Ex vivo detectable human CD8 T-cell responses to cancer-testis antigens.

Clinical trials have shown that strong tumor antigen-specific CD8 T-cell responses are difficult to induce but can be achieved for T-cells specific for melanoma differentiation antigens, upon repetitive vaccination with stable emulsions prepared with synthetic peptides and incomplete Freund's adjuvant. Here, we show in four melanoma patients that ex vivo detectable T-cells and thus strong T-cell responses can also be induced against the more universal cancer-testis antigens NY-ESO-1 and Mage-A10. Interestingly, all patients had ex vivo detectable T-cell responses against multiple antigens after serial vaccinations with three peptides emulsified in incomplete Freund's adjuvant. Antigen-specific T-cells displayed an activated phenotype and secreted IFNgamma. The robust immune responses provide a solid basis for further development of human T-cell vaccination.

Loss of mating flight and shift in the pattern of carbohydrate storage in sexuals of ants (Hymenoptera; Formicidae)

In ants, energy for flying is derived from carbohydrates (glycogen and free sugars). The amount of these substrates was compared in sexuals participating or not participating in mating flights. Results show that in participating females (Lasius niger, L. flavus, Myrmica scabrinodis, Formica rufa, F. polyctena, F. lugubris), the amount of carbohydrates, especially glycogen, was higher than in non-participating females (Cataglyphis cursor, Iridomyrmex humilis). Similarly, male C. cursor and I. humilis which fly, exhibit a much higher carbohydrate content than do the non-flying females of these species. Furthermore, the quantity of carbohydrates stored was generally higher in males than in females for each species. These results are discussed with regard to the loss of the nuptial flight by some species of ants.

Nutritional influences on lipogenesis and thermogenesis after a carbohydrate meal.

In vivo lipogenesis and thermogenesis were studied for 24 h after ingestion of 500 g of carbohydrate (CHO) in subjects who had consumed either a high-fat, a mixed, or a high-CHO diet during the 3-6 days preceding the test. CHO oxidation and conversion to fat was significantly less in the high-fat diet group (222 +/- 5 g) than in the mixed (300 +/- 13 g) or high-CHO diet (331 +/- 7 g) groups, resulting in a greater glycogen storage in the high-fat (278 +/- 6 g) than in the other two groups (197 +/- 11 and 170 +/- 2 g). Net lipogenesis occurred sooner and lasted longer in the high-CHO group, amounting to 0.8 +/- 0.5, 3.4 +/- 0.6, and 9 +/- 1 g of lipid synthesized in the high-fat, mixed, and high-CHO groups, respectively. The thermic effect of the CHO load was 5.2 +/- 0.5% on the high-fat, 6.5 +/- 0.4% on the mixed diet, and 8.6 +/- 0.4% on the high-CHO diet. Significant relationships were demonstrated between the postabsorptive nonprotein respiratory quotient and net lipogenesis after the CHO load (r = 0.82) and between net lipogenesis and the increase in energy expenditure (r = 0.71). It is concluded that the antecedent diet influences the amount of net lipogenesis and the magnitude of thermogenesis after a large CHO test meal. However, lipogenesis remains too limited even after such large CHO intakes to cause an increase in the body's fat content.

Patterns of protein and carbohydrate accumulation during somatic embryogenesis of Acca sellowiana

The aim of this work was to quantify the protein, starch and total sugars levels during histodifferentiation and development of somatic embryos of Acca sellowiana Berg. For histological observations, the samples were dehydrated in a battery of ethanol, embedded in historesin and stained with toluidine blue (morphology), coomassie blue (protein bodies) and periodic acid-Schiff (starch). Proteins were extracted using a buffer solution, precipitated using ethanol and quantified using the Bradford reagent. Total sugars were extracted using a methanol-chloroform-water (12:5:3) solution and quantified by a reaction with anthrone at 0.2%. Starch was extracted using a 30% perchloric acid solution and quantified by a reaction with anthrone at 0.2%. During the somatic embryogenesis' in vitro morphogenesis and differentiation processes, the total protein levels decreased and the soluble sugars levels increased during the first 30 days in culture and remained stable until the 120th day. On the other hand, total protein levels increased according to the progression in the developmental stages of the somatic embryos. The levels of total sugars and starch increased in the heart and cotyledonary stages, and decreased in the torpedo and pre-cotyledonary stages. These compounds play a central role in the development of somatic embryos of Acca sellowiana.

An analysis of the benefit of using HEV genotype 3 antigens in detecting anti-HEV IgG in a European population.

BACKGROUND: The benefit of using serological assays based on HEV genotype 3 in industrialised settings is unclear. We compared the performance of serological kits based on antigens from different HEV genotypes. METHODS: Taking 20 serum samples from patients in southwest France with acute HEV infection (positive PCR for HEV genotype 3) and 550 anonymised samples from blood donors in southwest Switzerland, we tested for anti-HEV IgG using three enzyme immunoassays (EIAs) (MP Diagnostics, Dia.Pro and Fortress) based on genotype 1 and 2 antigens, and one immunodot assay (Mikrogen Diagnostik recomLine HEV IgG/IgM) based on genotype 1 and 3 antigens. RESULTS: All acute HEV samples and 124/550 blood donor samples were positive with ≥1 assay. Of PCR-confirmed patient samples, 45%, 65%, 95% and 55% were positive with MP Diagnostics, Dia.Pro, Fortress and recomLine, respectively. Of blood donor samples positive with ≥1 assay, 120/124 (97%), were positive with Fortress, 19/124 (15%) were positive with all EIAs and 51/124 (41%) were positive with recomLine. Of 11/20 patient samples positive with recomLine, stronger reactivity for HEV genotype 3 was observed in 1/11(9%), and equal reactivity for both genotypes in 5/11 (45.5%). CONCLUSIONS: Although recomLine contains HEV genotype 3, it has lower sensitivity than Fortress in acute HEV infection and fails to identify infection as being due to this genotype in approximately 45% of patients. In our single blood donor population, we observe wide variations in measured seroprevalence, from 4.2% to 21.8%, depending on the assay used.

Carbohydrate and nitrogen stores in Festuca paniculata under mowing explain dominance in subalpine grasslands

Cessation of traditional management threatens semi-natural grassland diversity through the colonisation or increase of competitive species adapted to nutrient-poor conditions. Regular mowing is one practice that controls their abundance. This study evaluated the ecophysiological mechanisms limiting short- and long-term recovery after mowing for Festuca paniculata, a competitive grass that takes over subalpine grasslands in the Alps following cessation of mowing. We quantified temporal variations in carbon (C) and nitrogen (N) content, starch, fructan and total soluble sugars in leaves, stem bases and roots of F. paniculata during one growth cycle in mown and unmown fields and related them to the dynamics of soil mineral N concentration and soil moisture. Short-term results suggest that the regrowth of F. paniculata following mowing might be N-limited, first because of N dilution by C increments in the plant tissue, and second, due to low soil mineral N and soil moisture at this time of year. However, despite short-term effects of mowing on plant growth, C and N content and concentration at the beginning of the following growing season were not affected. Nevertheless, total biomass accumulation at peak standing biomass was largely reduced compared to unmown fields. Moreover, lower C storage capacity at the end of the growing season impacted C allocation to vegetative reproduction during winter, thereby dramatically limiting the horizontal growth of F. paniculata tussocks in the long term. We conclude that mowing reduces the growth of F. paniculata tussocks through both C and N limitation. Such results will help understanding how plant responses to defoliation regulate competitive interactions within plant communities.

CSP-A Model for In Vivo Presentation of Plasmodium berghei Sporozoite Antigens by Hepatocytes.

One target of protective immunity against the Plasmodium liver stage in BALB/c mice is represented by the circumsporozoite protein (CSP), and mainly involves its recognition by IFN-γ producing specific CD8+T-cells. In a previous in vitro study we showed that primary hepatocytes from BALB/c mice process Plasmodium berghei (Pb) CSP (PbCSP) and present CSP-derived peptides to specific H-2k(d) restricted CD8+T-cells with subsequent killing of the presenting cells. We now extend these observations to an in vivo infection model in which infected hepatocytes and antigen specific T-cell clones are transferred into recipient mice inducing protection from sporozoite (SPZ) challenge. In addition, using a similar protocol, we suggest the capacity of hepatocytes in priming of naïve T-cells to provide protection, as further confirmed by induction of protection after depletion of cross-presenting dendritic cells (DCs) by cytochrome c (cyt c) treatment or using traversal deficient parasites. Our results clearly show that hepatocytes present Plasmodium CSP to specific-primed CD8+T-cells, and could also prime naïve T-cells, leading to protection from infection. These results could contribute to a better understanding of liver stage immune response and design of malaria vaccines.

Studies of the association of the A, B and Lewis Blood group antigens with carcinoembryonic antigen (CEA).

Carcinoembryonic antigen (CEA) was purified from primary tumour or from hepatic metastases obtained from ten cases of carcinoma of the colon. In nine cases the blood group antigens A, B, Lea or Leb were detected in CEA preparations by the binding of 125I-labelled CEA by blood group antibodies. The extent of binding appeared to preclude simple contamination of CEA preparations by blood group glycoprotein. In all cases the blood group antigens detected were consistent with the patients' known blood groups. Blood group I and i activities were not detected. It is concluded that the determinants of A, B and Lewis antigens and of CEA share the same glycoprotein carrier molecules.

Total and exogenous carbohydrate oxidation in obese prepubertal children.

The aim was to explore whether the origin of carbohydrate oxidation (exogenous compared with endogenous carbohydrate) after consumption of a mixed meal was influenced by obesity in children. Ten obese prepubertal children 8 y of age (44.2 +/- 3.6 kg) were studied over 9.5 h and compared with eight normal-weight, matched control children (28.5 +/- 1.6 kg). They were fed a mixed meal containing naturally enriched [13C]carbohydrate (cane sugar and popcorn) providing 55% of the daily energy requirement as measured by 24-h resting metabolic rate. Total carbohydrate oxidation was calculated by indirect calorimetry (hood system) whereas exogenous carbohydrate oxidation was estimated from carbon dioxide production (VCO2), the isotopic enrichment of breath 13CO2, and the abundance of [13C]carbohydrate in the meal ingested. The time course of 13CO2 in breath-measured over 570 min-followed a similar pattern in both groups. Although total carbohydrate oxidation was not significantly different among the two groups, exogenous carbohydrate utilization was significantly greater (P < 0.03) and endogenous carbohydrate oxidation was significantly lower (P < 0.05) in obese compared with control children. In addition, the rate of exogenous carbohydrate oxidation expressed as a proportion of total carbohydrate oxidation was positively related to the body fat of the children (r = 0.68, P < 0.01). The study suggests that in the postprandial phase, a smaller proportion of carbohydrate oxidation is accounted for by glycogen breakdown in obese children. The sparing of endogenous glycogen may result from decreased glycogen turnover already present at an early age.

Gastrointestinal transit of extruded or pelletized diets in pacu fed distinct inclusion levels of lipid and carbohydrate

The objective of this work was to evaluate the effect of pelletized or extruded diets, with different levels of carbohydrate and lipid, on the gastrointestinal transit time (GITT) and its modulation in pacu (Piaractus mesopotamicus). One hundred and eighty pacu juveniles were fed with eight isonitrogenous diets containing two carbohydrate levels (40 and 50%) and two lipid levels (4 and 8%). Four diets were pelletized and four were extruded. Carbohydrate and lipid experimental levels caused no changes to the bolus transit time. However, the bolus permanence time was related to diet processing. Fish fed pelletized diets exhibited the highest gastrointestinal transit time. Regression analysis of bolus behavior for pelletized and extruded diets with 4% lipid depicted different fits. GITT regression analysis of fish fed 8% lipid was fitted to a cubic equation and displayed adjustments of food permanence, with enhanced utilization of the diets, either with extruded or pelletized diets. GITT of fish fed extruded diets with 4% lipid was adjusted to a linear equation. The GITT of pacu depends on the diet processing and is affected by dietary levels of lipid and carbohydrate.

Translation and assembly of HLA-DR antigens in Xenopus oocytes injected with mRNA from a human B-cell line.

HLA-DR antigens are polymorphic cell surface glycoproteins, expressed primarily in B lymphocytes and macrophages, which are thought to play an important role in the immune response. Two polypeptide chains, alpha and beta, are associated at the cell surface, and a third chain associates with alpha and beta intracellularly. RNA isolated from the human B-cell line Raji was injected in Xenopus laevis oocytes. Immunoprecipitates of translation products with several monoclonal antibodies revealed the presence of HLA-DR antigens similar to those synthesized in Raji cells. One monoclonal antibody was able to bind the beta chain after dissociation of the three polypeptide chains with detergent. The presence of all three chains was confirmed by two-dimensional gel electrophoresis. The glycosylation pattern of the three chains was identical to that observed in vivo, as evidenced in studies using tunicamycin, an inhibitor of N-linked glycosylation. The presence of alpha chains assembled with beta chains in equimolar ratio was further demonstrated by amino-terminal sequencing. An RNA fraction enriched for the three mRNAs, encoding alpha, beta, and intracellular chains, was isolated. This translation-assembly system and the availability of monoclonal antibodies make it possible to assay for mRNA encoding specific molecules among the multiple human Ia-like antigens.

Longitudinal analyses of immune responses to Plasmodium falciparum derived peptides corresponding to novel blood stage antigens in coastal Kenya

We have recently described 95 predicted alpha-helical coiled-coil peptides derived from putative Plasmodium falciparum erythrocytic stage proteins. Seventy peptides recognized with the highest level of prevalence by sera from three endemic areas were selected for further studies. In this study, we sequentially examined antibody responses to these synthetic peptides in two cohorts of children at risk of clinical malaria in Kilifi district in coastal Kenya, in order to characterize the level of peptide recognition by age, and the role of anti-peptide antibodies in protection from clinical malaria. Antibody levels from 268 children in the first cohort (Chonyi) were assayed against 70 peptides. Thirty-nine peptides were selected for further study in a second cohort (Junju). The rationale for the second cohort was to confirm those peptides identified as protective in the first cohort. The Junju cohort comprised of children aged 1-6 years old (inclusive). Children were actively followed up to identify episodes of febrile malaria in both cohorts. Of the 70 peptides examined, 32 showed significantly (p<0.05) increased antibody recognition in older children and 40 showed significantly increased antibody recognition in parasitaemic children. Ten peptides were associated with a significantly reduced odds ratio (OR) for an episode of clinical malaria in the first cohort of children and two of these peptides (LR146 and AS202.11) were associated with a significantly reduced OR in both cohorts. LR146 is derived from hypothetical protein PFB0145c in PlasmoDB. Previous work has identified this protein as a target of antibodies effective in antibody dependent cellular inhibition (ADCI). The current study substantiates further the potential of protein PFB0145c and also identifies protein PF11_0424 as another likely target of protective antibodies against P. falciparum malaria