vs. centrifugal evaporation) and vial types, i.e. polypropylene (PP) and glass. Under our experimental conditions, drying resulted in a decreased recovery of the model peptides by 10% on average. Insulin showed the lowest recovery value relative to the other model peptides. For both drying methods, recovery of the model peptides was increased when C(9)-Glu was present. Overall, the use of PP vials is proposed for freeze-drying, while glass vials are found to be more suitable for centrifugal evaporation. The presence of PEG 400 in PP vials caused significantly reduced recoveries for all model peptides using centrifugal evaporation, although this was not observed in glass vials. As a general conclusion, applying C(9)-Glu as an additive along with choosing appropriate vial type (i.e. PP for lyophilization and glass for centrifugal evaporation) can avoid or diminish peptide loss during the evaporation procedure.

Impurity profiling quality control testing of synthetic peptides using liquid chromatography-photodiode array-fluorescence and liquid chromatography-electrospray ionization-mass spectrometry: the obestatin case.

Following several conflicting publications, the inability to reproduce the original findings on in vitro obestatin binding and activation of GPR39 receptors was recently reported by its discoverers, and several hypotheses to rationalize these findings were proposed. Based on one of these postulations (i.e., presence of impurities), peptide identity and impurity profiles were thoroughly evaluated on obestatin peptides obtained from five different manufacturers, as used by the different research groups. We found that one of the products examined was in reality a totally different peptide and that the quality of two-thirds of the other peptides was insufficient for in vitro and in vivo experiments (i.e., peptide purity less than 95% and/or individual impurities exceeding 1%). These observations question the divergent conclusions reported in the literature about the activity of obestatin. Therefore, we strongly recommend appropriate quality control testing before using any peptides for biomedical research purposes.