Altered serological and cellular reactivity to H-2 antigens after target cell infection with vaccinia virus

MICE generate cytotoxic T lymphocytes (CTL) which are able to lyse virus infected target cells in vitro after infection with lymphocytic choriomeningitis virus (LCMV) and pox-viruses1−3. CTL kill syngeneic and semiallogenic infected cells but not allogenic infected targets. Target cell lysis in these systems seems to be restricted by H-2 antigens, especially by the K or D end of the major histocompatibility complex (MHC). In experiments where virus specific sensitised lymphocytes kill virus infected allogenic target cells4 the effector lymphocytes have not been characterised exactly. Recent investigations suggest that the active cell in this assay, at least in the measles infection, is a non-thymus derived cell (H. Kreth, personal communication). An H-2 restriction of cell mediated cytolysis (CMC) to trinitrophenol (TNP)-modified lymphocytes has also been described5. Zinkernagel and Doherty6 postulated that the CTL is directed against syngeneic H-2 antigens and viral antigens and they suggested an alteration of H-2 induced by the LCMV infection. Earlier7 we found a close topological relationship between H-2 antigens and the target antigen(s) responsible for CMC in the vaccinia system. Here we report experiments which were carried out to prove alteration of H-2 after infection of L-929 fibroblasts with vaccinia virus

Elevated Fibroblast growth factor 21 (FGF21) in obese, insulin resistant states is normalised by the synthetic retinoid Fenretinide in mice

The authors would like to thank undergraduate student Aleksandra Kowalczuk (University of Aberdeen) for assisting in experiments and Dr. Emma K. Lees (School of Health Sciences, Liverpool Hope University, Liverpool, UK) for invaluable discussions concerning the regulation of FGF21. We thank Dr. Calum Sutherland and Dr. Amy Cameron (both Division of Molecular and Clinical Medicine, Ninewells Hospital and Medical School, University of Dundee, Scotland, UK) for technical support and expertise in performing hepatocyte studies. Fenretinide was a generous gift of T. Martin (Johnson & Johnson, New Brunswick, NJ) and U. Thumeer (Cilag AG, Schaffhausen, Switzerland), for use completely without restriction or obligation. Quantitative-PCR was carried out using the qPCR Core Facility (Institute of Medical Sciences, University of Aberdeen). RNA-sequencing was carried out at the University of Aberdeen Centre for Genome Enabled Biology and Medicine. Pancreas histology was performed by Dr Linda Davidson (Department of Histology, Aberdeen Royal Infirmary, NHS Grampian, Foresterhill Health Campus, Aberdeen, UK). This study was supported by the British Heart Foundation Intermediate Basic Research Fellowship FS/09/026 to N. Mody, RCUK fellowship to MD, EFSD/Lilly Programme Grant to MD and N. Mody, Tenovus Scotland grants G10/04 and G14/14 to N. Mody, University of Aberdeen Centre for Genome Enabled Biology and Medicine (CGEBM) PhD studentship to N. Morrice and Biotechnology and Biological Sciences Research Council studentship to GDM.Peer reviewedPublisher PD

Influence of sex, age, pubertal maturation and body mass index on circulating white blood cell counts in healthy European adolescents—the HELENA study

Percentiles 10th, 25th, 50th, 75th and 90th are presented for circulating white blood cells (WBC), neutrophils, lymphocytes, monocytes, eosinophils and basophils in healthy European adolescents (12.5–17.5 years, n = 405, 48.9 % boys), considering age, sex, puberty and body mass index (BMI). CD3+ (mature T cells), CD4+ (T helper), CD8+ (T cytotoxic), CD16+56+ (natural killer), CD19+ (B cells), CD3+CD45RA+, CD4+CD45RA+, CD8+CD45RA+ (naïve), CD3+CD45RO+, CD4+CD45RO+ and CD8+CD45RO+ (memory) lymphocytes were also analysed by immunophenotyping. Girls presented higher WBC, neutrophil, CD3+CD45RO+ and CD4+CD45RO+ cell counts and CD3+/CD19+ ratio, and lower CD3+CD45RA+ and CD4+CD45RA+ counts than boys. Age was associated with higher neutrophil counts and CD3+/CD19+, and lower CD19+ counts; in boys, with lower CD3+CD45RA+, CD4+CD45RA+ and CD8+CD45RA+ counts as well; in girls, with higher WBC, CD3+CD45RO+ and CD4+CD45RO+ counts. Pubertal maturation in boys was associated with lower WBC and lymphocyte counts; in girls, with higher basophil, CD3+CD45RO+ and CD4+CD45RO+ values. BMI was associated with higher WBC counts; in boys, also with higher lymphocyte counts; in girls, with higher neutrophil, CD4+, CD3+CD45RO+ and CD4+CD45RO+ counts. Conclusion: Our study provides normative values for circulating immune cells in adolescents, highlighting the importance of considering sex, age, pubertal maturation and BMI when establishing reference ranges for WBC in paediatric populations

U.S. Physicians’ Views on Financing Options to Expand Health Insurance Coverage: A National Survey

Background: Physician opinion can influence the prospects for health care reform, yet there are few recent data on physician views on reform proposals or access to medical care in the United States. Objective: To assess physician views on financing options for expanding health care coverage and on access to health care. Design and Participants: Nationally representative mail survey conducted between March 2007 and October 2007 of U.S. physicians engaged in direct patient care. Measurements: Rated support for reform options including financial incentives to induce individuals to purchase health insurance and single-payer national health insurance; rated views of several dimensions of access to care. Main results: 1,675 of 3,300 physicians responded (50.8%). Only 9% of physicians preferred the current employer-based financing system. Forty-nine percent favored either tax incentives or penalties to encourage the purchase of medical insurance, and 42% preferred a government-run, taxpayer-financed single-payer national health insurance program. The majority of respondents believed that all Americans should receive needed medical care regardless of ability to pay (89%); 33% believed that the uninsured currently have access to needed care. Nearly one fifth of respondents (19.3%) believed that even the insured lack access to needed care. Views about access were independently associated with support for single-payer national health insurance. Conclusions: The vast majority of physicians surveyed supported a change in the health care financing system. While a plurality support the use of financial incentives, a substantial proportion support single payer national health insurance. These findings challenge the perception that fundamental restructuring of the U.S. health care financing system receives little acceptance by physicians

Metabonomic fingerprints of fasting plasma and spot urine reveal human pre-diabetic metabolic traits

Impaired glucose tolerance (IGT) which precedes overt type 2 diabetes (T2DM) for decades is associated with multiple metabolic alterations in insulin sensitive tissues. In an UPLC-qTOF-mass spectrometry-driven non-targeted metabonomics approach we investigated plasma as well as spot urine of 51 non-diabetic, overnight fasted individuals aiming to separate subjects with IGT from controls thereby identify pathways affected by the pre-diabetic metabolic state. We could clearly demonstrate that normal glucose tolerant (NGT) and IGT subjects clustered in two distinct groups independent of the investigated metabonome. These findings reflect considerable differences in individual metabolite fingerprints, both in plasma and urine. Pre-diabetes associated alterations in fatty acid-, tryptophan-, uric acid-, bile acid-, and lysophosphatidylcholine-metabolism, as well as the TCA cycle were identified. Of note, individuals with IGT also showed decreased levels of gut flora-associated metabolites namely hippuric acid, methylxanthine, methyluric acid, and 3-hydroxyhippuric acid. The findings of our non-targeted UPLC-qTOF-MS metabonomics analysis in plasma and spot urine of individuals with IGT vs NGT offers novel insights into the metabolic alterations occurring in the long, asymptomatic period preceding the manifestation of T2DM thereby giving prospects for new intervention targets

Neuronal Nitric Oxide Synthase-Rescue of Dystrophin/Utrophin Double Knockout Mice does not Require nNOS Localization to the Cell Membrane

Survival of dystrophin/utrophin double-knockout (dko) mice was increased by muscle-specific expression of a neuronal nitric oxide synthase (nNOS) transgene. Dko mice expressing the transgene (nNOS TG+/dko) experienced delayed onset of mortality and increased life-span. The nNOS TG+/dko mice demonstrated a significant decrease in the concentration of CD163+, M2c macrophages that can express arginase and promote fibrosis. The decrease in M2c macrophages was associated with a significant reduction in fibrosis of heart, diaphragm and hindlimb muscles of nNOS TG+/dko mice. The nNOS transgene had no effect on the concentration of cytolytic, CD68+, M1 macrophages. Accordingly, we did not observe any change in the extent of muscle fiber lysis in the nNOS TG+/dko mice. These findings show that nNOS/NO (nitric oxide)-mediated decreases in M2c macrophages lead to a reduction in the muscle fibrosis that is associated with increased mortality in mice lacking dystrophin and utrophin. Interestingly, the dramatic and beneficial effects of the nNOS transgene were not attributable to localization of nNOS protein at the cell membrane. We did not detect any nNOS protein at the sarcolemma in nNOS TG+/dko muscles. This important observation shows that sarcolemmal localization is not necessary for nNOS to have beneficial effects in dystrophic tissue and the presence of nNOS in the cytosol of dystrophic muscle fibers can ameliorate the pathology and most importantly, significantly increase life-span

Arginine Metabolism by Macrophages Promotes Cardiac and Muscle Fibrosis in mdx Muscular Dystrophy

Duchenne muscular dystrophy (DMD) is the most common, lethal disease of childhood. One of 3500 new-born males suffers from this universally-lethal disease. Other than the use of corticosteroids, little is available to affect the relentless progress of the disease, leading many families to use dietary supplements in hopes of reducing the progression or severity of muscle wasting. Arginine is commonly used as a dietary supplement and its use has been reported to have beneficial effects following short-term administration to mdx mice, a genetic model of DMD. However, the long-term effects of arginine supplementation are unknown. This lack of knowledge about the long-term effects of increased arginine metabolism is important because elevated arginine metabolism can increase tissue fibrosis, and increased fibrosis of skeletal muscles and the heart is an important and potentially life-threatening feature of DMD.We use both genetic and nutritional manipulations to test whether changes in arginase metabolism promote fibrosis and increase pathology in mdx mice. Our findings show that fibrotic lesions in mdx muscle are enriched with arginase-2-expressing macrophages and that muscle macrophages stimulated with cytokines that activate the M2 phenotype show elevated arginase activity and expression. We generated a line of arginase-2-null mutant mdx mice and found that the mutation reduced fibrosis in muscles of 18-month-old mdx mice, and reduced kyphosis that is attributable to muscle fibrosis. We also observed that dietary supplementation with arginine for 17-months increased mdx muscle fibrosis. In contrast, arginine-2 mutation did not reduce cardiac fibrosis or affect cardiac function assessed by echocardiography, although 17-months of dietary supplementation with arginine increased cardiac fibrosis. Long-term arginine treatments did not decrease matrix metalloproteinase-2 or -9 or increase the expression of utrophin, which have been reported as beneficial effects of short-term treatments.Our findings demonstrate that arginine metabolism by arginase promotes fibrosis of muscle in muscular dystrophy and contributes to kyphosis. Our findings also show that long-term, dietary supplementation with arginine exacerbates fibrosis of dystrophic heart and muscles. Thus, commonly-practiced dietary supplementation with arginine by DMD patients has potential risk for increasing pathology when performed for long periods, despite reports of benefits acquired with short-term supplementation

Relevance of laboratory testing for the diagnosis of primary immunodeficiencies: a review of case-based examples of selected immunodeficiencies

The field of primary immunodeficiencies (PIDs) is one of several in the area of clinical immunology that has not been static, but rather has shown exponential growth due to enhanced physician, scientist and patient education and awareness, leading to identification of new diseases, new molecular diagnoses of existing clinical phenotypes, broadening of the spectrum of clinical and phenotypic presentations associated with a single or related gene defects, increased bioinformatics resources, and utilization of advanced diagnostic technology and methodology for disease diagnosis and management resulting in improved outcomes and survival. There are currently over 200 PIDs with at least 170 associated genetic defects identified, with several of these being reported in recent years. The enormous clinical and immunological heterogeneity in the PIDs makes diagnosis challenging, but there is no doubt that early and accurate diagnosis facilitates prompt intervention leading to decreased morbidity and mortality. Diagnosis of PIDs often requires correlation of data obtained from clinical and radiological findings with laboratory immunological analyses and genetic testing. The field of laboratory diagnostic immunology is also rapidly burgeoning, both in terms of novel technologies and applications, and knowledge of human immunology. Over the years, the classification of PIDs has been primarily based on the immunological defect(s) ("immunophenotype") with the relatively recent addition of genotype, though there are clinical classifications as well. There can be substantial overlap in terms of the broad immunophenotype and clinical features between PIDs, and therefore, it is relevant to refine, at a cellular and molecular level, unique immunological defects that allow for a specific and accurate diagnosis. The diagnostic testing armamentarium for PID includes flow cytometry - phenotyping and functional, cellular and molecular assays, protein analysis, and mutation identification by gene sequencing. The complexity and diversity of the laboratory diagnosis of PIDs necessitates many of the above-mentioned tests being performed in highly specialized reference laboratories. Despite these restrictions, there remains an urgent need for improved standardization and optimization of phenotypic and functional flow cytometry and protein-specific assays. A key component in the interpretation of immunological assays is the comparison of patient data to that obtained in a statistically-robust manner from age and gender-matched healthy donors. This review highlights a few of the laboratory assays available for the diagnostic work-up of broad categories of PIDs, based on immunophenotyping, followed by examples of disease-specific testing

Stable Isotope Biogeochemistry of Seabird Guano Fertilization: Results from Growth Chamber Studies with Maize (Zea Mays)

Stable isotope analysis is being utilized with increasing regularity to examine a wide range of issues (diet, habitat use, migration) in ecology, geology, archaeology, and related disciplines. A crucial component to these studies is a thorough understanding of the range and causes of baseline isotopic variation, which is relatively poorly understood for nitrogen (δ(15)N). Animal excrement is known to impact plant δ(15)N values, but the effects of seabird guano have not been systematically studied from an agricultural or horticultural standpoint.This paper presents isotopic (δ(13)C and δ(15)N) and vital data for maize (Zea mays) fertilized with Peruvian seabird guano under controlled conditions. The level of (15)N enrichment in fertilized plants is very large, with δ(15)N values ranging between 25.5 and 44.7‰ depending on the tissue and amount of fertilizer applied; comparatively, control plant δ(15)N values ranged between -0.3 and 5.7‰. Intraplant and temporal variability in δ(15)N values were large, particularly for the guano-fertilized plants, which can be attributed to changes in the availability of guano-derived N over time, and the reliance of stored vs. absorbed N. Plant δ(13)C values were not significantly impacted by guano fertilization. High concentrations of seabird guano inhibited maize germination and maize growth. Moreover, high levels of seabird guano greatly impacted the N metabolism of the plants, resulting in significantly higher tissue N content, particularly in the stalk.The results presented in this study demonstrate the very large impact of seabird guano on maize δ(15)N values. The use of seabird guano as a fertilizer can thus be traced using stable isotope analysis in food chemistry applications (certification of organic inputs). Furthermore, the fertilization of maize with seabird guano creates an isotopic signature very similar to a high-trophic level marine resource, which must be considered when interpreting isotopic data from archaeological material

Ebola: translational science considerations

We are currently in the midst of the most aggressive and fulminating outbreak of Ebola-related disease, commonly referred to as “Ebola”, ever recorded. In less than a year, the Ebola virus (EBOV, Zaire ebolavirus species) has infected over 10,000 people, indiscriminately of gender or age, with a fatality rate of about 50%. Whereas at its onset this Ebola outbreak was limited to three countries in West Africa (Guinea, where it was first reported in late March 2014, Liberia, where it has been most rampant in its capital city, Monrovia and other metropolitan cities, and Sierra Leone), cases were later reported in Nigeria, Mali and Senegal, as well as in Western Europe (i.e., Madrid, Spain) and the US (i.e., Dallas, Texas; New York City) by late October 2014. World and US health agencies declared that the current Ebola virus disease (EVD) outbreak has a strong likelihood of growing exponentially across the world before an effective vaccine, treatment or cure can be developed, tested, validated and distributed widely. In the meantime, the spread of the disease may rapidly evolve from an epidemics to a full-blown pandemic. The scientific and healthcare communities actively research and define an emerging kaleidoscope of knowledge about critical translational research parameters, including the virology of EBOV, the molecular biomarkers of the pathological manifestations of EVD, putative central nervous system involvement in EVD, and the cellular immune surveillance to EBOV, patient-centered anthropological and societal parameters of EVD, as well as translational effectiveness about novel putative patient-targeted vaccine and pharmaceutical interventions, which hold strong promise, if not hope, to curb this and future Ebola outbreaks. This work reviews and discusses the principal known facts about EBOV and EVD, and certain among the most interesting ongoing or future avenues of research in the field, including vaccination programs for the wild animal vectors of the virus and the disease from global translational science perspective