HIPs and HIP-reactive T cells.

Mounting evidence implicates hybrid insulin peptides (HIPs) as important autoantigens in the development of type 1 diabetes (T1D). These fusion peptides formed between insulin and other pancreatic beta cell-derived peptides contain non-genomically encoded amino acid sequences, making them plausible targets for autoreactive T cells in T1D. HIPs are detectable by mass spectrometry in human and murine islets and are targeted by diabetes-inducing T cells in non-obese diabetic mice as well as by T cells isolated from the residual pancreatic islets of human organ donors with T1D. The discovery of HIPs comes with numerous new challenges, as well as opportunities to study the pathogenesis of T1D. Here we review the original discovery of HIPs and describe recent studies investigating the role of HIP-reactive T cells in the development of diabetes. We also discuss potential mechanisms that may be responsible for the generation of HIPs in beta cells and describe challenges that need to be addressed in the field of mass spectrometry to enable the discovery of new HIPs. The identification of these potentially disease-driving antigens in T1D is of key interest to the field as it may provide new tools to predict, prevent and potentially reverse the disease.

Identification of Hybrid Insulin Peptides (HIPs) in Mouse and Human Islets by Mass Spectrometry.

We recently discovered hybrid insulin peptides (HIPs) as a novel class of post-translationally modified peptides in murine-derived beta cell tumors, and we demonstrated that these molecules are autoantigens in type 1 diabetes (T1D). A HIP consists of an insulin fragment linked to another secretory granule peptide via a peptide bond. We verified that autoreactive CD4 T cells in both mouse and human autoimmune diabetes recognize these modified peptides. Here, we use mass spectrometric analyses to confirm the presence of HIPs in both mouse and human pancreatic islets. We also present criteria for the confident identification of these peptides. This work supports the hypothesis that HIPs are autoantigens in human T1D and provides a foundation for future efforts to interrogate this previously unknown component of the beta cell proteome.

Are English CT departments and radiographers prepared for the morbidly obese patient?

INTRODUCTION: Morbid obesity is increasing in England, as is the use of CT scanning. All CT scanners have weight and body width limits. It is imperative that the radiographer performing the scan is aware of these limits, particularly in an emergency. This study aim was to determine whether radiographers are aware of their scanner limits, where they may be able to send a patient who exceeds these limits and whether a formal protocol exists. The secondary aim of the study was to determine capacities of scanners in acute trusts throughout England. METHODS: CT radiographers from 86 English Hospital Trusts with Emergency Departments were contacted and asked questions regarding their CT scanners and their practice of CT scanning morbidly obese patients. RESULTS: 21% of CT radiographers did not know the maximum width capacity of their scanner. Only 24% knew where a nearby larger capacity scanner was located and only 3% had a formal protocol for scanning obese patients. Weight capacities ranged from 147 to 305 kg. Width capacities ranged from 55 to 100 cm. 70% had weight capacity 226 kg or less and 70% had size capacity of 78 cm or less. CONCLUSION: Patients over 226 kg or 78 cm may not be accommodated in most (70%) trusts in England. Lack of knowledge of scanner capacities and alternative scanners for morbidly obese patients could have consequences for these patients, particularly in an emergency. The authors advise that all acute trusts have a protocol regarding CT scanning morbidly obese to prevent delays in accessing imaging.

Best practices for germ-free derivation and gnotobiotic zebrafish husbandry.

All animals are ecosystems with resident microbial communities, referred to as microbiota, which play profound roles in host development, physiology, and evolution. Enabled by new DNA sequencing technologies, there is a burgeoning interest in animal-microbiota interactions, but dissecting the specific impacts of microbes on their hosts is experimentally challenging. Gnotobiology, the study of biological systems in which all members are known, enables precise experimental analysis of the necessity and sufficiency of microbes in animal biology by deriving animals germ-free (GF) and inoculating them with defined microbial lineages. Mammalian host models have long dominated gnotobiology, but we have recently adapted gnotobiotic approaches to the zebrafish (Danio rerio), an important aquatic model. Zebrafish offer several experimental attributes that enable rapid, large-scale gnotobiotic experimentation with high replication rates and exquisite optical resolution. Here we describe detailed protocols for three procedures that form the foundation of zebrafish gnotobiology: derivation of GF embryos, microbial association of GF animals, and long-term, GF husbandry. Our aim is to provide sufficient guidance in zebrafish gnotobiotic methodology to expand and enrich this exciting field of research.

Effect of early grain feeding of beef steers on postabsorptive metabolism.

The objective of this study was to determine the effect of early weaning followed by a period of high-grain feeding on plasma acetate kinetics and signaling protein phosphorylation in LM tissue of growing steers. We hypothesized that early grain feeding would result in altered cell signaling and acetate use to support observed improvements in carcass gain and marbling. Fall-born Angus × Simmental steers were weaned at 106 ± 4 d of age (early weaned [EW]; n = 6) and fed a high-grain diet for 148 d or remained with their dams (normal weaned [NW]; n = 6) on pasture until weaning at 251 ± 5 d of age. Both treatments were subsequently combined and grazed on mixed summer pasture to 394 ± 5 d of age followed by a feedlot ration until harvest at 513 ± 5 d of age. Longissimus muscle tissue biopsies were collected at 253 ± 5 and 394 ± 5 d of age and at harvest. Total and phosphorylated forms of 5' adenosine monophosphate-activated protein kinase (AMPK) and downstream proteins of the mammalian target of rapamycin signaling pathway were determined by western blotting. Eight steers were used to assess acetate clearance at different age points via a bolus infusion of acetate (4 mmol/kg of BW). Early weaned steers had greater (P < 0.05) ADG than NW steers during the early grain feeding period. Phosphorylated to total ratios of ribosomal protein S6 (rpS6) and ribosomal protein S6 kinase 1 (S6K1) were significantly different during the early grain feeding period. Phosphorylated to total ratios of S6K1, rpS6, acetyl-CoA carboxylase, and 4E binding protein 1 and the absolute amount of phosphorylated AMPK were correlated with ADG, explaining 46% of the variance. Acetate clearance rates were less (P < 0.05) and synthesis rates were greater (P = 0.06) in EW steers during early grain feeding. Acetate synthesis rates were also greater (P < 0.05) in NW steers at harvest, suggesting a permanent shift in the gut microflora or gut function in response to the treatment. Neither treatment nor acetate infusion significantly affected plasma glucose or insulin concentrations. Plasma ß-hydroxybutyric acid concentrations increased with acetate infusion (P < 0.05). Based on these results, altered cell signaling during the early grain feeding period likely mediated increased protein deposition, leading to increased carcass weights, but observed changes in acetate appearance and clearance rates do not appear to explain the observed differences in intramuscular fat deposition during the terminal feeding period.

Acetate and glucose incorporation into subcutaneous, intramuscular, and visceral fat of finishing steers.

The objectives of this study were to assess the effects of early grain feeding on acetate and glucose turnover rates and acetate and glucose preference for palmitate synthesis by subcutaneous fat (SCF), intramuscular fat (IMF), and visceral fat (VF) in finishing steers. Sixteen Angus × Simmental steers were used in the study; 8 were early weaned (EW) and fed a high-grain diet immediately after weaning for 100 or 148 d, and 8 remained with their dams on pasture until weaning at 202 ± 5 or 253 ± 5 d of age. Normal weaned (NW) and EW animals were combined and grazed to 374 ± 5 or 393 ± 5 d of age, when they were placed on a corn silage-based finishing ration until they achieved a SCF thickness of 1.0 to 1.2 cm (494 ± 17 d of age for EW steers and 502 ± 12 d of age for NW steers). Immediately before harvest, steers were continuously infused for 12 h with [2H3] acetate (1.63 mmol/min; n = 8) or [U-13C6] glucose (0.07 mmol/min; n = 8). Blood samples were collected before initiation of infusions and at the end of the infusion from 8 animals or at 1-h intervals for the first 11 h and at 15-min intervals for the last hour of infusion for the other 8 animals. Adipose tissue samples from SCF, IMF, and VF depots were collected at harvest, and lipids were extracted. Plasma enrichments of acetate and glucose and palmitate enrichment in each depot were used to calculate plasma turnover rates and fractional synthesis rates (FSR; % per h) of palmitate from each isotope. Early weaned steers had greater marbling scores compared to NW steers ( P< 0.05). Plasma turnover rates and FSR for EW and NW steers were similar except for SCF, where a greater FSR from acetate was observed for EW steers. It is possible the greater FSR for SCF was due to harvesting the animals at a slightly more advanced stage of conditioning as evidenced by the trend for greater 12th rib fat (P = 0.07). Plasma acetate turnover and palmitate FSR from acetate were much greater (P < 0.05) than the corresponding rates from glucose, supporting the primary role of acetate as an energy source and the primary substrate for lipid synthesis across fat depots. However, FSR from acetate and glucose were not different among depots, suggesting that any potential effects of dietary starch on differential deposition of energy in SCF and IMF are not substrate driven.

Controlled formation and reflection of a bright solitary matter-wave.

Bright solitons are non-dispersive wave solutions, arising in a diverse range of nonlinear, one-dimensional systems, including atomic Bose-Einstein condensates with attractive interactions. In reality, cold-atom experiments can only approach the idealized one-dimensional limit necessary for the realization of true solitons. Nevertheless, it remains possible to create bright solitary waves, the three-dimensional analogue of solitons, which maintain many of the key properties of their one-dimensional counterparts. Such solitary waves offer many potential applications and provide a rich testing ground for theoretical treatments of many-body quantum systems. Here we report the controlled formation of a bright solitary matter-wave from a Bose-Einstein condensate of (85)Rb, which is observed to propagate over a distance of ∼1.1 mm in 150 ms with no observable dispersion. We demonstrate the reflection of a solitary wave from a repulsive Gaussian barrier and contrast this to the case of a repulsive condensate, in both cases finding excellent agreement with theoretical simulations using the three-dimensional Gross-Pitaevskii equation.

Magnetic transport apparatus for the production of ultracold atomic gases in the vicinity of a dielectric surface.

We present an apparatus designed for studies of atom-surface interactions using quantum degenerate gases of (85)Rb and (87)Rb in the vicinity of a room temperature dielectric surface. The surface to be investigated is a super-polished face of a glass Dove prism mounted in a glass cell under ultra-high vacuum. To maintain excellent optical access to the region surrounding the surface, magnetic transport is used to deliver ultracold atoms from a separate vacuum chamber housing the magneto-optical trap (MOT). We present a detailed description of the vacuum apparatus highlighting the novel design features; a low profile MOT chamber and the inclusion of an obstacle in the transport path. We report the characterization and optimization of the magnetic transport around the obstacle, achieving transport efficiencies of 70% with negligible heating. Finally, we demonstrate the loading of a hybrid optical-magnetic trap with (87)Rb and the creation of Bose-Einstein condensates via forced evaporative cooling close to the dielectric surface.

Preliminary examination of integrated vector management in a tropical rainforest area of Cameroon.

In the tropical rainforest area of Cameroon, people are affected by blackflies (Simulium spp.) and mosquitoes (Anopheles spp). Use of insecticide-treated bed nets (ITNs) has been promoted to protect vulnerable groups from mosquito bites, whereas historically indoor residual spraying (IRS) was the primary intervention. In a malaria-endemic area, a pilot study examined different mosquito control interventions applied to entire villages to assess their impact on vectors, malaria incidence and the quality of life of the communities. The Sanaga River near these villages was treated with insecticide to kill blackfly larvae. A medical survey of the six villages had shown that 20% of the population suffered from malaria, while 50% were infected with onchocerciasis and 5% with Loa loa. IRS+ITN using ICON CS (lambda-cyhalothrin capsule suspension formulation) or improved screening of houses combined with outdoor misting reduced the numbers of mosquitoes collected from exit traps compared to the other treatments. More sporozoites were detected in mosquitoes sampled in exit traps in the untreated village than in the treated villages. Malaria incidence several months after treatments was not significantly different from pre-treatment levels. Blackfly adult populations were reduced for several weeks following larvicide application but recovered when treatment was halted.

Use of transferrin-iron enterobactin complexes as the source of iron by serum-exposed bacteria.

Two smooth and six rough strains of Salmonella typhimurium with progressively smaller amounts of sugar and protein in their outer membrane were tested for degree of virulence in normal and iron-injected mice and for ability to acquire iron in mammalian sera. The rate of mortality showed that bacterial virulence for mice was lowered with progressive decrease of outer-membrane sugar and protein. Iron injections increased the rate of mortality in mice infected either with smooth strains or with superficially rough strains but were without effect in mice infected with deep rough strains. In in vitro experiments, iron promoted with equal effectiveness the growth of all serum-exposed bacterial strains, whereas enterobactin (E) was much more effective in promoting the growth of smooth and superficial rough than in promoting that of deep rough strains. Various experiments showed that deep rough strains cannot grow in E-supplemented serum because they are not able to use the transferrin-iron-E complexes that E forms with transferrin-iron. This failure to use transferrin-iron-E complexes by deep rough strains was found to be due to the inability of these strains to absorb iron containing complexes to their outer membrane. Adsorption studies with chemically treated bacteria showed that the receptor of transferrin-iron-E or E-iron complexes is a protein of the outer membrane of bacterial cells.

Virulence-associated acquisition of iron in mammalian serum by Escherichia coli.

Effects of iron on the growth of avirulent and virulent strains of Escherichia coli were tested in mice and in mammalian sera. Infection of the animals with iron increased mortality rates in mice infected with the avirulent strain to levels found in mice infected with the virulent strain. In vitro experiments showed that bacteria deprived of iron in bovine or human sera or milk or in chicken egg white stopped miltiplication and died in a very short time. These antibacterial effects were neutralized effectively with the addition of exogenous iron or the iron-binding bacterial product, enterochelin. In contrast to avirulent bacteria, which were effectively inhibited in mammalian serum, virulent bacteria were able to obtain iron and multiply. The ability of virulent bacteria to grow in mammalian serum is being attributed to the presence of iron-binding enterochelin and lipopolysaccharide in large amounts on the cell walls of virulent bacteria.