Mitogen induced cellular cytotoxic activity as a monocyte function in human peripheral blood preparations

The effector cell population in man responsible for mitogen induced cellular cytotoxicity (MICC) of chicken erythrocytes was investigated using several separation techniques, including free flow electrophoresis. Electrophoresis produced substantial monocyte enrichment in some fractions with substantial depletion in others. MICC activity was seen to correlate with monocyte content in these fractions. Furthermore, removal of phagocytic cells with carbonyl iron and removal of adherent cells on plastic petri dishes depleted preparations of MICC activity. Thus it is suggested that under conditions described in this paper, the effector cell of the MICC assay in man appears to be a monocyte. This MICC effector cell was shown to be different from the effector cell in the natural killing (NK cells) of RPMI 6410 cells.     

Structure and mechanism of galactose oxidase: catalytic role of tyrosine 495

 The catalytic mechanism of the copper-containing enzyme galactose oxidase involves a protein radical on Tyr272, one of the equatorial copper ligands. The first step in this mechanism has been proposed to be the abstraction of a proton from the alcohol substrate by Tyr495, the axial copper ligand that is weakly co-ordinated to copper. In this study we have generated and studied the properties of a Y495F variant to test this proposal. X-ray crystallography reveals essentially no change from wild-type other than loss of the tyrosyl hydroxyl group. Visible spectroscopy indicates a significant change in the oxidised Y495F compared to wild-type with loss of a broad 810-nm peak, supporting the suggestion that this feature is due to inter-ligand charge transfer via the copper. The presence of a peak at 420 nm indicates that the Y495F variant remains capable of radical formation, a fact supported by EPR measurements. Thus the significantly reduced catalytic efficiency (1100-fold lower k _cat / K _m) observed for this variant is not due to an inability to generate the Tyr272 radical. By studying azide-induced pH changes, it is clear that the reduced catalytic efficiency is due mainly to the inability of Y495F to accept protons. This provides definitive evidence for the key role of Tyr495 in the initial proton abstraction step of the galactose oxidase catalytic mechanism. Received: 17 December 1996 / Accepted: 12 March 1997     

Can CD44 Be a Mediator of Cell Destruction? The Challenge of Type 1 Diabetes

CD44 is a multi-functional receptor with multiple of isoforms engaged in modulation of cell trafficking and transmission of apoptotic signals. We have previously shown that injection of anti-CD44 antibody into NOD mice induced resistance to type 1 diabetes (T1D). In this communication we describe our efforts to understand the mechanism underlying this effect. We found that CD44-deficient NOD mice develop stronger resistance to T1D than wild-type littermates. This effect is not explained by the involvement of CD44 in cell migration, because CD44-deficient inflammatory cells surprisingly had greater invasive potential than the corresponding wild type cells, probably owing to molecular redundancy. We have previously reported and we show here again that CD44 expression and hyaluronic acid (HA, the principal ligand for CD44) accumulation are detected in pancreatic islets of diabetic NOD mice, but not of non-diabetic DBA/1 mice. Expression of CD44 on insulin-secreting β cells renders them susceptible to the autoimmune attack, and is associated with a diminution in β-cells function (e.g., less insulin production and/or insulin secretion) and possibly also with an enhanced apoptosis rate. The diabetes-supportive effect of CD44 expression on β cells was assessed by the TUNEL assay and further strengthened by functional assays exhibiting increased nitric oxide release, reduced insulin secretion after glucose stimulation and decreased insulin content in β cells. All these parameters could not be detected in CD44-deficient islets. We further suggest that HA-binding to CD44-expressing β cells is implicated in β-cell demise. Altogether, these data agree with the concept that CD44 is a receptor capable of modulating cell fate. This finding is important for other pathologies (e.g., cancer, neurodegenerative diseases) in which CD44 and HA appear to be implicated.     

Effectiveness of a Universal Parental Support Programme to Promote Healthy Dietary Habits and Physical Activity and to Prevent Overweight and Obesity in 6-Year-Old Children: The Healthy School Start Study,a Cluster-Randomised Controlled Trial

ObjectiveTo develop and evaluate the effectiveness of a parental support programme to promote healthy dietary and physical activity habits and to prevent overweight and obesity in Swedish children.MethodsA cluster-randomised controlled trial was carried out in areas with low to medium socio-economic status. Participants were six-year-old children (n = 243) and their parents. Fourteen pre-school classes were randomly assigned to intervention (n = 7) and control groups (n = 7). The intervention lasted for 6 months and included: 1) Health information for parents, 2) Motivational Interviewing with parents and 3) Teacher-led classroom activities with children. Physical activity was measured by accelerometry, dietary and physical activity habits and parental self-efficacy through a questionnaire. Body weight and height were measured and BMI standard deviation score was calculated. Measurements were conducted at baseline, post-intervention and at 6-months follow-up. Group differences were examined using analysis of covariance and Poisson regression, adjusted for gender and baseline values.ResultsThere was no significant intervention effect in the primary outcome physical activity. Sub-group analyses showed a significant gender-group interaction in total physical activity (TPA), with girls in the intervention group demonstrating higher TPA during weekends (p = 0.04), as well as in sedentary time, with boys showing more sedentary time in the intervention group (p = 0.03). There was a significantly higher vegetable intake (0.26 servings) in the intervention group compared to the control group (p = 0.003). At follow-up, sub-group analyses showed a sustained effect for boys. The intervention did not affect the prevalence of overweight or obesity.ConclusionsIt is possible to influence vegetable intake in children and girls’ physical activity through a parental support programme. The programme needs to be intensified in order to increase effectiveness and sustain the effects long-term. These findings are an important contribution to the further development of evidence-based parental support programmes to prevent overweight and obesity in children.

Trial Registration

Controlled-trials.com ISRCTN32750699     

Genome-wide analyses implicate 33 loci in heritable dog osteosarcoma, including regulatory variants near CDKN2A/B

Background

Canine osteosarcoma is clinically nearly identical to the human disease, but is common and highly heritable, making genetic dissection feasible.

Results

Through genome-wide association analyses in three breeds (greyhounds, Rottweilers, and Irish wolfhounds), we identify 33 inherited risk loci explaining 55% to 85% of phenotype variance in each breed. The greyhound locus exhibiting the strongest association, located 150 kilobases upstream of the genes CDKN2A/B, is also the most rearranged locus in canine osteosarcoma tumors. The top germline candidate variant is found at a >90% frequency in Rottweilers and Irish wolfhounds, and alters an evolutionarily constrained element that we show has strong enhancer activity in human osteosarcoma cells. In all three breeds, osteosarcoma-associated loci and regions of reduced heterozygosity are enriched for genes in pathways connected to bone differentiation and growth. Several pathways, including one of genes regulated by miR124, are also enriched for somatic copy-number changes in tumors.

Conclusions

Mapping a complex cancer in multiple dog breeds reveals a polygenic spectrum of germline risk factors pointing to specific pathways as drivers of disease.     

Identification of genomic regions associated with phenotypic variation between dog breeds using selection mapping

The extraordinary phenotypic diversity of dog breeds has been sculpted by a unique population history accompanied by selection for novel and desirable traits. Here we perform a comprehensive analysis using multiple test statistics to identify regions under selection in 509 dogs from 46 diverse breeds using a newly developed high-density genotyping array consisting of >170,000 evenly spaced SNPs. We first identify 44 genomic regions exhibiting extreme differentiation across multiple breeds. Genetic variation in these regions correlates with variation in several phenotypic traits that vary between breeds, and we identify novel associations with both morphological and behavioral traits. We next scan the genome for signatures of selective sweeps in single breeds, characterized by long regions of reduced heterozygosity and fixation of extended haplotypes. These scans identify hundreds of regions, including 22 blocks of homozygosity longer than one megabase in certain breeds. Candidate selection loci are strongly enriched for developmental genes. We chose one highly differentiated region, associated with body size and ear morphology, and characterized it using high-throughput sequencing to provide a list of variants that may directly affect these traits. This study provides a catalogue of genomic regions showing extreme reduction in genetic variation or population differentiation in dogs, including many linked to phenotypic variation. The many blocks of reduced haplotype diversity observed across the genome in dog breeds are the result of both selection and genetic drift, but extended blocks of homozygosity on a megabase scale appear to be best explained by selection. Further elucidation of the variants under selection will help to uncover the genetic basis of complex traits and disease.