E-LDL upregulates TOSO expression and enhances the survival of human macrophages

Uptake of modified lipoproteins by macrophages causes foam cell formation and promotes atherosclerosis. Atherogenic lipoproteins are cytotoxic and induce cell death under certain conditions but may also enhance macrophage survival. Macrophages treated with enzymatically modified LDL (E-LDL) were subjected to GeneChip analysis and the antiapoptotic gene TOSO was found induced. TOSO mRNA is upregulated and apoptosis is reduced in E-LDL but not in oxidized LDL (Ox-LDL) loaded macrophages. FLIP(L) abundance was suggested to mediate the antiapoptotic properties of TOSO; however, FLIP(L) was not changed. Ox-LDL is internalized predominantly by scavenger receptors such as CD36 while E-LDL particles are preferentially internalized by Fc- and complement-receptor dependent phagocytosis and internalization of phagobeads by macrophages upregulates TOSO. In COS-7 cells however, phagocytotic activity was not affected by TOSO. These data indicate that E-LDL-generated foam cells are protected from cell death most likely through the expression of TOSO by a FLIP(L) independent mechanism.     

Warm temperature accelerates short photoperiod-induced growth cessation and dormancy induction in hybrid poplar (Populus × spp.)

There is increasing evidence that temperature, in addition to photoperiod, may be an important factor regulating bud dormancy. The impact of temperature during growth cessation, dormancy development, and subsequent cold acclimation was examined in four hybrid poplar clones with contrasting acclimation patterns: ‘Okanese’—EARLY, ‘Walker’—INT1, ‘Katepwa’—INT2, and ‘Prairie Sky’—LATE. Four day–night temperature treatments (13.5/8.5, 18.5/13.5, 23.5/8.5, and 18.5/3.5°C) were applied during a 60-day induction period to reflect current and predicted future annual variation in autumn temperature for Saskatoon, SK. Warm night temperature (18.5/13.5°C) strongly accelerated growth cessation, dormancy development, and cold acclimation in all four clones. Day temperature had the opposite effect of night temperature. Day and night temperatures appeared to act antagonistically against each other during growth cessation and subsequent dormancy development and cold acclimation. Growth cessation, dormancy development, and cold acclimation in EARLY and LATE were less affected by induction temperature than INT1 and INT2 suggesting that genotypic variations exist in response to temperature. Separating specific phenological stages and the impact by temperature on each clone revealed the complexity of fall phenological changes and their interaction with temperature. Most importantly, future changes in temperature may affect time to growth cessation, subsequently altering the depth of dormancy and cold hardiness in hybrid poplar.     

Human immunodeficiency virus-specific gamma interferon enzyme-linked immunospot assay responses targeting specific regions of the proteome during primary subtype C infection are poor predictors of the course of viremia and set point

It is unknown whether patterns of human immunodeficiency virus (HIV)-specific T-cell responses during acute infection may influence the viral set point and the course of disease. We wished to establish whether the magnitude and breadth of HIV type 1 (HIV-1)-specific T-cell responses at 3 months postinfection were correlated with the viral-load set point at 12 months and hypothesized that the magnitude and breadth of HIV-specific T-cell responses during primary infection would predict the set point. Gamma interferon (IFN-γ) enzyme-linked immunospot (ELISPOT) assay responses across the complete proteome were measured in 47 subtype C HIV-1-infected participants at a median of 12 weeks postinfection. When corrected for amino acid length and individuals responding to each region, the order of recognition was as follows: Nef > Gag > Pol > Rev > Vpr > Env > Vpu > Vif > Tat. Nef responses were significantly (P < 0.05) dominant, targeted six epitopic regions, and were unrelated to the course of viremia. There was no significant difference in the magnitude and breadth of responses for each protein region with disease progression, although there was a trend of increased breadth (mean, four to seven pools) in rapid progressors. Correlation of the magnitude and breadth of IFN-γ responses with the viral set point at 12 months revealed almost zero association for each protein region. Taken together, these data demonstrate that the magnitude and breadth of IFN-γ ELISPOT assay responses at 3 months postinfection are unrelated to the course of disease in the first year of infection and are not associated with, and have low predictive power for, the viral set point at 12 months.     

Glutamate metabolism in HIV-1 infected macrophages: Role of HIV-1 Vpr

HIV-1 infected macrophages play a significant role in the neuropathogenesis of AIDS. HIV-1 viral protein R (Vpr) not only facilitates HIV-1 infection but also contribute to long-lived persistence in macrophages. Our previous studies using SILAC-based proteomic analysis showed that the expression of critical metabolic enzymes in the glycolytic pathway and tricarboxylic acid (TCA) cycle were altered in response to Vpr expression in macrophages. We hypothesized that Vpr-induced modulation of glycolysis and TCA cycle regulates glutamate metabolism and release in HIV-1 infected macrophages.

We assessed the amount of specific metabolites induced by Vpr and HIV-1 in macrophages at the intracellular and extracellular level in a time-dependent manner utilizing multiple reaction monitoring (MRM) targeted metabolomics. In addition, stable isotope-labeled glucose and an MRM targeted metabolomics assay were used to evaluate the de novo synthesis and release of glutamate in Vpr overexpressing macrophages and HIV-1 infected macrophages, throughout the metabolic flux of glycolytic pathway and TCA cycle activation.

The metabolic flux studies demonstrated an increase in glucose uptake, glutamate release and accumulation of α-ketoglutarate (α-KG) and glutamine in the extracellular milieu in Vpr expressing and HIV-1 infected macrophages. Interestingly, glutamate pools and other intracellular intermediates (glucose-6-phosphate (G6P), fructose-6-phosphate (F6P), citrate, malate, α-KG, and glutamine) showed a decreased trend except for fumarate, in contrast to the glutamine accumulation observed in the extracellular space in Vpr overexpressing macrophages.

Our studies demonstrate that dysregulation of mitochondrial glutamate metabolism induced by Vpr in HIV-1 infected macrophages commonly seen, may contribute to neurodegeneration via excitotoxic mechanisms in the context of NeuroAIDS.     

Population size and structure of the African Softshell Turtle,Trionyx triunguis,in Dalaman,southwestern Turkey

We assessed the size of the population of the African Softshell Turtle, Trionyx triunguis, in Dalaman (Mu?la, Turkey), which is considered to be the largest population of the species in the Mediterranean, by using the Jolly-Seber mark-recapture method. A total of 415 individuals were caught during the summer months of 2009 to 2011, of which 148 were recaptures. From 267 marked individuals, 148 (55%) were male, 69 (26%) were female and 50 (19%) were juveniles and subadults of indeterminate sex. The male:female ratio was calculated to be 2.14:1. By using Model A' of the Jolly-Seber mark-recapture method, which includes deaths but no immigration, the mean population size in Kükürt and Küçükdalyan (Karg?n) lakes, together with Tersakan and Ta?l?çay creeks, were estimated to be 396±36. Based on this estimation, the population density was calculated to be 14 turtles/ha. Approximately 67.4% of the estimated population was marked. The mean capture probability (p) and mean survival ratio (Φ) were 0.094±0.009 and 0.957±0.076, respectively. Our results showed that the species’ population size is larger than previously estimated based on visual counts. The status “Vulnerable” C2a of the IUCN Red Data Book categories seems therefore more appropriate for the Mediterranean subpopulation of Trionyx triunguis than “Endangered” C2a.     

Nucleotide sequence encoding the 5'' end of Xenopus laevis 18S rRNA.

We have sequenced a region of cloned Xenopus laevis ribosomal DNA encompassing the last 24 nucleotides of the external transcribed spacer and the first 275 nucleotides of the 18S gene. The start of the 18S gene was identified by correlating the results obtained from RNA hybridization and fingerprinting with the DNA sequence. This 5' region of 18S rRNA contains five 2'-O-methyl groups and at least six pseudouridine residues. Several of these modified nucleotides are clustered into a relatively short region from nucleotides 99-124. Nucleotides 227-250 constitute a distinctive sequence of 24 consecutive G and C residues. Comparison with the first 160 nucleotides of a yeast 18S gene (25) reveals three blocks of high sequence homology separated by two short tracts where homology is low or absent. The external transcribed spacer sequences diverge widely from within a few nucleotides of the start of the 18S gene.     

Biosynthesis of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) and characterisation of its blend with oil palm empty fruit bunch fibers

Poly(3-hydroxybutyrate-co-38 mol%-3-hydroxyvalerate) [P(3HB-co-38mol%-3HV)] was produced by Cupriavidus sp. USMAA2-4 in the presence of oleic acid and 1-pentanol. Due to enormous production of empty fruit bunch (EFB) in the oil palm plantation and high production cost of P(3HB-co-3HV), oil palm EFB fibers were used for biocomposites preparation. In this study, maleic anhydride (MA) and benzoyl peroxide (DBPO) were used to improve the miscibility between P(3HB-co-3HV) and EFB fibers. Introduction of MA into P(3HB-co-3HV) backbone reduced the molecular weight and improved the thermal stability of P(3HB-co-3HV). Thermal stability of P(3HB-co-3HV)/EFB composites was shown to be comparable to that of commercial packaging product. Composites with 35% EFB fibers content have the highest tensile strength compared to 30% and 40%. P(3HB-co-3HV)/EFB blends showed less chemicals leached compared to commercial packaging.     

cMyc is a principal upstream driver of beta-cell proliferation in rat insulinoma cell lines and is an effective mediator of human beta-cell replication

Adult human β-cells replicate slowly. Also, despite the abundance of rodent β-cell lines, there are no human β-cell lines for diabetes research or therapy. Prior studies in four commonly studied rodent β-cell lines revealed that all four lines displayed an unusual, but strongly reproducible, cell cycle signature: an increase in seven G(1)/S molecules, i.e. cyclins A, D3, and E, and cdk1, -2, -4, and -6. Here, we explore the upstream mechanism(s) that drive these cell cycle changes. Using biochemical, pharmacological and molecular approaches, we surveyed potential upstream mitogenic signaling pathways in Ins 1 and RIN cells. We used both underexpression and overexpression to assess effects on rat and human β-cell proliferation, survival and cell cycle control. Our results indicate that cMyc is: 1) uniquely up-regulated among other candidates; 2) principally responsible for the increase in the seven G(1)/S molecules; and, 3) largely responsible for proliferation in rat β-cell lines. Importantly, cMyc expression in β-cell lines, although some 5- to 7-fold higher than normal rat β-cells, is far below the levels (75- to 150-fold) previously associated with β-cell death and dedifferentiation. Notably, modest overexpression of cMyc is able to drive proliferation without cell death in normal rat and human β-cells. We conclude that cMyc is an important driver of replication in the two most commonly employed rat β-cell lines. These studies reverse the current paradigm in which cMyc overexpression is inevitably associated with β-cell death and dedifferentiation. The cMyc pathway provides potential approaches, targets, and tools for driving and sustaining human β-cell replication.