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To exert their therapeutic effects, nanoparticles (NPs) often need to travel into the tissues composed of multilayered cells. Accumulative evidence has revealed the crucial role of transcellular transport route (entry into one cell, exocytosis, and re-entry into another) in this process. While NP endocytosis and subcellular transport are intensively characterized, the exocytosis and re-entry steps are poorly understood, which becomes a barrier for NP delivery into complex tissues. Here, the authors term the exocytosis and re-entry steps together as intercellular exchange. A collagen-based three-dimension assay is developed to specifically quantify the intercellular exchange of NPs, and distinguish the contributions of several potential mechanisms. The authors show that NPs can be exocytosed freely or enclosed inside extracellular vesicles (EVs) for re-entry, while direct cell-cell contact is hardly involved. EVs account for a significant fraction of NP intercellular exchange, and its importance in NP transport is demonstrated in vitro and in vivo. While freely released NPs engage with the same receptors for re-entry, EV-enclosed ones bypass this dependence. These studies provide an easy and precise system to investigate the intercellular exchange stage of NP delivery, and shed the first light in the importance of EVs in NP transport between cells and into complex tissues., (© 2022 The Authors. Advanced Science published by Wiley-VCH GmbH.)

Idiopathic pulmonary fibrosis (IPF) is a lethal lung disease targeting the alveolar gas exchange apparatus, leading to death by asphyxiation. IPF progresses on a tissue scale through aberrant matrix remodeling, enhanced cell contraction, and subsequent microenvironment densification. Although two pharmaceuticals modestly slow progression, IPF patient survival averages less than 5 years. A major impediment to therapeutic development is the lack of high-fidelity models that account for the fibrotic microenvironment. Our goal is to create a three-dimensional (3D) platform to enable lung fibrosis studies and recapitulate IPF tissue features. We demonstrate that normal lung fibroblasts encapsulated in collagen microspheres can be pushed toward an activated phenotype, treated with FDA-approved therapies, and their fibrotic function quantified using imaging assays (extracellular matrix deposition, contractile protein expression, and microenvironment compaction). Highlighting the system's utility, we further show that fibroblasts isolated from IPF patient lungs maintain fibrotic phenotypes and manifest reduced fibrotic function when treated with epigenetic modifiers. Our system enables enhanced screening due to improved predictability and fidelity compared to 2D systems combined with superior tractability and throughput compared to 3D systems., (© 2021 Author(s).)

Yes-associated protein (YAP) and PDZ-binding motif (TAZ) have emerged as important regulators of pathologic fibroblast activation in fibrotic diseases. Agonism of Gαs-coupled G protein coupled receptors (GPCRs) provides an attractive approach to inhibit the nuclear localization and function of YAP and TAZ in fibroblasts that inhibits or reverses their pathological activation. Agonism of the dopamine D1 GPCR has proven effective in preclinical models of lung and liver fibrosis. However, the molecular mechanisms coupling GPCR agonism to YAP and TAZ inactivation in fibroblasts remain incompletely understood. Here, using human lung fibroblasts, we identify critical roles for the cAMP effectors EPAC1/2, the small GTPase RAP2c, and the serine/threonine kinase MAP4K7 as the essential elements in the downstream signaling cascade linking GPCR agonism to LATS1/2-mediated YAP and TAZ phosphorylation and nuclear exclusion in fibroblasts. We further show that this EPAC/RAP2c/MAP4K7 signaling cascade is essential to the effects of dopamine D1 receptor agonism on reducing fibroblast proliferation, contraction, and extracellular matrix production. Targeted modulation of this cascade in fibroblasts may prove a useful strategy to regulate YAP and TAZ signaling and fibroblast activities central to tissue repair and fibrosis., (© 2021 Wiley Periodicals LLC.)

This quality improvement project used the Model for Improvement including the Plan-Do-Study-Act cycle of change framework to educate pediatric intensive care unit (PICU) nurses on risk factors for pediatric pressure injuries and prevention strategies, improve turning compliance for PICU patients, and implement an electronic trigger to order nutrition consultations on all patients with a Braden Q score less than 16. The quality improvement project decreased preventable patient harm to PICU patients by decreasing the pressure injury incidence rate from 8% to 3% in the 6-week time period., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Impact Statement: The described microtissue-microwell workflow is uniquely suited for high-throughput study of extracellular matrix (ECM) remodeling at the molecular, cellular, and tissue levels and demonstrates possibilities of studying progressive, heterogeneous diseases in a way that is meaningful for drug discovery and development. We outline several assays that can be utilized in studying tissue-level diseases and functions that involve cell-ECM interactions and ECM remodeling (e.g., cancer, fibrosis, wound healing) in pursuit of an improved three-dimensional cell culturing system. Finally, we demonstrate the ability to cryopreserve cells encapsulated in microtissue constructs while remaining highly viable, proliferative, and retaining cell functions that are involved in ECM remodeling.

A critical role of vascular endothelium is as a semi-permeable barrier, dynamically regulating the flux of solutes between blood and the surrounding tissue. Existing platforms that quantify endothelial function in vitro are either significantly throughput limited or overlook physiologically relevant extracellular matrix (ECM) interactions and thus do not recapitulate in vivo function. Leveraging droplet microfluidics, we developed a scalable platform to measure endothelial function in nanoliter-volume, ECM-based microtissues. In this study, we describe our high-throughput method for fabricating endothelial-coated collagen microtissues that incorporate physiologically relevant cell-ECM interactions. We showed that the endothelial cells had characteristic morphology, expressed tight junction proteins, and remodeled the ECM via compaction and deposition of basement membrane. We also measured macromolecular permeability using two optical modalities, and found the cell layers: (1) had permeability values comparable to in vivo measurements and (2) were responsive to physiologically-relevant modulators of endothelial permeability (TNF-α and TGF-β). This is the first demonstration, to the authors' knowledge, of high-throughput assessment (n > 150) of endothelial permeability on natural ECM. Additionally, this technology is compatible with standard cell culture equipment (e.g. multi-well plates) and could be scaled up further to be integrated with automated liquid handling systems and automated imaging platforms. Overall, this platform recapitulates the functions of traditional transwell inserts, but extends application to high-throughput studies and introduces new possibilities for interrogating cell-cell and cell-matrix interactions.

Background: Appendiceal cancer (AC) patients treated with cytoreductive surgery (CRS) and hyperthermic intraperitoneal chemotherapy (HIPEC) often demonstrate an unpredictable variability in their survival outcomes. Biomarkers predictive of CRS/HIPEC efficacy could better guide treatment decisions. We hypothesized that variation in the transcriptional programming of AC tumors might distinguish molecular subtypes with differential outcomes after CRS/HIPEC., Study Design: Gene expression profiles of 2 AC cohorts were analyzed using Affymetrix whole-genome expression microarrays. Hierarchical clustering methods, Kaplan-Meier analysis, and Cox regression models were used to discover and validate prognostic molecular subtypes of AC. Gene set enrichment analysis was used to infer pathologic attributes of the molecular subtypes., Results: Unsupervised hierarchical clustering analysis of tumor expression profiles revealed a 139-gene cassette that distinguished 2 molecular subtypes (based on low vs high expression of the gene cassette) with statistically significant survival differences (disease-specific survival, p = 0.0075; progression-free survival, p = 0.0072). In a second AC cohort, the 139-gene cassette reproducibly partitioned tumors into subtypes with significant survival differences. Tumors showing high relative expression of the genes comprising the cassette associated with poor survival outcomes (disease-specific survival, p = 0.047; progression-free survival, p = 0.0079), and exhibited gene expression patterns enriched for oncogenic processes and pathways. The prognostic value of the molecular subtypes was specific for low-grade appendiceal tumors (disease-specific survival, p = 0.028; progression-free survival, p = 0.0016), and remained significant in the presence of conventional prognostic markers, including grade, surgical resection score, Eastern Cooperative Oncology Group status, and age., Conclusions: The 139-gene cassette can have actionable clinical utility for identifying low-grade appendiceal tumor molecular subtypes predictive of therapeutic efficacy of CRS/HIPEC., (Copyright © 2016 American College of Surgeons. Published by Elsevier Inc. All rights reserved.)

Background: Cytoreductive surgery (CRS) and hyperthermic intraperitoneal chemotherapy (HIPEC) have shown variability in survival outcomes when used to treat peritoneal surface disease (PSD) from appendiceal and colorectal cancers. The primary goal of this study was to examine outcomes for high-grade appendiceal (HGA) and high-grade colonic primaries after CRS-HIPEC to determine if a significant difference exists between the two groups., Methods: A retrospective analysis of patients with peritoneal dissemination from appendiceal and colonic primaries were identified in a prospectively maintained database of 1,223 CRS-HIPEC procedures performed between 1991 and 2015. Patient demographics, performance status resection status, tumor grade, nodal status, morbidity, mortality, and survival were reviewed with biopsy-proven PSD being classified according to primary site. Univariate and multivariate analyses were performed, and outcomes compared., Results: The study identified 171 CRS-HIPEC procedures for 165 patients: 110 (66.7%) for HGA and 55 (33.3%) for high-grade colonic lesions. Observed median disease-free survival (DFS) and overall survival (OS) for both groups were the same at14.4 and 18 months, respectively. Median survival according to resection status for R0/R1, R2a, and R2b/c were 36, 15.6, and 8.4 months (P<0.0001). Median OS for those who received preoperative chemotherapy versus those who did not were 14.4 and 20.4 months, respectively (P=0.01). For those who received preoperative chemotherapy, no difference was apparent in the DFS interval (P=0.34). Multivariate predictors of OS included resection status (P<0.0001) and lymph node involvement (P=0.0005)., Conclusions: Preoperative chemotherapy offered no clear DFS or OS benefit, for HGA or high-grade colon cancer patients. Complete cytoreduction offered the greatest survival benefit to both groups with a correlating drop in survival to resection status. Outcomes for high grade appendiceal cancer are remarkably similar to colon cancer.

The free amino acids (FAA) profile was determined for newly fertilized eggs and resultant larvae from wild-caught red snapper Lutjanus campechanus induced to spawn with hCG. Yolk sac and oil globule volumes of eggs and larvae were monitored over time from digital photographs. FAA profiles of the eggs and larvae were measured in picomoles (pmol) of FAA/mg of eggs by HPLC. Newly fertilized eggs had a mean total FAA content of 21.72 +/- 3.55 nmoles/egg (92.81 +/- 9.71 nmoles/mg eggs). Leucine, valine, lysine, and isoleucine were the most abundant essential FAA comprising 35.9% of the total FAA. Alanine, serine, asparagine, and glycine were the most abundant non-essential FAA comprising 34.2% of the total FAA. At 24 h post-hatch (hph) the mean total FAA had decreased by 81% since egg fertilization. The bulk of the FAA decrease was between the time of hatch and 12 hph. Only 8.5 +/- 1.5% of the initial concentration in fertilized eggs of isoleucine, 9.7 +/- 2.5% of arginine, and 9.9 +/- 2.0% of threonine remained at 12 hph. Among the non-essential FAA, alanine dropped the most by 12 hph with 4.6% of the concentration found in a recently fertilized egg remaining, while cysteine had increased 254.7 +/- 26.2%. The yolk sac volume decreased rapidly in the first 12 hph and was further reduced 77.0 +/- 2.5% from 12 to 24 hph. The oil globule depletion rate was a more linear decline from fertilized egg to 36 hph.

The objective of this study was to determine the possibility of replacing soybean meal (SBM) in finisher pig diets completely with hydrolyzed feather meal (FM). Forty-five gilts and 45 castrated males (57.8+/-0.8 kg of BW; 3 gilts or 3 castrated males/pen) were randomly assigned to 5 finisher-1 diets with 6 pens per diet. At an average pen BW of 81.0+/-1.4 kg, pigs were offered finisher-2 diets. Corn-SBM, finisher-1 and finisher-2 positive control (PC) diets were formulated to contain 6.1 and 4.7 g of apparent ileal digestible (AID) Lys/kg, respectively. Corn-FM negative control (NC) diets were formulated to be iso-N to the PC diets. The NC diets were supplemented with Lys and Trp (NRC; NC + Lys and Trp) to alleviate apparent deficiencies based on AID AA in FM reported by the 1998 NRC. In addition, the NC diets were supplemented further with the third limiting AA, Thr (40+3AA; NC + Lys, Trp, and Thr), and fourth and fifth limiting AA, His and Ile (40+5AA; NC + Lys, Trp, Thr, His, and Ile), based on the assumption that the apparent ileal digestibility of all indispensable AA in FM is 40%. Pigs had ad libitum access to feed and water. At the end of the study (112.1+/-1.8 kg), blood samples were collected from all pigs before slaughter. As expected, overall ADFI, AID Lys (representing indispensable AA) intake (LysI), ADG, and G:F were greater and BW gain:LysI (G:LysI) was less in pigs fed the PC diet than those fed the NC diet (P<0.001). Pigs fed the NRC diet grew slower than those fed the PC diet (P=0.004), and their G:LysI tended to be less than those fed the PC (P=0.083) or the 40+3AA and 40+5AA diets (P=0.094). Greater fat-free lean percentage (P=0.005), fat-free lean gain (LG; P<0.001), and LG:F (P<0.001) were observed in pigs fed the PC diet compared with those fed the NC diet, but their LG:F or LG:LysI was similar to those fed the NRC diet. However, LG:F (P=0.030) and LG:LysI (P=0.028) were greater in pigs fed the 40+3AA and the 40+5AA diets than those fed the NRC diet. Pigs fed the 40+5AA diet tended to utilize AID Lys more efficiently for LG than those fed the 40+3AA diet (P=0.068). Subjective meat color (P<0.001), firmness (P=0.004), and marbling (P<0.001) scores were greater in pigs fed the 40+5AA diet than those fed the 40+3AA diet. Diets seemed to have no clear effect on the weight of liver, kidney, and heart or serum urea N and triacylglycerides. Pigs fed the PC diet had more serum albumin and total protein (P<0.001), but less glucose (P=0.031) and cholesterol (P<0.001) than those fed the NC diet. Serum total protein was greater (P=0.031) in pigs fed the 40+5AA diet than those fed the 40+3AA diet. The results indicated that the corn-FM diets were not as effective as the corn-SBM diet in supporting BW gain of finisher pigs. However, the results seemed to indicate that pigs fed the corn-FM diet supplemented with the necessary AA can utilize feed and AA for BW gain and lean gain as efficiently as those fed the corn-SBM diet. Further research is needed to explore fully the optimal AA supplementation strategy to effectively utilize FM for pig production.

Atmospheric ammonia (NH(3)) is a concern because of its environmental impact. The greatest contribution to atmospheric NH(3) comes from agricultural sources. This study quantified NH(3) volatilization from cattle defecation and urination on pasture under field conditions in Auburn, Alabama. Treatments consisted of beef feces, dairy feces, dairy urine, and a control. The experiment was conducted during four seasons from June 2003 to April 2004. Fresh feces or urine was applied onto grass swards, and NH(3) volatilization was measured up to 14 d after application using an inverted chamber method. Dairy urine was the only significant source of NH(3). Ammonia nitrogen (N) loss differed among seasons, ranging from 1.8% in winter to 20.9% during the warmer summer months. Cumulative volatilization was best described in this experiment by the equation % NH(3)-N loss = N(max) (1 - e(-ct))(i). The highest rate of NH(3) volatilization generally occurred within 24 h. This study suggests that NH(3) volatilization from cattle urine on pasture is significant and varies with season, whereas NH(3) volatilization from cattle feces is negligible.

A feeding trial was designed to examine the effects of copper sulfate pentahydrate (CuSO(4).5H(2)O) on the fatty acid composition and oxidative stability in muscle and adipose tissues of Boer x Spanish goat kids. Fifteen (n = 5 per treatment) goats were fed 0, 100, or 200 mg of supplemental Cu per day as copper sulfate for 98 d. The animals were slaughtered, and LM, s.c. adipose from the sternal region, and mesenteric adipose tissues were collected. Total lipids were extracted with chloroform:methanol (2:1), methylated and isolated via GLC from all tissues. The subsequent peaks were then positively identified by mass spectrometry. Thiobarbituric acid-reactive substances were measured also. In s.c. adipose, dietary Cu significantly decreased C14:0 (P = 0.03) and C16:0 (P = 0.01). In muscle, C15:0 (P = 0.03) was linearly increased by Cu. Dietary Cu supplementation did not influence oxidative stability in goat muscle or s.c. adipose. Copper supplementation at 200 mg/d resulted in a significant increase in malondialdehyde in mesenteric adipose (P = 0.01) compared with the 0 or 100 mg/d groups. These results indicate that lipid composition may differ from depot to depot and that depending on the depot, dietary Cu seems to elicit a variable response on the fatty acid composition.

Sixteen multiparous Jersey cows were assigned at calving to one of 4 dietary treatments. An 18% crude protein (CP) diet was fed as a total mixed ration through 30 d in milk (DIM), and beginning at 31 DIM a 9, 12, 15, or 18% CP diet was fed through 58 DIM (depletion). All cows were then fed the 18% CP diet until 84 DIM (repletion). Muscle biopsies were taken under local anesthesia at 49 and 84 DIM from the semitendinosus muscle. Milk production, DMI, and milk component contents were measured. Calpain and calpastatin contents of muscle taken at biopsy were evaluated using Western blotting techniques. Milk production and milk protein content were reduced during the depletion period by decreasing dietary protein. Diet had no effect on milk fat content or DMI. During repletion, DMI was affected by dietary treatment. Western blots of muscle extracts indicated no differences in calpain content at any stage of the experiment or in calpastatin content of muscle at 49 DIM. However, at 84 DIM, calpastatin (135 kDa) was lower or undetectable in cows fed either the 9 or 12% CP diets from 31 to 59 DIM. Bands for a 110-kDa degradation product of calpastatin were present in some cows fed the 9, 12, and 15% CP diets during the depletion period. Results indicate a change in skeletal muscle calpain/calpastatin proteolytic system during protein repletion following depletion with diets of less than 15% CP during early to peak lactation in dairy cows.

The objectives were to determine effects of prepartum protein intake and dietary amino acid balance on production, adaptations in body fat and protein, amino acid concentrations, and, indirectly, body protein breakdown in early lactation. Multiparous Holstein cows (n = 42) were fed diets containing 11 or 14% crude protein with or without 20 g/d of methionine hydroxy analog for 21 d prepartum and then fed a common diet of 17% crude protein for 120 d postpartum, with or without 50 g/d of methionine hydroxy analog. Dry matter intake postpartum averaged 25.4 kg and milk production 41.6 kg. Cows fed the 14% CP diet ate 0.7 kg more dry matter and gave 1.7 kg more milk than those fed the 11% diet postpartum, but this difference was not significant. Cows fed methionine hydroxy analog prepartum lost less body protein from -14 to 60 d in milk. From d 60 to 120, body fat increased 8.5 and 11.5 kg for low and high protein groups and body protein increased 0.5 and 1.0 kg. Serum concentrations of branched chain amino acids fell 17% in the first few weeks postpartum, lysine fell 15%, histidine fell 16%, methionine increased 20%, and cysteine increased 30%. The ratio of serum 3-methylhistidine to creatinine was determined to indicate muscle protein degradation. An increase in this ratio at 7 d postpartum indicated increased body protein breakdown, there was no effect of prepartum ration. Increased protein intake prepartum may allow more feed intake and milk production postpartum, and supplementing a methionine analog on a ration already balanced in methionine by contemporary models may spare body protein.

Eighteen male English Pointers (2-4 years of age, 23.94+/-0.54 kg body weight) were allotted to three diet and two physical conditioning groups to evaluate the effect of level and source of dietary fat on the olfactory acuity of canine athletes subjected to treadmill exercise. Diet groups (6 dogs/diet) consisted of commercially prepared diets (minimum of 26% crude protein) containing 12% fat as beef tallow (A), 16% fat provided by equivalent amounts of beef tallow and corn oil (B), or 16% fat provided by equivalent amounts of beef tallow and coconut oil (C). This dietary formulation resulted in approximately 60% of the total fatty acid being saturated for diets A and C, while approximately 72% of the total fatty acids were unsaturated in diet B. One-half of the dogs within each dietary group were subjected to treadmill exercise 3 times per week for 30 min (8.05 km/h, 0% grade) for 12 weeks. All dogs were subjected to a submaximal exercise stress test (8.05 km/h, 10% slope for 60 min) every four weeks beginning at week 0. Olfactory acuity was measured utilizing behavioral olfactometry before and after each physical stress test. Non-conditioned (NON) dogs displayed a greater decrease (P<0.05) in olfactory acuity following exercise, while physically conditioned (EXE) dogs did not show a change from pre-test values. A diet by treatment interaction (P<0.10) was detected over the course of the study. NON dogs fed coconut oil had decreased odorant-detecting capabilities when week 4 values were compared with week 12 values. Feeding a diet that is predominately high in saturated fat may affect the odorant-detecting capabilities of working dogs. Additionally, these data indicate that utilization of a moderate physical conditioning program can assist canine athletes in maintaining olfactory acuity during periods of intense exercise.

Effects of dietary ascorbyl-2-phosphate on immune function after a 210-km trip were measured in 18 Holstein heifers. After transport on d 0, 10 g of ascorbyl-2-phosphate each were added to the diets of 10 heifers, whereas eight heifers were fed a control diet. Plasma cortisol concentrations increased by an average of 25.6 microgram/ml on d 0 following transport, but by d 7 after transport had decreased to pretransport levels. Average daily gain was lower in heifers fed ascorbyl-2-phosphate from d 28 to 49 d after transport, but did not differ over the entire study. Feeding ascorbyl-2-phosphate maintained plasma ascorbate concentrations on d 7 post-transport, which decreased in control heifers. Plasma keyhole limpet hemocyanin antibody titers were significantly higher in control heifers from d 7 to 49. Mononuclear leukocyte proliferation responses were decreased on d 0 in lymphocytes stimulated by mitogens, with pokeweed mitogen-stimulated cells showing less of a response than cells stimulated by the other mitogens. In the absence of mitogens, dietary ascorbyl-2-phosphate increased basal 3H-methyl thymidine incorporation by cultured lymphocytes. Across diets and mitogens, lymphocytes treated with cortisol showed decreased 3H-methyl thymidine incorporation. Transportation acted as a stressor, as evidenced by the increased plasma cortisol levels at d 0 immediately after transport, but immunological effects were not apparent by d 7. Feeding ascorbyl-2-phosphate maintained plasma ascorbate concentrations on d 7, but had negative effects on immune responses posttransport.

Thirty-six adult female Beagles averaging 2 (young-adult) or 8 (geriatric) yr of age were used to assess the effects of graded levels of dietary protein (16, 24, or 32%) on endocrine-controlled regulation of whole-body protein turnover. Rates of whole-body protein synthesis (WBPS) and whole-body protein degradation (WBPD) were estimated using orally administered 15N-glycine and total excreta collection. Although N balance was similar for all dogs, N flux through the metabolic pool increased linearly (P < 0.05) as protein intake increased. Rates of WBPS, WBPD, or the difference between them were not influenced by age (P > 0.10). A quadratic increase (P < 0.05) in WBPS and WBPD was observed in response to dietary protein. Serum insulin-like growth factor-I (IGF-I), IGF-I-binding protein 3, and total IGF-I-binding proteins were higher (P < 0.05) in geriatric dogs than in young-adult dogs regardless of protein intake. These results indicate that dietary protein in excess of 16% may not be required to maintain N balance in young-adult and aging dogs despite the linear increase in N flux through the metabolic pool. Furthermore, age-induced changes in endocrine functionality may differ between dogs and other species.

Fifty-five multiparous Holstein dairy cows were used to evaluate the singular and combined effects of somatotropin and monensin treatments during the late dry period on postpartum metabolism and production. Treatments were 1) control (C); 2) injection of exogenous bovine somatotropin (bST); 3) TMR top dressed with 300 mg of monensin/day (M); and 4) monensin and somatotropin in combination (bST+M) during the last 28 days before expected parturition. A 500-mg subcutaneous injection of sustained release somatotropin was administered adjacent to the tail head at d -28 and -14 relative to expected calving. Diet and management were the same for all cows after parturition. Production and intake were measured daily until 63 d in milk. Milk composition, blood metabolites, and body weight and condition score were measured weekly. Prepartum glucose, nonesterified fatty acid, and blood urea nitrogen concentrations were not different among treatments. Cows on the M treatment tended to have greater dry matter intake postpartum than those on the C treatment and 30% lower plasma nonesterified fatty acid concentrations during wk 1 postpartum than all other treatments. Milk yield and milk fat yield were not different among treatments, but milk fat percent tended to be lower for the bST+M treatment than the C treatment. Changes in plasma amino acid concentrations were consistent with mobilization of skeletal muscle protein, possibly for use in gluconeogenesis. Results from this study provide evidence that prepartum feeding of monensin reduced plasma nonesterified fatty acid concentrations and may improve glucose metabolism of the periparturient dairy cow.

Sixty-four individually housed pigs were used to investigate the effect of amino acid content of finisher diets on growth performance of pigs subjected to marginal dietary amino acid restrictions (80% of the 1988 NRC lysine recommendation) during the grower phase. In each of the two trials, low- and high-amino-acid grower diets (.421 and .765 g lysine/MJ DE, respectively) and four finisher diets (.421, .516, .612, and .707 g lysine/MJ DE) were randomly assigned within sex to 16 gilts and 16 castrated males weighing 23.0 +/- 2.0 kg in a 2 x 4 factorial arrangement of treatments. The average weight of pigs after a completion of diet change was 50.4 +/- 2.1 kg. All pigs were slaughtered at an average weight of 105.2 +/- 4.1 kg. Ultrasound backfat thickness was measured at the time of diet change and before slaughter. Pigs were allowed ad libitum access to feed and water. During the grower phase, pigs fed the high-amino-acid diet grew faster (P < .001) and more efficiently (P < .001) and had less ultrasound backfat (P < .001) than those fed the low-amino-acid diet. The grower diet had no effect on weight gain during the finisher phase. Consequently, pigs fed the high-amino-acid grower diet had better overall weight gain (P < .01) than those fed the low-amino-acid diet. The rate of lean accretion was, however, similar between the two groups of pigs. Furthermore, pigs fed the low-amino-acid grower diet seemed to have better carcass quality, as indicated by less ultrasound backfat (P < .01) and larger carcass longissimus muscle area (P < .05). Average and 10th rib carcass backfat decreased linearly (P < or = .05) and lean accretion rate improved linearly (P < .05) as the amino acid content of finisher diets increased, but there was no grower x finisher diet interaction in these and other response criteria. Although no evidence of compensatory weight gain was observed, it is possible that compensatory lean tissue growth may have occurred in pigs subjected to early amino acid restrictions at the expense of fatty tissue growth.

Whole-body influx and efflux of K(+) were determined for 25-day-old striped bass,Morone saxatilis, in conditions that simulated harvesting fish from ponds. During the first 5h in fresh water with combined high NaCl (80 mM) and low Ca(2+) (0.12 mM) concentrations, a combination that is acutely lethal to this age of striped bass, K(+) influx for fish in 0.07 mM K(+) was 21±1.7 (SEM) compared to 3.4±0.33 nmol g(-1) h(-1) for fish in water with low Na(+) (0.25 mM) or high Ca(2+) (2.5 mM) concentrations. Influx of K(+) was inhibited during the first few hours after fish were placed in flux chambers. Potassium efflux as percentage of(42)K lost per hour was two-fold higher from fish in the high Na-low Ca treatment compared to fish in low concentrations of Na(+) or high concentrations of Ca(2+). Potassium efflux was probably much greater than influx, but exact values for efflux could not be calculated from the data available. Survival of fish in water with high Na-low Ca was not increased by addition of KCl to the water, indicating that the net loss of K(+) was probably not the cause of death.

Twelve abomasally cannulated wether lambs were fed isocaloric diets containing 9, 12 or 15% crude protein to determine insulin-like growth factor-I (IGF-I) responses to altered abomasal nitrogen flow and nitrogen status. Lambs were offered 1100 g/d of their respective diets. Voluntary feed consumption was not affected by nitrogen intake. Ruminal and total tract digestibilities of dry matter, organic matter and nitrogen increased linearly (P < 0.05) with increased dietary nitrogen. Abomasal flows of total, bacterial and rumen escape nitrogen increased (linear, P < 0.01), whereas dry matter and organic matter flows decreased (linear, P < 0.01). Total amino acid flow was greater (linear, P < 0.01) in lambs fed additional nitrogen due to increased (linear, P < 0.01) flows of essential and nonessential amino acids. Nitrogen retention and blood urea nitrogen increased linearly (P < 0.01). Serum IGF-I concentrations and relative hybridization intensity of hepatic IGF-I mRNA increased (linear, P < 0.05) as lambs consumed more nitrogen. Serum IGF-I and hepatic IGF-I mRNA were correlated positively (P < 0.05) with nitrogen intake and abomasal flows of nitrogen and various amino acids. These data provide evidence of a relationship between abomasal amino acid flow, as influenced by nitrogen intake, and hepatic gene expression and serum concentrations of IGF-I in growing lambs.

Twenty-four (24) mature, mixed breed, healthy dogs weighing from 14.6 kg to 27.6 kg were used to study the effects of various steroids on the olfactory function of the dog using olfactory detection threshold as an index. Two odorants were used, viz; benzaldehyde and eugenol. Of the various steroids used, only dexamethasone produced classical signs of Cushing's syndrome in the dogs. However, all dogs that received either dexamethasone alone or hydrocortisone plus DOCA exhibited a significant elevation in the olfactory detection threshold for both odorants without any observable structural alteration of the olfactory tissue using light microscopy. On the other hand, neither DOCA, hydrocortisone alone, nor any of the vehicles used in the study significantly altered the olfactory function of the dogs. The results show that Cushing's syndrome can be experimentally produced in dogs using exogenous steroids and that this condition diminishes the olfactory capability of the dog without producing classical signs of the disease.

Twenty-seven multiparous Holstein cows averaging 120.7 DIM, 9 per diet, were assigned at random to 20, 17, or 14% ADF diets of corn, corn silage, and soybean meal, following a covariant period when all cows received the 14% ADF diet. An 18-d period (period 1) of cool weather preceded the onset of high environmental temperature from June 1988 to the end of July 1988 (period 2). Mean daily maximum air temperatures were 35.2, 36.8, and 34.7 degrees C for the covariant period and for periods 1 and 2, respectively. Minimum temperatures were 14.5, 16.0, and 21.5 degrees C. Milk production during both periods was higher, and the decrease in milk production associated with the onset of hot weather was lower, in cows fed the 14% ADF diet. Intake was not affected by diet during period 1 but was lower in cows fed the 17 and 14% ADF diets in period 2 relative to period 1. At any given environmental temperature, DMI was higher in cows fed lower ADF diets. Although DMI declined more rapidly with increasing daily minimum temperature with lower ADF diets, milk production was less sensitive to daily minimum temperature in cows fed the 14% ADF diet.

A study was initiated to determine whether development of a functional ruminant digestive system was associated with alterations in plasma growth hormone (GH) concentration. Holstein bull calves were fed milk or milk with grain until studied at the age of 1 month (n = 12). Calves placed on pasture with some grain supplementation were studied at the age of 3 months (n = 6) to determine plasma GH concentration in an animal with fully developed ruminant metabolism. Blood samples were taken at 10-minute intervals for 5 hours, followed by administration of bovine GH-releasing factor (0.075 micrograms/kg of body weight) and subsequent blood sample collection for 1 hour. On the following day, a blood sample was collected via jugular cannula, clonidine (10 micrograms/kg) was administered, and blood samples were subsequently obtained. Data indicated that milk-fed calves had higher mean plasma GH concentration than did either milk/grain-fed or older calves. The difference in mean plasma GH concentration was related to higher secretory pulse amplitude. Pituitary responses to bovine GH-releasing factor did not differ among the 3 groups, but response to clonidine were greater in milk-fed calves than in calves of the other groups. These data indicate that the change from a nonruminant to a ruminant-type gastrointestinal tract, perhaps attributable to subsequent changes in metabolism, may induce changes in hypothalmic function to decrease GH concentration.

Multiparous Holstein cows at six universities were utilized to examine effects of ruminally protected methionine and lysine on lactational performance. Three hundred and four cows began the study; 259 cows were included in the production analysis. Following a 21-d standardization period, cows received a basal diet of corn silage and ground corn supplemented with one of five dietary treatments, which were supplements of soybean meal or corn gluten meal, the latter with zero and three combinations of protected methionine and lysine (methionine; methionine and lysine; methionine and double (2x) lysine). Treatment effects were evaluated during early, mid, late, and total lactation (22 to 112, 113 to 224, 225 to 280, and 22 to 280 d postpartum, respectively). On a DM basis, ratios of forage to concentrate (50:50, 60:40, and 70:30) increased, and dietary CP (16.0, 14.5, and 13.0%) decreased during the three periods of lactation. Amount of amino acid supplementation also decreased (15, 12, and 9 g/d methionine; 20, 16, and 12 g/d lysine; and 40, 32, and 24 g/d 2x lysine) with period of lactation. Actual and least squares means for milk, FCM, and milk protein yields were greater for soybean than for corn gluten meal during early, mid, and total lactation. In addition, these variables responded linearly to lysine in early lactation. Response to lysine was quadratic during mid and total lactation for these variables. Differences in nutrient intake explained production responses to protein sources but not to lysine. Serum amino acid responses primarily reflected differences in dietary protein source and rumen-protected amino acid.

The effect of housing calves in indoor metal pens (1.2 m x 1.2 m) or commercial calf hutches was determined on plasma concentration of cortisol, antibodies, and ascorbate. Six calves per treatment were deprived of colostrum, assigned randomly to treatment, and fed a commercial milk replacer until 56 d of age. Calves housed in hutches had higher plasma IgG concentrations than calves in pens at 42 and 56 d of age. Housing had no effect on plasma IgM concentration. Antibody titers to keyhole limpet hemocyanin injected at 14 and 28 d of age were higher in hutch-housed calves from 21 to 56 d of age. Calves housed in hutches also had lower plasma cortisol concentrations, although cortisol decreased with age in both treatment groups. Plasma ascorbate and ascorbate plus dehydroascorbate were higher in hutch-housed calves. Regression analysis indicated a positive relationship between plasma ascorbate and IgG in metal penhoused calves and a negative relationship in hutch-housed calves. Housing in 1.2 m x 1.2 m metal pens decreases cortisol, plasma ascorbate, IgG, and specific antibody titers in dairy calves relative to calves housed in hutches.

Effect of feeding high fat diets on peripheral plasma concentrations of immunoreactive insulin, glucagon, and growth hormone following intravenous glucose challenge (100 mg D-glucose/kg body weight) at 50 and 100 d of lactation in 16 multiparous Holstein cows was evaluated. The high fat diet contained 18.5% whole cottonseed on a dry matter basis as the source of extra dietary fat. Feeding the high fat diet had no apparent effect on energy balance. Basal plasma insulin and insulin:glucagon ratio were increased in cows fed the high fat diet relative to those of controls. Glucagon, insulin, and insulin:glucagon ratio response to glucose challenge were not affected by diet. Response of growth hormone to glucose challenge at 50 d of lactation was depressed in cows fed the high fat diet. Plasma glucose peaked at greater concentrations in cows fed the high fat diet. Feeding a high fat diet to dairy cows appears to increase basal insulin concentration and insulin:glucagon ratio, which has actions opposed to glucose synthesis. Thus, endocrine effects of a high fat diet appear to favor decreased milk production.

Early lactation in the dairy cow is associated with an increased demand for energy that can only be met by hormone-mediated partitioning of nutrients. The purpose of this study was to determine adaptive responses of basal and glucose-stimulated glucagon (IRG), insulin (IRI), and growth hormone (GH) concentrations to early lactation. Blood was collected via jugular cannulas from nonpregnant nonlactating (NPNL) cows and cows 14 days antepartum (AP) and 5 and 30 days postpartum (PP). Basal concentrations of IRI decreased with lactation, IRG was essentially unchanged, and GH was increased with lactation. The molar IRI/IRG (I/G) ratio was decreased from 4.6 at day 14 AP to 1.3 at day 30 PP. The effects of exogenous glucose (0.56 mM/kg body wt) on IRI were greatest during pregnancy and declined with lactation. IRG responses to glucose were similar except in NPNL cows, which had a minimal but prolonged IRG inhibition, GH responses to glucose infusion were absent in NPNL cows and were most significant at 5 days PP. The decreased I/G and elevated GH concentrations suggests that both IRG and GH play a role in nutrient partitioning during early lactation.

Colostrum-fed, colostrum-deprived, and colostrum-fed and colostrum-deprived calves fed ascorbic acid (1.75 g/d) in whole, raw milk to 6 wk of age were sampled from 0 to 8 wk of age in order to determine whether ascorbate supplementation would increase plasma Ig concentrations, antibody response to immunization, and disease resistance. Plasma IgG concentrations were lower at 14 and 28 d of age in calves fed ascorbate compared with plasma concentrations in calves not receiving ascorbate supplementation, irrespective of colostrum feeding. Colostrum feeding had no effect on antibody titer to keyhole limpet hemocyanin at any age, but ascorbate-supplemented calves had lower plasma antibody titers to keyhole limpet hemocyanin at 35 and 56 d of age. Calves fed ascorbate had lower clinical scores for diarrhea. Dietary ascorbate does not appear to be immunostimulatory in dairy calves to 56 d of age and appeared to inhibit antibody synthesis. However, at 14 d of age there was an interaction of ascorbate supplementation and colostrum feeding; plasma IgG concentrations were higher in colostrum-deprived calves fed ascorbate then in colostrum-deprived calves not fed ascorbate.

The objective was to determine the efficacy of a blend of ammonium salts of the volatile fatty acids, isobutyric, 2-methylbutyric, isovaleric, and valeric as a supplement to diets for dairy cows. Treatments of 0 (control) or 120 (supplemented) g/cow of the blend were fed daily from approximately 3 wk prepartum through a complete lactation. Five trials were conducted concurrently with a total of 116 multiparous Holstein cows. Dietary ingredients or combinations of ingredients differed in each of the trials. Diets contained either 1) corn gluten meal and urea, 2) soybean meal, or 3) cottonseed meal as the primary grain source of crude protein. The forage portion of the diets contained corn silage in combination with one or more of the following: alfalfa hay, alfalfa haylage, or wheat silage. Cows fed the supplement produced more milk and fat-corrected milk than the control cows for the 305-d lactation on four of the five diets, resulting in an average increase of 1.7 kg/d or 7%. Feed intake of cows on the supplemented diet was generally similar or lower than intake of the control cows throughout lactation, indicating that increased milk yield was associated with improved feed utilization. Percent milk fat was similar for cows on the supplemented diet, but fat yield was higher. Percent milk protein was lower for supplemented cows, but protein yield was about the same for both treatments because of higher milk yield. Health and reproduction were similar for all cows.

Concentrations of insulin, glucagon, growth hormone, adrenocorticotropin, and cortisol were determined in plasma samples obtained at 20-min intervals for 6 h from low and high producing dairy cows at d 30 and 90 postpartum. Four nonpregnant, nonlactating cows also were sampled. Insulin concentrations were reduced at d 30 in both groups of lactating cows compared with concentrations in nonlactating cows; glucagon concentrations were unchanged. The molar insulin: glucagon was reduced at d 30 in both groups and at d 90 for low, but not high producers. Growth hormone concentrations were higher at d 30 in high producers than at d 90, in low producers at d 30, and higher than in nonlactating cows. Cortisol concentrations were lower in high producing cows at d 30 than at d 90 or in nonlactating cows due to a reduced pulse amplitude. No differences were observed for adrenocorticotropin. Reduced molar insulin: glucagon may be an integral response of the cow to lactation, while the difference in the insulin: glucagon for high and low producers at d 90 postpartum may indicate a continued need for a gluconeogenic stimulus in low producers. The elevated growth hormone and low cortisol concentrations likely participate in the enhanced production observed in high producing dairy cows.

Altered concentrations of metabolic hormones have been suggested as important mediators of energy partitioning during early lactation. This study was initiated to determine the effects of propionate (1.0 mmol/kg body weight) infusion on plasma concentrations of glucagon, insulin, growth hormone, propionate and glucose at 14 days ante-partum (AP) and days 5 and 30 postpartum (PP). No differences were seen in propionate concentrations between sampling days. Glucose concentrations were elevated following propionate infusion in pregnant cows but were not elevated in the PP cows. Insulin responses to propionate infusion did not differ between days while the glucagon response was blunted at day 5 PP. Basal glucagon concentrations were elevated between days 5 and 30 PP, insulin concentrations were unchanged between days, while the molar insulin/glucagon ratio was decreased during early lactation. Basal growth hormone (GH) concentrations were elevated between day 14 AP and day 30 PP. GH responsiveness to declining propionate concentrations was greatest at day 5 PP. These data further suggest a role for glucagon as well as GH in nutrient partitioning during early lactation.

Dairy cows in early lactation were reported to secrete growth hormone in response to declining glucose concentrations at day 5 but not day 30 post-partum, whereas GH responses to TRH were reported to be enhanced after day 30 post partum. The present study examined GH response following glucose infusion and the effect of GHRH as well as effects of SRIH on GHRH-stimulated GH release. Declining plasma glucose concentrations after glucose infusion stimulated GH release at day 5 post partum but not in nonpregnant, nonlactating cows or in cows at days 30 and 90 post partum. GHRH stimulated GH release on all days tested, but the response was highest at day 30 post partum when compared to other days. Somatostatin infusion inhibited GHRH effects on GH concentrations only at day 30 post partum and in nonpregnant, nonlactating cows. Thus, a differential response of the GH regulatory system could be demonstrated between days 5 and 30 post partum utilizing different stimuli. Evaluation of plasma glucose and free fatty acid concentrations on days 5, 10, 20, and 30 post partum revealed a progressive decrease in FFA but not glucose as lactation progressed. Decreased plasma FFA concentrations were paralleled by a decrease in basal GH, somatomedin-C and epinephrine. Thus, a decline in FFA may be responsible for the disparity between effects of GHRH and glucose on GH release between days 5 and 30 post partum.

Efficacy of multiple and single intramammary infusions of benzathine cloxacillin (500 mg/quarter) were compared for treatment of mastitis during the dry period. Treatments were three infusions, one each at 0, 7, and 14 d into the dry period; one infusion at drying off; and no infusions. Cows were grouped (30/treatment) based on previous mastitis history, mean monthly SCC, mature equivalent milk production, and pathogen detected pretreatment. Infected quarters/cow at parturition, posttreatment were 1.26, 1.6, and 2.35 for multiple, single, and no infusion groups, respectively. Milk production and somatic cell count posttreatment were not affected by treatment. Across all genera of microorganism, 75.5, 73.6, and 52.8% of infections were eliminated in the multiple, single, and no infusion groups, respectively. Primary effect of multiple infusions with cloxacillin in the dry period was to prevent new streptococcal infections. Sensitivity tests on staphylococcal isolates indicated that cloxacillin was still an effective dry cow treatment after 7 yr of use in the herd. Resistance to cloxacillin in approximately half the posttreatment staphylococcal isolates appeared to be nonenzyme (penicillinase) dependent. Multiple dry cow treatments with cloxacillin do not offer any advantage over a single treatment.

Ten calves less than 14 h of age (average 8.6 h) were anaesthetized, and the intestine was ligated into segments 10 cm in length at 3-cm intervals beginning 1.8 m anterior of the ileocecal junction and proceeding proximally. Seven treatments were assigned in random order to segments in three successive sections of the small intestine. Segments received 1 ml of viable bacteria of intestinal origin, autoclaved bacteria of intestinal origin, or sterile microbiological broth at zero time; then after 4 h they were injected with iodine-125 labeled gamma-globulin. After an additional 1.5 h, the experiment was ended and uptake assayed. Two treatments measured anaerobic microbial growth after 4 h incubation with 1 ml of either sterile broth or live bacteria culture. Residual [iodine-125] gamma-globulin was measured in segments receiving 1 ml of sterile broth or live bacteria culture with 5.5-h incubation followed by 15-s exposure to labeled gamma-globulin. Uptake was lowest in segments receiving live bacteria as compared to segments receiving sterile inocula. Number of bacteria per gram of tissue was correlated negatively with uptake. Low corticosteroids in serum were associated with low uptake of gamma-globulin.

Twenty-one Holstein bull calves were randomly assigned at birth to 3 groups. Two groups (each of 7 calves) were raised as follows: fed a milk diet alone or fed milk with grain supplementation after 2 weeks of age; studies were done when calves reached 4 weeks of age. The 3rd group was fed on milk with grain supplementation until weaning after which the calves were maintained on grain and pasture. These calves (older calves) were studied at 12 weeks of age. Either propionate (0.28 mmol/kg) or glucose (0.56 mmol/kg) was injected IV in a random order. Samples of blood were obtained from the calves before and immediately after injections were done and at 2, 5, 10, 15, 30, 45, and 60 minutes after secretagogue injection. Plasma was examined for glucose by a glucose oxidase procedure and for immunoreactive insulin (IRI) and glucagon (IRG) by radioimmunoassay. The IRI response to the injection of glucose was greater in older calves (P less than 0.02). Patterns of IRI secretion, as determined by heterogeneity of regression, showed age differences for both secretagogues (P less than 0.05). Base-line IRG was greater in milk/grain-fed calves than in milk-fed calves (P less than 0.05). Mean IRG response to propionate injection was higher (P less than 0.05) in milk/grain-fed calves than in milk-fed calves. Plasma glucose concentration increased in older calves, but decreased in milk-fed calves after propionate injection. The data indicate that maturation in the ruminant is accompanied by altered regulation of insulin and glucagon secretion.(ABSTRACT TRUNCATED AT 250 WORDS)

Eight lactating Holstein dairy cows were fed corn silage-based diets with or without whole cottonseed at 18.5% of the dietary dry matter. At 42 days postpartum, a pulse injection of 100 mg glucose/kg body weight was given intravenously and plasma glucose concentration was monitored for 45 min. At 50 d postpartum, biopsies of adipose tissue and mammary tissue were taken and tissue slices were incubated in vitro with either uniform carbon-14 glucose or 1-carbon-14 palmitate. Basal concentration of plasma glucose was not affected by diet, although apparent distribution volume of glucose did seem to decrease due to feeding whole cottonseed. Feeding whole cottonseed decreased uptake of glucose and palmitate in both adipose tissue and mammary tissue and also decreased oxidation of glucose to carbon dioxide in both tissues. Palmitate oxidation was not affected by diet. Incorporation of carbon-14 from glucose into adipose tissue lipids was decreased in cows fed whole cottonseed. Results indicate that fat supplementation in the form of whole cottonseed may decrease palmitate incorporation and glucose utilization for glycerol and reducing equivalent synthesis in both adipose tissue and mammary tissue of lactating dairy cows.

A study was conducted to determine the effect of transportation stress on ovarian function in superovulated heifers. Thirty cyclic Hereford heifers of similar age and weight and in good body condition were randomly assigned to control and stress-treated groups. All animals received two daily injections of 5 mg follicle stimulating hormone (FSH) for 4 d beginning on Day 10 to 12 of the estrous cycle. A blood sample was collected at each FSH injection. On the fourth day of injections, heifers were given 25 mg prostaglandin F(2)alpha (PGF(2)alpha) in the morning and a second injection of 15 mg PGF(2)alpha in the afternoon. During superovulation, the stressed heifers were transported to a different location every 12 h whereas control animals remained at the pretrial site. Following 4 d of intermittent transporting and FSH treatment, stress-treated heifers were recombined with control animals. Ovaries were examined 8 d following the onset of standing estrus to determine length, width, thickness, and number of corpora lutea (CL). Peripheral plasma levels of cortisol were higher in the stressed group (P< 0.1). Least squares means for numbers of CL were 20.4 +/- 2.1 and 15.4 +/- 1.7 for control and treated heifers, respectively (P< 0.1). There were no treatment differences (P> 0.1) between length, width, or thickness of ovaries when the number of CL was held constant. These data suggest that stress of the type, intensity, and duration imposed in this study increased plasma levels of cortisol and reduced ovulation rate as determined by CL formation in superovulated heifers.

With 32 lactating Holstein cows we evaluated physiological effects of gossypol and cyclopropenoid fatty acids in diets containing 18.5% whole cottonseed (dry matter) based on corn, corn silage, and soybean meal. All cows consumed a control diet for the first 2 wk of lactation and then were assigned to either control or whole cottonseed diet for the remainder of their lactation. Milk production, milk fat and protein percentages, and daily dry matter intake were measured. Dry matter intakes were less for cows consuming the whole cottonseed diet, but net energy intake was similar for all diets. Milk fat from cows consuming whole cottonseed contained detectable concentrations of cyclopropene fatty acids. Total lipid in plasma, total serum cholesterol, serum gossypol, and apparent liver gossypol concentrations were greater in cows fed whole cottonseed. Gossypol and cyclopropenoid fatty acids appeared to be absorbed from the gut of cows fed whole cottonseed. Small amounts of gossypol in serum and liver tissue and small amounts of cyclopropene fatty acids in adipose tissue lipids and milk fat indicate a need to elucidate the significance of these physiologically active compounds in the human diet and their biological effects on lactating dairy cows.