Effects of Feeding Selenium-Enriched Alfalfa Hay on Immunity and Health of Weaned Beef Calves

Previously, we reported that feeding selenium (Se)-enriched forage improves antibody titers in mature beef cows, and whole-blood Se concentrations and growth rates in weaned beef calves. Our current objective was to test whether beef calves fed Se-enriched alfalfa hay during the transition period between weaning and movement to a feedlot also have improved immune responses and slaughter weights. Recently weaned beef calves (n?=?60) were fed an alfalfa-hay-based diet for 7 weeks, which was harvested from fields fertilized with sodium selenate at 0, 22.5, 45.0, or 89.9 g Se/ha. All calves were immunized with J-5 Escherichia coli bacterin. Serum was collected for antibody titers 2 weeks after the third immunization. Whole-blood neutrophils collected at 6 or 7 weeks were evaluated for total antioxidant potential, bacterial killing activity, and expression of genes associated with selenoproteins and innate immunity. Calves fed the highest versus the lowest level of Se-enriched alfalfa hay had higher antibody titers (P?=?0.02), thioredoxin reductase-2 mRNA levels (P?=?0.07), and a greater neutrophil total antioxidant potential (P?=?0.10), whereas mRNA levels of interleukin-8 receptor (P?=?0.02), l-selectin (P?=?0.07), and thioredoxin reductase-1 (P?=?0.07) were lower. In the feedlot, calves previously fed the highest-Se forage had lower mortality (P?=?0.04) and greater slaughter weights (P?=?0.02). Our results suggest that, in areas with low-forage Se concentrations, feeding beef calves Se-enriched alfalfa hay during the weaning transition period improves vaccination responses and subsequent growth and survival in the feedlot.     

Effect of Feeding Selenium-Fertilized Alfalfa Hay on Performance of Weaned Beef Calves

Selenium (Se) is an essential micronutrient in cattle, and Se-deficiency can affect morbidity and mortality. Calves may have greater Se requirements during periods of stress, such as during the transitional period between weaning and movement to a feedlot. Previously, we showed that feeding Se-fertilized forage increases whole-blood (WB) Se concentrations in mature beef cows. Our current objective was to test whether feeding Se-fertilized forage increases WB-Se concentrations and performance in weaned beef calves. Recently weaned beef calves (n = 60) were blocked by body weight, randomly assigned to 4 groups, and fed an alfalfa hay based diet for 7 wk, which was harvested from fields fertilized with sodium-selenate at a rate of 0, 22.5, 45.0, or 89.9 g Se/ha. Blood samples were collected weekly and analyzed for WB-Se concentrations. Body weight and health status of calves were monitored during the 7-wk feeding trial. Increasing application rates of Se fertilizer resulted in increased alfalfa hay Se content for that cutting of alfalfa (0.07, 0.95, 1.55, 3.26 mg Se/kg dry matter for Se application rates of 0, 22.5, 45.0, or 89.9 g Se/ha, respectively). Feeding Se-fertilized alfalfa hay during the 7-wk preconditioning period increased WB-Se concentrations (P _Linear<0.001) and body weights (P _Linear = 0.002) depending upon the Se-application rate. Based upon our results we suggest that soil-Se fertilization is a potential management tool to improve Se-status and performance in weaned calves in areas with low soil-Se concentrations.     

Scalable economic extracellular synthesis of CdS nanostructured particles by a non-pathogenic thermophile

We report microbially facilitated synthesis of cadmium sulfide (CdS) nanostructured particles (NP) using anaerobic, metal-reducing Thermoanaerobacter sp. The extracellular CdS crystallites were <10 nm in size with yields of ~3 g/L of growth medium/month with demonstrated reproducibility and scalability up to 24 L. During synthesis, Thermoanaerobacter cultures reduced thiosulfate and sulfite salts to H₂S, which reacted with Cd²⁺ cations to produce thermodynamically favored NP in a single step at 65 °C with catalytic nucleation on the cell surfaces. Photoluminescence (PL) analysis of dry CdS NP revealed an exciton-dominated PL peak at 440 nm, having a narrow full width at half maximum of 10 nm. A PL spectrum of CdS NP produced by dissimilatory sulfur reducing bacteria was dominated by features associated with radiative exciton relaxation at the surface. High reproducibility of CdS NP PL features important for scale-up conditions was confirmed from test tubes to 24 L batches at a small fraction of the manufacturing cost associated with conventional inorganic NP production processes.     

Regulation of integrin function by CD47 ligands. Differential effects on alpha vbeta 3 and alpha 4beta1 integrin-mediated adhesion

We examined the regulation of alpha4beta1 integrin function in melanoma cells and T cells by ligands of CD47. A CD47 antibody (B6H12) that inhibited alphavbeta3-mediated adhesion of melanoma cells induced by CD47-binding peptides from thrombospondin-1 directly stimulated alpha4beta1-mediated adhesion of the same cells to vascular cell adhesion molecule-1 and N-terminal regions of thrombospondin-1 or thrombospondin-2. B6H12 also stimulated alpha4beta1- as well as alpha2beta1- and alpha5beta1-mediated adhesion of CD47-expressing T cells but not of CD47-deficient T cells. alpha4beta1 and CD47 co-purified as a detergent-stable complex on a CD47 antibody affinity column. CD47-binding peptides based on C-terminal sequences of thrombospondin-1 also specifically enhanced adhesion of melanoma cells and T cells to alpha4beta1 ligands. Unexpectedly, activation of alpha4beta1 function by the thrombospondin-1 CD47-binding peptides also occurred in CD47-deficient T cells. CD47-independent activation of alpha4beta1 required the Val-Val-Met (VVM) motif of the peptides and was sensitive to inhibition by pertussis toxin. These results indicate that activation of alpha4beta1 by the CD47 antibody B6H12 and by VVM peptides occurs by different mechanisms. The antibody directly activates a CD47-alpha4beta1 complex, whereas VVM peptides may target an unidentified Gi-linked receptor that regulates alpha4beta1.     

The cytoplasmic tyrosines of integrin subunit beta1 are involved in focal adhesion kinase activation

We have previously shown that mutation of the two tyrosines in the cytoplasmic domain of integrin subunit beta1 (Y783 and Y795) to phenylalanines markedly reduces the capability of beta1A integrins to mediate directed cell migration. In this study, beta1-dependent cell spreading was found to be delayed in GD25 cells expressing beta1A(Y783/795F) compared to that in wild-type GD25-beta1A. Focal adhesion kinase (FAK) tyrosine phosphorylation and activation were severely impaired in response to beta1-dependent adhesion in GD25-beta1A(Y783/795F) cells compared to that in wild-type GD25-beta1A or mutants in which only a single tyrosine was altered (beta1A(Y783F) or beta1A(Y795F)). Phosphorylation site-specific antibodies selective for FAK phosphotyrosine 397 indicated that the defect in FAK phosphorylation via beta1A(Y783/795F) lies at the level of the initial autophosphorylation step. Indeed, beta1A-dependent tyrosine phosphorylation of tensin and paxillin was lost in the beta1A(Y783/795F) cells, consistent with the impairment in FAK activation. In contrast, p130(CAS) overall tyrosine phosphorylation was unaffected by the beta1 mutations. Despite the defect in beta1-mediated FAK activation, FAK was still localized to focal adhesions. Taken together, the phenotype of the GD25-beta1A(Y783/795F) cells resembles, but is distinct from, the phenotype observed in FAK-null cells. These observations argue that tyrosines 783 and 795 within the cytoplasmic tail of integrin subunit beta1A are critical mediators of FAK activation and cell spreading in GD25 cells.     

Specific interactions between F1 adhesin of Streptococcus pyogenes and N-terminal modules of fibronectin

Protein F1 is a surface protein of Streptococcus pyogenes that mediates high affinity binding to fibronectin (Fn) and facilitates S. pyogenes adherence and penetration into cells. The smallest portion of F1 known to retain the full binding potential of the intact protein is a stretch of 49 amino acids known as the functional upstream domain (FUD). Synthetic and recombinant versions of FUD were labeled with fluorescein isothiocyanate and used in fluorescence anisotropy experiments. These probes bound to Fn or the 70-kDa fragment of Fn with dissociation constants of 8-30 nm. Removal of the N-terminal seven residues of FUD did not cause a change in binding affinity. Further N- or C-terminal truncations resulted in complete loss of binding activity. Analysis of recombinant versions of the 70-kDa fragment that lacked one or several type I modules indicates that residues 1-7 of the 49-mer bind to type I modules I1 and I2 of the 27-kDa subfragment and the C-terminal residues bind to modules I4 and I5. Fluorescein isothiocyanate-labeled 49-mer also bound with lower affinity to large Fn fragments that lack the five type I modules of the 27-kDa fragment but contain the other seven type 1 modules of Fn. These results indicate that, although FUD has a general affinity for type I modules, high affinity binding of FUD to Fn is mediated by specific interactions with N-terminal type I modules.     

Absence of Vitamin K-Dependent γ-Carboxylation in Human Periostin Extracted from Fibrotic Lung or Secreted from a Cell Line Engineered to Optimize γ-Carboxylation

Periostin (PN, gene name POSTN) is an extracellular matrix protein that is up-regulated in bronchial epithelial cells and lung fibroblasts by TH-2 cytokines. Its paralog, TGF-β-induced protein (βig-h3, gene name TGFBI), is also expressed in the lung and up-regulated in bronchial myofibroblasts by TGF-β. PN and βig-h3 contain fasciclin 1 modules that harbor putative recognition sequences for γ-glutamyl carboxylase and are annotated in UniProt as undergoing vitamin K-dependent γ-carboxylation of multiple glutamic acid residues. γ-carboxylation profoundly alters activities of other proteins subject to the modification, e.g., blood coagulation factors, and would be expected to alter the structure and function of PN and βig-h3. To analyze for the presence of γ-carboxylation, proteins extracted from fibrotic lung were reacted with monoclonal antibodies specific for PN, βig-h3, or modification with γ-carboxyglutamic acid (Gla). In Western blots of 1-dimensional gels, bands stained with anti-PN or -βig-h3 did not match those stained with anti-Gla. In 2-dimensional gels, anti-PN-positive spots had pIs of 7.0 to >8, as expected for the unmodified protein, and there was no overlap between anti-PN-positive and anti-Gla-positive spots. Recombinant PN and blood coagulation factor VII were produced in HEK293 cells that had been transfected with vitamin K 2, 3-epoxide reductase C1 to optimize γ-carboxylation. Recombinant PN secreted from these cells did not react with anti-Gla antibody and had pIs similar to that found in extracts of fibrotic lung whereas secreted factor VII reacted strongly with anti-Gla antibody. Over 67% coverage of recombinant PN was achieved by mass spectrometry, including peptides with 19 of the 24 glutamates considered targets of γ-carboxylation, but analysis revealed no modification. Over 86% sequence coverage and three modified glutamic acid residues were identified in recombinant fVII. These data indicate that PN and βig-h3 are not subject to vitamin K-dependent γ-carboxylation.     

Integrated Healthcare Delivery: A Qualitative Research Approach to Identifying and Harmonizing Perspectives of Integrated Neglected Tropical Disease Programs

BackgroundWhile some evidence supports the beneficial effects of integrating neglected tropical disease (NTD) programs to optimize coverage and reduce costs, there is minimal information regarding when or how to effectively operationalize program integration. The lack of systematic analyses of integration experiences and of integration processes may act as an impediment to achieving more effective NTD programming. We aimed to learn about the experiences of NTD stakeholders and their perceptions of integration.MethodologyWe evaluated differences in the definitions, roles, perceived effectiveness, and implementation experiences of integrated NTD programs among a variety of NTD stakeholder groups, including multilateral organizations, funding partners, implementation partners, national Ministry of Health (MOH) teams, district MOH teams, volunteer rural health workers, and community members participating in NTD campaigns. Semi-structured key informant interviews were conducted. Coding of themes involved a mix of applying in-vivo open coding and a priori thematic coding from a start list.FindingsIn total, 41 interviews were conducted. Salient themes varied by stakeholder, however dominant themes on integration included: significant variations in definitions, differential effectiveness of specific integrated NTD activities, community member perceptions of NTD programs, the influence of funders, perceived facilitators, perceived barriers, and the effects of integration on health system strength. In general, stakeholder groups provided unique perspectives, rather than contrarian points of view, on the same topics. The stakeholders identified more advantages to integration than disadvantages, however there are a number of both unique facilitators and challenges to integration from the perspective of each stakeholder group.ConclusionsQualitative data suggest several structural, process, and technical opportunities that could be addressed to promote more effective and efficient integrated NTD elimination programs. We highlight a set of ten recommendations that may address stakeholder concerns and perceptions regarding these key opportunities. For example, public health stakeholders should embrace a broader perspective of community-based health needs, including and beyond NTDs, and available platforms for addressing those needs.