Effect of the pelleting process on diet formulations with varying levels of crystalline amino acids and reducing sugars on digestibility in growing pigs.

The objective of this experiment was to determine the effects of pelleting on the standardized ileal digestibility (SID) of amino acids (AA) and crude protein (CP) in diets with or without increased concentrations of free AA and reducing sugars (RS). Eight individually housed, ileal cannulated barrows (initially 31.4 kg) were allotted to an 8 × 8 Latin square with eight diets and eight 7-d periods with ileal digesta collected on days 6 and 7. Treatments were arranged in a 2 × 2 × 2 factorial with the main effects of diet form (mash or pellet), crystalline AA (low or high), or RS (low or high), provided by distillers dried grains with solubles and bakery meal. Diets were pelleted to achieve a hot pellet temperature of 85 to 88 °C. Data were analyzed as a Latin square design using the GLIMMIX procedure of SAS 9.4. A feed form × RS interaction (P < 0.026) for SID of tryptophan was observed. Feeding pelleted low RS diets increased SID of tryptophan compared with mash high and low RS diets, and pelleted high RS diets. For the main effects of feed form, the SID of total AA, CP, and indispensable AA was greater (P < 0.042) in pelleted diets compared with mash diets. For the main effects of crystalline AA, pigs fed high crystalline AA had increased (P = 0.007) SID of tryptophan and decreased (P = 0.050) SID of histidine compared with those fed low crystalline AA diets. For the main effects of RS, high RS diets had decreased (P < 0.05) SID of total AA, CP, and indispensable AA compared with low RS diets. In conclusion, pelleting diets increased AA digestibility, and pelleting diets with increased crystalline AA or RS did not affect the improvement in AA digestibility from pelleting. Diets formulated with high crystalline AA had increased SID of tryptophan. Formulating diets with high RS resulted in decreased AA digestibility compared with corn-soybean meal-based diets.

The objective of this study was to determine the effects of pelleting on the standardized ileal digestibility (SID) of amino acids (AA) in diets with or without increased concentrations of free AA and reducing sugars (RS). Treatments were arranged in a 2 × 2 × 2 factorial with the main effects of diet form (mash or pellet), crystalline AA (low or high), or RS (low or high), provided by dried distillers grains with solubles and bakery meal. A total of 8 illeal cannulated barrows were fed treatments in an 8 × 8 Latin square design. Results indicated that there was no evidence of interactions between diet types and diet form, indicating that increasing amounts of crystalline AA and RS did not reduce amino acid digestibility when pelleting diets. Additionally, pelleting diets resulted in increased amino acid digestibility compared to mash diets. Diets formulated with 20% dried distillers grains with solubles and 15% bakery resulted in decreased amino acid digestibility compared with the corn­soybean meal-based diets. Crystalline amino acid concentration did not influence amino acid digestibility of indispensable AA, except for SID of tryptophan which was increased in diets with higher concentrations of crystalline AA.

Prenatal inflammation reprograms hyperactive ILC2s that promote allergic lung inflammation and airway dysfunction.

Allergic asthma is a chronic respiratory disease that initiates in early life, but causal mechanisms are poorly understood. Here we examined how prenatal inflammation shapes allergic asthma susceptibility by reprogramming lung immunity from early development. Induction of Type I interferon-mediated inflammation during development provoked expansion and hyperactivation of group 2 innate lymphoid cells (ILC2s) seeding the developing lung. Hyperactivated ILC2s produced increased IL-5 and IL-13, and were associated with acute Th2 bias, eosinophilia, and decreased Tregs in the lung. The hyperactive ILC2 phenotype was recapitulated by adoptive transfer of a fetal liver precursor following exposure to prenatal inflammation, indicative of developmental programming. Programming of ILC2 function and subsequent lung immune remodeling by prenatal inflammation led to airway dysfunction at baseline and in response to papain, indicating increased asthma susceptibility. Our data provide a link by which developmental programming of progenitors by early-life inflammation drives lung immune remodeling and asthma susceptibility through hyperactivation of lung-resident ILC2s. One Sentence Summary: Prenatal inflammation programs asthma susceptibility by inducing the production of hyperactivated ILC2s in the developing lung.

Both maternal IFNγ exposure and acute prenatal infection with Toxoplasma gondii activate fetal hematopoietic stem cells.

Infection directly influences adult hematopoietic stem cell (HSC) function and differentiation, but the fetal hematopoietic response to infection during pregnancy is not well-studied. Here, we investigated the fetal hematopoietic response to maternal infection with Toxoplasma gondii (T. gondii), an intracellular parasite that elicits Type II IFNγ-mediated maternal immunity. While it is known that maternal infection without direct pathogen transmission can affect fetal immune development, the effects of maternal IFNγ on developing HSCs and the signals that mediate these interactions have not been investigated. Our investigation reveals that the fetal HSCs respond to T. gondii infection with virulence-dependent changes in proliferation, self-renewal potential, and lineage output. Furthermore, maternal IFNγ crosses the fetal-maternal interface, where it is perceived by fetal HSCs. By comparing the effects of maternal IFNγ injection with maternal T. gondii infection, we reveal that the effects of IFNγ treatment mimic some aspects of the fetal HSC response to infection. Moreover, our findings illuminate that the fetal HSC response to prenatal infection is distinct from the adult HSC response to IFNγ-induced inflammation. Altogether, our data disentangle the role of infection-induced inflammatory cytokines in driving the expansion of downstream hematopoietic progenitors.

Nutritional value of a new source of cheese coproduct fed to weanling pigs.

Three experiments were conducted to test the hypothesis that values for standardized ileal digestibility (SID) of amino acids (AA) and metabolizable energy (ME) in a the cheese coproduct are greater than in fish meal or enzyme-treated soybean meal (ESBM). The second objective was to test the hypothesis that pigs fed a diet containing cheese coproduct will have growth performance that is not different from that of pigs fed other sources of protein. In experiment 1, eight ileal-cannulated barrows (11.0 ± 0.4 kg) were allotted to a replicated 4 × 4 Latin square design with four diets and four periods and two pigs per diet in each period. The four diets included an N-free diet and three diets that contained ESBM, fish meal, or the cheese coproduct as the source of AA. Results indicated that the cheese coproduct had greater (P < 0.05) SID of most AA compared with ESBM and fish meal. In experiment 2, 32 weanling barrows (14.0 ± 1.1 kg) were housed individually in metabolism crates and randomly allotted to one of four diets. A corn-based diet and three diets that contained corn and ESBM, fish meal, or cheese coproduct were formulated. Feces and urine were collected quantitatively. The ME in cheese coproduct was greater (P < 0.05) than in ESBM and fish meal. In experiment 3, 128 weaned pigs (6.2 ± 0.6 kg) were allotted to a randomized complete block design with four treatments and 8 replicate pens per diet. Phase 1 diets that contained 0%, 6.65%, 7.35%, or 14% cheese coproduct were fed from days 1 to 14 and a common phase 2 diet without cheese coproduct was fed from days 15 to 28. Individual pig weights were recorded at the beginning of the experiment, on days 14 and 28, and daily feed allotments were also recorded. Two blood samples were collected from 1 pig per pen on day 14 to analyze for blood urea N, albumin, total plasma protein, peptide YY, immunoglobulin G, tumor necrosis factor-α, interleukin-6, and interleukin-10. No differences were observed in average daily gain among treatments, but there was a tendency (P < 0.10) for total protein on day 14 to increase as cheese coproduct increased in the diets. In conclusion, the cheese coproduct used in this experiment has a greater SID of AA and greater ME than ESBM and fish meal and the cheese coproduct may be included in prestarter diets for weanling pigs without negatively impacting growth performance or indicators of intestinal health.

Milk proteins are highly digestible and have an excellent balance of indispensable amino acids (AA), but the price of dairy products are expensive compared with other protein sources. However, cheese that cannot be used for human consumption may be used in diets for pigs, but there is limited information about the nutritional value of cheese coproducts. Therefore, three experiments were conducted to determine the standardized ileal digestibility of AA, and the metabolizable energy (ME) of a cheese coproduct and effects of inclusion of different levels of cheese coproduct in phase 1 (1 to 14 d postweaning) diets for weanling pigs. Results demonstrated that the cheese coproduct has an excellent digestibility of AA and due to its greater ME than fish meal and enzyme-treated soybean meal, cheese coproduct can be used to increase the energy density of diets for weanling pigs without affecting the health or growth performance of pigs.

B-1 plasma cells require non-cognate CD4 T cell help to generate a unique repertoire of natural IgM.

Evolutionarily conserved, "natural" (n)IgM is broadly reactive to both self and foreign antigens. Its selective deficiency leads to increases in autoimmune diseases and infections. In mice, nIgM is secreted independent of microbial exposure to bone marrow (BM) and spleen B-1 cell-derived plasma cells (B-1PC), generating the majority of nIgM, or by B-1 cells that remain non-terminally differentiated (B-1sec). Thus, it has been assumed that the nIgM repertoire is broadly reflective of the repertoire of body cavity B-1 cells. Studies here reveal, however, that B-1PC generate a distinct, oligoclonal nIgM repertoire, characterized by short CDR3 variable immunoglobulin heavy chain regions, 7-8 amino acids in length, some public, many arising from convergent rearrangements, while specificities previously associated with nIgM were generated by a population of IgM-secreting B-1 (B-1sec). BM, but not spleen B-1PC, or B-1sec also required the presence of TCRαß CD4 T cells for their development from fetal precursors. Together, the studies identify important previously unknown characteristics of the nIgM pool.

Isolation and Characterization of Fetal Liver Hematopoietic Stem Cells.

Hematopoietic stem cells (HSCs) are responsible for the generation and maintenance of pools of multipotent precursors that ultimately give rise to all fully differentiated blood and immune cells. Proper identification and isolation of HSCs for functional analysis has greatly facilitated our understanding of both normal and abnormal adult hematopoiesis. Whereas adult hematopoiesis in mice and humans is driven by quiescent HSCs that reside almost exclusively within the bone marrow (BM), developmental hematopoiesis is characterized by a series of transient progenitors driving waves of increasingly mature hematopoietic cell production that occur across multiple anatomical sites. These waves of hematopoietic cell production are also responsible for the generation of distinct immune cell populations during development that persist into adulthood and contribute uniquely to adult immunity. Therefore, methods to properly isolate and characterize fetal progenitors with high purity across development become increasingly important not only for defining developmental hematopoietic pathways, but also for understanding the contribution of developmental hematopoiesis to the immune system. Here, we describe and discuss methods and considerations for the isolation and characterization of HSCs from the fetal liver, the primary hematopoietic organ during fetal development.

- Invited Review - Mineral composition and phosphorus digestibility in feed phosphates fed to pigs and poultry.

Phosphorus (P) is a macro mineral needed for bone mineralization and cell membrane structure and P is also involved in several fundamental pathways of metabolism in the body. Because of the low concentration and digestibility of P in plant ingredients that are the main components of diets for poultry and pigs, feed phosphates are usually included in diets in addition to the P contributed by plant ingredients. The most widely used feed phosphates in poultry and swine diets are dicalcium phosphate (DCP) and monocalcium phosphate (MCP), but tricalcium phosphate (TCP), monosodium phosphate (MSP), and magnesium phosphate (MgP) may be used as well. Because feed phosphates are mostly produced from rock phosphate, feed phosphates have impurities that contain minerals other than P. Concentrations of P in feed phosphates range from 14.8% (MgP) to 25.7% (MSP). The standardized total tract digestibility (STTD) of P in pigs ranges from 71% (TCP) to 95% (MSP). The STTD of Ca and the standardized ileal digestibility (SID) of P and Ca in feed phosphates fed to pigs and poultry have been determined only in a few experiments. Available data indicate that the STTD of Ca and SID of P in MCP are greater than in DCP in both poultry and pigs, but the SID of Ca is similar between DCP and MCP fed to broilers. Information on mineral concentrations and digestibility values in feed phosphates is needed in diet formulation for pigs and poultry, but if diets are formulated to contain equal concentrations of digestible P and Ca, it is unlikely that animal performance will be impacted by the source of feed phosphates used in the diet.

Prenatal inflammation perturbs murine fetal hematopoietic development and causes persistent changes to postnatal immunity.

Adult hematopoietic stem and progenitor cells (HSPCs) respond directly to inflammation and infection, causing both acute and persistent changes to quiescence, mobilization, and differentiation. Here we show that murine fetal HSPCs respond to prenatal inflammation in utero and that the fetal response shapes postnatal hematopoiesis and immune cell function. Heterogeneous fetal HSPCs show divergent responses to maternal immune activation (MIA), including changes in quiescence, expansion, and lineage-biased output. Single-cell transcriptomic analysis of fetal HSPCs in response to MIA reveals specific upregulation of inflammatory gene profiles in discrete, transient hematopoietic stem cell (HSC) populations that propagate expansion of lymphoid-biased progenitors. Beyond fetal development, MIA causes the inappropriate expansion and persistence of fetal lymphoid-biased progenitors postnatally, concomitant with increased cellularity and hyperresponsiveness of fetal-derived innate-like lymphocytes. Our investigation demonstrates how inflammation in utero can direct the output and function of fetal-derived immune cells by reshaping fetal HSC establishment.

Effects of microbial phytase on standardized total tract digestibility of phosphorus in feed phosphates fed to growing pigs.

An experiment was conducted to test the hypothesis that the apparent total tract digestibility (ATTD) and the standardized total tract digestibility (STTD) of P in feed phosphates are increased by microbial phytase when fed to growing pigs. Monocalcium phosphate (MCP), monosodium phosphate (MSP), and magnesium phosphate (MgP) from volcanic deposits were used in the experiment. Three corn-soybean meal based diets that contained 0, 500, or 4,000 units of microbial phytase (FTU), but no feed phosphates, were formulated. Nine additional diets were formulated by adding each of the three feed phosphates to the three basal diets. A P-free diet was also formulated to estimate the basal endogenous loss of P, and therefore, 13 diets were used in the experiment. A total of 117 growing barrows (initial body weight: 15.56 ±â€…1.68 kg) were allotted to the 13 diets with 9 pigs per diet. Pigs were housed individually in metabolism crates equipped with a feeder and a nipple drinker. Installation of a screen floor under the slatted floor allowed for collection of feces. Diets were fed for 10 d, with the initial 5 d being a period of adaptation to the diet followed by a collection period of 4 d. During the experiment, pigs were fed equal amounts of feed twice daily at 0800 and 1600 h. Results indicated that the ATTD and STTD of P in all diets increased with the inclusion of 500 or 4,000 FTU, but the ATTD and STTD of P in the feed phosphates were not affected by the inclusion of phytase. This indicates that the increases in ATTD and STTD of P that were observed in the mixed diets when phytase was used were due to the release of P from phytate in corn and soybean meal and not from an increase in digestibility of P in feed phosphates. However, MgP had a lower (P < 0.05) ATTD and STTD of P than MCP and MSP. In conclusion, microbial phytase does not increase the digestibility of P in MCP, MSP, or MGP, but the digestibility of P in MgP is less than in MCP and MSP.

Microbial phytase increases the digestibility by pigs of phytate-bound P in feed ingredients of plant origin, but digestible P can also be increased in diets by the addition of feed phosphates due to their high digestibility of P and lack of phytate. However, it is possible that the phytate from plant ingredients complexes with P from feed phosphates, resulting in a lower digestibility of P, but research to address this possibility has not been reported. Therefore, the hypothesis was that phytase can increase the digestibility of P in feed phosphates fed to pigs. Monocalcium phosphate (MCP), monosodium phosphate (MSP), and magnesium phosphate (MgP) were the three feed phosphates used in the experiment and the three ingredients were included in corn-soybean meal based diets. Results indicated that the inclusion of phytase increased the digestibility of P in the diets, but there was no indication that phytase affected the digestibility of P from any of the three feed phosphates, which indicates that the increase in digestibility of P likely was due to the release of P from plant ingredients in the diets. However, the digestibility of P was lower in MgP compared with MCP and MSP.

BiVO4 Photoanodes for TEMPO-Mediated Benzyl Alcohol Oxidation in Organic Media.

The photoelectrochemical production of fuels, exemplified by light-driven water splitting to hydrogen and oxygen, offers a sustainable option to offset dependence on fossil fuels. A low-cost, efficient, and stable photoelectrochemical approach to solar fuels remains elusive but using similar materials and photoelectrodes for chemical production or biomass conversion offers an appealing alternative. This work reports a facile method for fabricating pristine (undoped) BiVO4 photoanodes to carry out TEMPO-mediated benzyl alcohol oxidation to benzaldehyde in organic media (TEMPO=2,2,6,6-tetramethylpiperidinyl-N-oxyl). The best performing BiVO4 photoanode studied here gave a faradaic efficiency (FE) of 85±5% for benzaldehyde formation in the presence of TEMPO and pyridine during a 2.5-hour reaction. Compared with direct electrocatalytic conversion under the same conditions, light capture and conversion by the BiVO4 surface decreased the required applied bias by 46 %. To our knowledge, this is the first report of visible light assisted, TEMPO-mediated benzyl alcohol oxidation using pristine BiVO4 photoanodes in organic media.

Aortic intimal resident macrophages are essential for maintenance of the non-thrombogenic intravascular state.

Leukocytes and endothelial cells frequently cooperate to resolve inflammatory events. In most cases, these interactions are transient in nature and triggered by immunological insults. Here, we report that in areas of disturbed blood flow, aortic endothelial cells permanently and intimately associate with a population of specialized macrophages that are recruited at birth from the closing ductus arteriosus and share the luminal surface with the endothelium becoming interwoven in the tunica intima. Anatomical changes that affect hemodynamics, like in patent ductus arteriosus, alter macrophage seeding to coincide with regions of disturbed flow. Aortic resident macrophages expand in situ via direct cell renewal. Induced-depletion of intimal macrophages led to thrombin-mediated endothelial cell contraction, progressive fibrin accumulation and formation of microthrombi that, once dislodged, caused blockade of vessels in several organs. Together the findings revealed that intravascular resident macrophages are essential to regulate thrombin activity and clear fibrin deposits in regions of disturbed blood flow.

A "Switch" in Time through Genes Aligned: Unraveling the Genomic Landscape of HSC Development.

Seeking to define the "switch" from fetal to adult hematopoiesis, Li et al. (2020) performed extensive genomic and epigenomic profiling of hematopoietic stem and progenitor cells across ontogeny (as explored in this issue of Cell Stem Cell). Gradual and stochastic changes in genomic and epigenomic regulation suggest the absence of any specific regulator.

Evaluation of the Edman degradation product of vancomycin bonded to core-shell particles as a new HPLC chiral stationary phase.

A modified macrocyclic glycopeptide-based chiral stationary phase (CSP), prepared via Edman degradation of vancomycin, was evaluated as a chiral selector for the first time. Its applicability was compared with other macrocyclic glycopeptide-based CSPs: TeicoShell and VancoShell. In addition, another modified macrocyclic glycopeptide-based CSP, NicoShell, was further examined. Initial evaluation was focused on the complementary behavior with these glycopeptides. A screening procedure was used based on previous work for the enantiomeric separation of 50 chiral compounds including amino acids, pesticides, stimulants, and a variety of pharmaceuticals. Fast and efficient chiral separations resulted by using superficially porous (core-shell) particle supports. Overall, the vancomycin Edman degradation product (EDP) resembled TeicoShell with high enantioselectivity for acidic compounds in the polar ionic mode. The simultaneous enantiomeric separation of 5 racemic profens using liquid chromatography-mass spectrometry with EDP was performed in approximately 3 minutes. Other highlights include simultaneous liquid chromatography separations of rac-amphetamine and rac-methamphetamine with VancoShell, rac-pseudoephedrine and rac-ephedrine with NicoShell, and rac-dichlorprop and rac-haloxyfop with TeicoShell.

Effective methodologies for enantiomeric separations of 150 pharmacology and toxicology related 1°, 2°, and 3° amines with core-shell chiral stationary phases.

Core-shell particles (superficially porous particles, SPPs) have been proven to provide high-throughput and effective separations of a variety of chiral molecules. However, due to their limited commercialization, many separations have not been reported with these stationary phases. In this study, four SPP chiral stationary phases (CSPs) were utilized for the enantiomeric separation of 150 chiral amines. These amines encompass a variety of structural and drug classes, which are particularly important to the pharmaceutical industry and in forensics. This comprehensive evaluation demonstrates the power of these CSPs and the ease of method development and optimization. The CSPs used in this study included the macrocyclic glycopeptide-based CSPs (VancoShell and NicoShell), the cyclodextrin-based CSP (CDShell-RSP), and the cyclofructan-based CSP (LarihcShell-P). These CSPs offered versatility for a variety of applications and worked in a complementary fashion to baseline separate all 150 amines. The LarihcShell-P was highly effective for separating primary amines. VancoShell, NicoShell, and CDShell-RSP were useful for separating all types of amines. These CSPs are multi-modal and can be utilized with mass spectrometry compatible solvents. Eighteen racemic controlled substances were simultaneously baseline separated in a single liquid chromatography-mass spectrometry (LC-MS) analysis. Details in high-performance liquid chromatography (HPLC) parameters will be discussed as well as the improved chromatographic performance afforded by the SPP CSPs.

A comprehensive methodology for the chiral separation of 40 tobacco alkaloids and their carcinogenic E/Z-(R,S)-tobacco-specific nitrosamine metabolites.

The predominant enantiomer of nicotine found in nature is (S)-nicotine and its pharmacology has been widely established. However, pharmacologic information concerning individual enantiomers of nicotine-related compounds is limited. Recently, a modified macrocyclic glycopeptide chiral selector was found to be highly stereoselective for most tobacco alkaloids and metabolites. This study examines the semi-synthetic and native known macrocyclic glycopeptides for chiral recognition, separation, and characterization of the largest group of nicotine-related compounds ever reported (tobacco alkaloids, nicotine metabolites and derivatives, and tobacco-specific nitrosamines). The enantioseparation of nicotine is accomplished in less than 20s for example. All liquid chromatography separations are mass spectrometry compatible for the tobacco alkaloids, as well as their metabolites. Ring-closed, cyclized structures were identified and separated from their ring-open, straight chain equilibrium structures. Also, E/Z-tobacco-specific nitrosamines and their enantiomers were directly separated. E/Z isomers also are known to have different physical and chemical properties and biological activities. This study provides optimal separation conditions for the analysis of nicotine-related isomers, which in the past have been reported to be ineffectively separated which can result in inaccurate results. The methodology of this study could be applied to cancer studies, and lead to more information about the role of these isomers in other diseases and as treatment for diseases.

Macrocyclic glycopeptide chiral selectors bonded to core-shell particles enables enantiopurity analysis of the entire verubecestat synthetic route.

Verubecestat is an inhibitor of ß-site amyloid precursor protein cleaving enzyme 1 (BACE1) being evaluated in clinical trials for the treatment of Alzheimer's disease. Synthetic route development involves diastereoselective transformations with a need for enantiomeric excess (ee) determination of each intermediate and final active pharmaceutical ingredient (API). The analytical technical package of validated methods relies on enantioselective SFC and RPLC separations using multiple 3 and 5 µm coated polysaccharide-based chiral stationary phases (CSPs) and mobile phases combinations. Evaluation of recently developed chiral columns revealed a single chiral selector (Teicoplanin) bonded to 2.7 µm core-shell particles using H3PO4 in H2O/ACN and triethylammonium acetate: methanol based eluents at different isocratic compositions allowed good enatioseparation of all verubecestat intermediates. EE determination of verubecestat is easily performed on NicoShell, another macrocyclic glycopeptide chiral selector bonded to 2.7 µm superficially porous particles. This approach enables fast and reliable enantiopurity analysis of the entire verubecestat synthetic route using only two chiral columns and mobile phases on a conventional HPLC system, simplifying technical package preparation, method validation and transfer to manufacturing facilities.

Evaluation of nicotine in tobacco-free-nicotine commercial products.

Recently, a variety of new tobacco-free-nicotine, TFN, products have been commercialized as e-liquids. Tobacco-derived nicotine contains predominantly (S)-(-)-nicotine, whereas TFN products may not. The TFN products are said to be cleaner, purer substances, devoid of toxic components that come from the tobacco extraction process. A variety of commercial tobacco and TFN products were analyzed to identify the presence and composition of each nicotine enantiomer. A rapid and effective enantiomeric separation of nicotine has been developed using a modified macrocyclic glycopeptide bonded to superficially porous particles. The enantiomeric assay can be completed in <2 min with high resolution and accuracy using high performance liquid chromatography with electrospray ionization mass spectrometry. The results of this study suggest the need for pharmacological studies of (R)-(+)-nicotine, which is present in much greater quantities in commercial TFN products compared to commercial tobacco-derived products. Such studies are required by the FDA for new enantiomeric pharmacological products. Copyright © 2016 John Wiley & Sons, Ltd.