Extracellular hemoglobin combined with an O2-generating material overcomes O2 limitation in the bioartificial pancreas

The bioartificial pancreas encapsulating pancreatic islets in immunoprotective hydrogel is a promising therapy for Type 1 diabetes. As pancreatic islets are highly metabolically active and exquisitely sensitive to hypoxia, maintaining O₂ supply after transplantation remains a major challenge. In this study, we address the O₂ limitation by combining silicone-encapsulated CaO₂ (silicone-CaO₂) to generate O₂ with an extracellular hemoglobin O₂-carrier coencapsulated with islets. We showed that the hemoglobin improved by 37% the O₂-diffusivity through an alginate hydrogel and displayed antioxidant properties neutralizing deleterious reactive O₂ species produced by silicone-CaO₂. While the hemoglobin alone failed to maintain alginate macroencapsulated neonate pig islets under hypoxia, silicone-CaO₂ alone or combined to the hemoglobin restored islet viability and insulin secretion and prevented proinflammatory metabolism (PTGS2 expression). Interestingly, the combination took the advantages of the two individual strategies, improved neonate pig islet viability and insulin secretion in normoxia, and VEGF secretion and PDK1 normalization in hypoxia. Moreover, we confirmed the specific benefits of the combination compared to silicone-CaO₂ alone on murine pseudo-islet viability in normoxia and hypoxia. For the first time, our results show the interest of combining an O₂ provider with hemoglobin as an effective strategy to overcome O₂ limitations in tissue engineering.     

Cutting edge: characterization of allorestricted and peptide-selective alloreactive T cells using HLA-tetramer selection

The vast majority of alloreactive T cells recognize foreign MHC molecules in a peptide-dependent manner. A subpopulation of these peptide-dependent alloreactive T cells is peptide-specific and contains T cells that are of interest for tumor immunotherapy. Allorestricted T cells (i.e., peptide-specific and alloreactive) specific for tumor-associated Ags can be raised in vitro. However, it is technically difficult to distinguish between peptide-specific and peptide-nonspecific alloreactive T cells by functional assays in vitro. Here we show for the first time that allorestricted T cells specifically bind HLA-peptide tetrameric complexes, as nominal Ag-specific T cells would do. In consequence, fluorescent HLA-peptide tetrameric complexes can be used for sorting and cloning of allorestricted CTLs specific for a peptide of interest. We also show by the mean of HLA-peptide tetramers the existence of peptide-selective alloreactive T cells that recognize a conformation on the foreign-MHC brought about by some but not all peptides bound.     

Could the egg itself be the source of the oviposition deterrent marker in the ectoparasitoid Dinarmus basalis?

Females of Dinarmus basalis, a solitary ectoparasitoid, strongly avoid superparasitising hosts bearing eggs which are 20 or more hours old. Our objective was to determine the mechanism by which the factors inducing host discrimination and such an oviposition deterring effect act, and the origin of the deterrent. This study showed that the information is carried by the parasitised host. It is acquired by a transfer of molecules from the wasp egg onto the host and it requires contact between egg and host for at least 4 h. The factors transferred do not, as commonly reported in other species, come from the female's genital apparatus but from the live egg itself. Consequently, the D. basalis egg directly participates in the success of parasitism by releasing factors which elicit the oviposition host deterring effect. The originality and the ecological importance of these results are discussed.     

Prevalence and seasonal variations of six bee viruses in Apis mellifera L. and Varroa destructor mite populations in France

A survey of six bee viruses on a large geographic scale was undertaken by using seemingly healthy bee colonies and the PCR technique. Samples of adult bees and pupae were collected from 36 apiaries in the spring, summer, and autumn during 2002. Varroa destructor samples were collected at the end of summer following acaricide treatment. In adult bees, during the year deformed wing virus (DWV) was found at least once in 97% of the apiaries, sacbrood virus (SBV) was found in 86% of the apiaries, chronic bee paralysis virus (CBPV) was found in 28% of the apiaries, acute bee paralysis virus (ABPV) was found in 58% of the apiaries, black queen cell virus (BQCV) was found in 86% of the apiaries, and Kashmir bee virus (KBV) was found in 17% of the apiaries. For pupae, the following frequencies were obtained: DWV, 94% of the apiaries; SBV, 80% of the apiaries; CBPV, none of the apiaries; ABPV, 23% of the apiaries; BQCV, 23% of the apiaries; and KBV, 6% of the apiaries. In Varroa samples, the following four viruses were identified: DWV (100% of the apiaries), SBV (45% of the apiaries), ABPV (36% of the apiaries), and KBV (5% of the apiaries). The latter findings support the putative role of mites in transmitting these viruses. Taken together, these data indicate that bee virus infections occur persistently in bee populations despite the lack of clinical signs, suggesting that colony disease outbreaks might result from environmental factors that lead to activation of viral replication in bees.     

Purification, cloning, and sequencing of a 3,5-dichlorophenol reductive dehalogenase from Desulfitobacterium frappieri PCP-1

A membrane-associated 3,5-dichlorophenol reductive dehalogenase was isolated from Desulfitobacterium frappieri PCP-1. The highest dehalogenase activity was observed with the biomass cultured at 22 degrees C, compared to 30 and 37 degrees C, where the cell suspensions were 2.2 and 9.6 times less active, respectively. The reductive dehalogenase was purified 12.7-fold to apparent homogeneity. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a single band with an apparent molecular mass of 57 kDa. Its dechlorinating activity was not inhibited by sulfate and nitrate but was completely inhibited by 2.5 mM sulfite and 10 mM KCN. A mixture of iodopropane and titanium citrate caused a light-reversible inhibition of the dechlorinating activities, suggesting the involvement of a corrinoid cofactor. Several polychlorophenols were dechlorinated at the meta and para positions. The apparent K(m) for 3,5-dicholorophenol was 49.3 +/- 3.1 microM at a methyl viologen concentration of 2 mM. Six internal tryptic peptides were sequenced by mass spectrometry. One open reading frame (ORF) was found in the Desulfitobacterium hafniense genome containing these peptide sequences. This ORF corresponds to a gene coding for a CprA-type reductive dehalogenase. The corresponding ORF (named cprA5) in D. frappieri PCP-1 was cloned and sequenced. The cprA5 gene codes for a 548-amino-acid protein that contains a twin-arginine-type signal for secretion. The gene product has a cobalamin binding site motif and two iron-sulfur binding motifs and shows 66% identity (76 to 77% similarity) with some tetrachloroethene reductive dehalogenases. This is the first CprA-type reductive dehalogenase that can dechlorinate chlorophenols at the meta and para positions.     

Trophic interactions in a high arctic snow goose colony

We examined the role of trophic interactions in structuringa high arctic tundra community characterized by a large breedingcolony of greater snow geese (Chen caerulescens atlantica).According to the exploitation ecosystem hypothesis of Oksanenet al. (1981), food chains are controlled by top-down interactions.However, because the arctic primary productivity is low, herbivorepopulations are too small to support functional predator populationsand these communities should thus be dominated by the plant/herbivore trophic-level interaction. Since 1990, we have beenmonitoring annual abundance and productivity of geese, the impactof goose grazing, predator abundance (mostly arctic foxes, Alopexlagopus) and the abundance of lemmings, the other significantherbivore in this community, on Bylot Island, Nunavut, Canada.Goose grazing consistently removed a significant proportionof the standing crop (     

Annotated expressed sequence tags for studies of the regulation of reproductive modes in aphids

The damaging effect of aphids to crops is largely determined by the spectacular rate of increase of populational expansion due to their parthenogenetic generations. Despite this, the molecular processes triggering the transition between the parthenogenetic and sexual phases between their annual life cycle have received little attention. Here, we describe a collection of genes from the cereal aphid Rhopalosiphum padi expressed during the switch from parthenogenetic to sexual reproduction. After cDNA cloning and sequencing, 726 expressed sequence tags (EST) were annotated. The R. padi EST collection contained a substantial number (139) of bacterial endosymbiont sequences. The majority of R. padi cDNAs encoded either unknown proteins (56%) or housekeeping polypeptides (38%). The large proportion of sequences without similarities in the databases is related to both their small size and their high GC content, corresponding probably to the presence of 5'-unstranslated regions. Fifteen genes involved in developmental and differentiation events were identified by similarity to known genes. Some of these may be useful candidates for markers of the early steps of sexual differentiation.