Sweet Killing in Obesity and Diabetes: The Metabolic Role of the BH3-only Protein BIM

Diabetes is a metabolic disorder affecting more than 400 million individuals and their families worldwide. The major forms of diabetes (types 1 and 2) are characterized by pancreatic β-cell dysfunction and, in some cases, loss of β-cell mass causing hyperglycemia due to absolute or relative insulin deficiency. The BCL-2 homology 3 (BH3)-only protein BIM has a wide role in apoptosis induction in cells. In this review, we describe the apoptotic mechanisms mediated by BIM activation in β cells in obesity and both forms of diabetes. We focus on molecular pathways triggered by inflammation, saturated fats, and high levels of glucose. Besides its role in cell death, BIM has been implicated in the regulation of mitochondrial oxidative phosphorylation and cellular metabolism in hepatocytes. BIM is both a key mediator of pancreatic β-cell death and hepatic insulin resistance and is thus a potential therapeutic target for novel anti-diabetogenic drugs. We consider the implications and challenges of targeting BIM in the treatment of the disease.     

Expression of recombinant human granulocyte colony-stimulating factor in CHO dhfr cells: new insights into the in vitro amplification expression system

The in vitro amplification method for heterologous gene expression in mammalian cells is based on the stable transfection of cells with long, linear DNA molecules having several copies of complete expression units, coding for the gene of interest, linked to one terminal unit, coding for the selectable marker. DNA concatenamers containing additional expression units can also be prepared: we exploited this feature by co-polymerizing expression units coding for granulocyte colony-stimulating factor (G-CSF) with cassettes for dihydrofolate reductase (DHFR) and for neomycin (Nm) resistance, as selectable markers. We were thus able to obtain high level production of G-CSF in chinese hamster ovary (CHO) dhfr^- cells by combining in vitro amplification to just one step of in vivo amplification. This approach required a considerably shorter time than the classical, stepwise amplification by methotrexate.     

Structural studies of novel glycoconjugates from polymerized allergens (allergoids) and mannans as allergy vaccines

Immunotherapy for treating IgE-mediated allergies requires high doses of the corresponding allergen. This may result in undesired side effects and, to avoid them, hypoallergenic allergens (allergoids) polymerized with glutaraldehyde are commonly used. Targeting allergoids to dendritic cells to enhance cell uptake may result in a more effective immunotherapy. Allergoids coupled to yeast mannan, as source of polymannoses, would be suitable for this purpose, since mannose-binding receptors are expressed on these cells. Conventional conjugation procedures of mannan to proteins use oxidized mannan to release reactive aldehydes able to bind to free amino groups in the protein; yet, allergoids lack these latter because their previous treatment with glutaraldehyde. The aim of this study was to obtain allergoids conjugated to mannan by an alternative approach based on just glutaraldehyde treatment, taking advantage of the mannoprotein bound to the polymannose backbone. Allergoid-mannan glycoconjugates were produced in a single step by treating with glutaraldehyde a defined mixture of allergens derived from Phleum pratense grass pollen and native mannan (non-oxidized) from Saccharomyces cerevisae. Analytical and structural studies, including 2D-DOSY and ¹H-¹³C HSQC nuclear magnetic resonance spectra, demonstrated the feasibility of such an approach. The glycoconjugates obtained were polymers of high molecular weight showing a higher stability than the native allergen or the conventional allergoid without mannan. The allergoid-mannan glycoconjugates were hypoallergenic as detected by the IgE reactivity with sera from grass allergic patients, even with lower reactivity than conventional allergoid without mannan. Thus, stable hypoallergenic allergoids conjugated to mannan suitable for using in immunotherapy can be achieved using glutaraldehyde. In contrast to mannan oxidation, the glutaraldehyde approach allows to preserve mannoses with their native geometry, which may be functionally important for its receptor-mediated recognition.     

Genome divergence and diversification within a geographic mosaic of coevolution

Despite substantial interest in coevolution's role in diversification, examples of coevolution contributing to speciation have been elusive. Here, we build upon past studies that have shown both coevolution between South Hills crossbills and lodgepole pine (Pinus contorta), and high levels of reproductive isolation between South Hills crossbills and other ecotypes in the North American red crossbill (Loxia curvirostra) complex. We used genotyping by sequencing to generate population genomic data and applied phylogenetic and population genetic analyses to characterize the genetic structure within and among nine of the ecotypes. Although genome‐wide divergence was slight between ecotypes (F_ST = 0.011–0.035), we found evidence of relative genetic differentiation (as measured by F_ST) between and genetic cohesiveness within many of them. As expected for nomadic and opportunistic breeders, we detected no evidence of isolation by distance. The one sedentary ecotype, the South Hills crossbill, was genetically most distinct because of elevated divergence at a small number of loci rather than pronounced overall genome‐wide divergence. These findings suggest that mechanisms related to recent local coevolution between South Hills crossbills and lodgepole pine (e.g. strong resource‐based density dependence limiting gene flow) have been associated with genome divergence in the face of gene flow. Our results further characterize a striking example of coevolution driving speciation within perhaps as little as 6000 years.     

Secretion of plasminogen activators and their inhibitors in corneal fibroblasts. Modification of this secretion in Schnyder's lens corneal dystrophy

In this study, we have shown that the secretion of plasminogen activators (PA) by corneal fibroblasts from Schnyder dystrophy is much lower than that secreted by normal corneal fibroblasts. This defective secretion of PA was noted both in the supernatant of cell culture and on the cell surface. This anomaly is found in cultured fibroblasts and therefore is not related to environment modification. Since PA was implicated in remodeling of extracellular matrix, we hypothesized that this anomaly should be responsible for the corneal dystrophy.     

Comparative studies of Trypanosoma vespertilionis Battaglia and Trypanosoma dionisii Bettencourt & Fran?a.

In diphasic blood agar media Trypanosoma vespertilionis developed spheroid clusters as compared to rather long, sausage-shaped (sometimes branched) clusters formed by Trypanosoma dionisii. The former species attained a greater population density (approximately 6 X 10(7) organisms/ml) than the latter (approximately 2 X 10(7) organisms/ml). Greater numbers of epimastigotes, some in active binary divisions, were observed during the logarithmic phase of growth, and morphologic changes occurred during cultivation which correlated with increased acidity and a depletion of glucose. Maximum numbers of trypomastigote forms were found during the stationary and early death phases. Most of the forms observed after 20 days were sphaeromastigotes. Glucose concentrations decreased to 0 M in T. vespertilionis and to 4.4 X 10(-5) M in T. dionisii cultures during the stationary and death phases. By the 12th day of incubation cultures of T. vespertilionis were more acid (pH 5.5) than those of T. dionisii vespertilionis and T. dionisii contained common and specific antigens. At least 2-3 common antigens were detected in extracts reacted against heterologous antisera. Specific antigens were observed as nonidentical lines formed by extracts reacted against homologous and heterologous antisera and with antisera absorbed with heterologous antigens. At least 2 specific antigens were evident in extracts of T. vespertilionis and 1 in extracts of T. dionisii.