The hepcidin-binding site on ferroportin is evolutionarily conserved

Mammalian iron homeostasis is regulated by the interaction of the liver-produced peptide hepcidin and its receptor, the iron transporter ferroportin. Hepcidin binds to ferroportin resulting in degradation of ferroportin and decreased cellular iron export. We identify the hepcidin-binding domain (HBD) on ferroportin and show that a synthetic 19 amino acid peptide corresponding to the HBD recapitulates the characteristics and specificity of hepcidin binding to cell-surface ferroportin. The binding of mammalian hepcidin to ferroportin or the HBD shows an unusual temperature dependency with an increased rate of dissociation at temperatures below 15°C. The increased rate of dissociation is due to temperature- dependent changes in hepcidin structure. In contrast, hepcidin from poikilothermic vertebrates, such as fish or frogs, binds the HBD in a temperature-independent fashion. The affinity of hepcidin for the HBD permits a rapid, sensitive assay of hepcidin from all species and yields insights into the evolution of hepcidin.     

Characterization of enhanced monovalent and bivalent thrombin DNA aptamer binding using single molecule force spectroscopy

Thrombin aptamer binding strength and stability is dependent on sterical parameters when used for atomic force microscopy sensing applications. Sterical improvements on the linker chemistry were developed for high-affinity binding. For this we applied single molecule force spectroscopy using two enhanced biotinylated thrombin aptamers, BFF and BFA immobilized on the atomic force microscopy tip via streptavidin. BFF is a dimer composed of two single-stranded aptamers (aptabody) connected to each other by a complementary sequence close to the biotinylated end. In contrast, BFA consists of a single DNA strand and a complementary strand in the supporting biotinylated part. By varying the pulling velocity in force-distance cycles the formed thrombin-aptamer complexes were ruptured at different force loadings allowing determination of the energy landscape. As a result, BFA aptamer showed a higher binding force at the investigated loading rates and a significantly lower dissociation rate constant, k_off, compared to BFF. Moreover, the potential of the aptabody BFF to form a bivalent complex could clearly be demonstrated.     

Inflammatory cytokines in vitro production are associated with Ala16Val superoxide dismutase gene polymorphism of peripheral blood mononuclear cells

Obesity is considered a chronic low-grade inflammatory state associated with a chronic oxidative stress caused by superoxide production (O(2)(-)). The superoxide dismutase manganese dependent (SOD2) catalyzes O(2)(-) in H(2)O(2) into mitochondria and is encoded by a single gene that presents a common polymorphism that results in the replacement of alanine (A) with a valine (V) in the 16 codon. This polymorphism has been implicated in a decreased efficiency of SOD2 transport into targeted mitochondria in V allele carriers. Previous studies described an association between VV genotype and metabolic diseases, including obesity and diabetes. However, the causal mechanisms to explain this association need to be more elucidated. We postulated that the polymorphism could influence the inflammatory response. To test our hypothesis, we evaluated the in vitro cytokines production by human peripheral blood mononuclear cells (PBMCs) carrier's different Ala16Val-SOD2 genotypes (IL-1, IL-6, IL-10, TNF-α, IFN-γ). Additionally, we evaluated if the culture medium glucose, enriched insulin, could influence the cytokine production. Higher levels of proinflammatory cytokines were observed in VV-PBMCs when compared to AA-PBMCs. However, the culture medium glucose and enriched insulin did not affect cytokine production. The results suggest that Ala16Val-SOD2 gene polymorphism could trigger the PBMCs proinflammatory cytokines level. However, discerning if a similar mechanism occurs in fat cells is an open question.     

Enhanced antioxidant formula based on a selenium-supplemented carotenoid-producing yeast biomass

Carotenoid-producing yeast species such as Rhodotorula glutinis and Sporobolomyces roseus efficiently accumulated selenium from the growth medium. It was observed that incorporation of selenium into yeast cells during the growth inhibited production of beta-carotenoid and other carotenoid precursors (torularhodin and torulene). The yeasts with high content of the carotenoid pigments and selenium may be used for the preparation of a new type of antioxidant formula that could be directly applied for various human and animal diets. We have demonstrated that such a formula can only be produced by separate processes of the cultivation of red yeasts and a subsequent sorption of selenium into the cells.     

Immune and viral profile from tolerance to hepatitis B surface antigen clearance: a longitudinal study of vertically hepatitis B virus-infected children on combined therapy

The aim of the study was to investigate longitudinally hepatitis B virus (HBV)-specific T-cell reactivity and viral behavior versus treatment response in tolerant children during combined antiviral therapy. Twenty-three children with infancy-acquired hepatitis B (HBeAg(+)) belonging to a published pilot study of 1-year treatment with lamivudine/alpha interferon (IFN-α) were investigated. Five seroconverted to anti-HBs (responders). Nine were HLA-A2(+) (4 responders and 5 nonresponders). Mutations within the HBV core gene were determined at baseline in liver and in serial serum samples by direct sequencing at baseline; during treatment week 2 (TW2), TW9, TW28, and TW52; and after follow-up week 24 (FUW24) and FUW52. HBV-specific reactivity was evaluated by T-cell proliferation with 16 HBV core 20-mer overlapping peptides and by HLA-A2-restricted core(18-27) pentamer staining and CD8(+) IFN-γ enzyme-linked immunospot (ELISPOT) assay. HBV core-specific T-cell proliferative and CD8 responses were more vigorous and broader among responders than among nonresponders at TW28 and TW52, while the number of mutations within HBV core gene immunodominant epitopes was lower at TW28 and was negatively associated with HBV-specific T-cell proliferative responses at both time points. The HBV DNA viral load was negatively associated with HBV-specific T-cell proliferative and CD8 responses during treatment, especially at TW28. Treatment-induced transition from immunotolerance to HBV immune control is characterized by the emergence of efficient virus-specific immune responses capable of restraining mutations and preventing viral evasion.     

In chronic fatigue syndrome, the decreased levels of omega-3 poly-unsaturated fatty acids are related to lowered serum zinc and defects in T cell activation

There is now evidence that major depression is accompanied by decreased levels of omega3 poly-unsaturated fatty acids (PUFA), such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). There is a strong comorbidity between major depression and chronic fatigue syndrome (CFS). The present study has been carried out in order to examine PUFA levels in CFS. In twenty-two CFS patients and 12 normal controls we measured serum PUFA levels using gas chromatography and mass spectrometry. We found that CFS was accompanied by increased levels of omega6 PUFAs, i.e. linoleic acid and arachidonic acid (AA), and mono-unsaturated fatty acids (MUFAs), i.e. oleic acid. The EPA/AA and total omega3/omega6 ratios were significantly lower in CFS patients than in normal controls. The omega3/omega6 ratio was significantly and negatively correlated to the severity of illness and some items of the FibroFatigue scale, i.e. aches and pain, fatigue and failing memory. The severity of illness was significantly and positively correlated to linoleic and arachidonic acid, oleic acid, omega9 fatty acids and one of the saturated fatty acids, i.e. palmitic acid. In CFS subjects, we found significant positive correlations between the omega3/omega6 ratio and lowered serum zinc levels and the lowered mitogen-stimulated CD69 expression on CD3+, CD3+ CD4+, and CD3+ CD8+ T cells, which indicate defects in early T cell activation. The results of this study show that a decreased availability of omega3 PUFAs plays a role in the pathophysiology of CFS and is related to the immune pathophysiology of CFS. The results suggest that patients with CFS should respond favourably to treatment with--amongst other things--omega3 PUFAs, such as EPA and DHA.     

Analysis of SNPs and Haplotypes in Vitamin D Pathway Genes and Renal Cancer Risk

In the kidney vitamin D is converted to its active form. Since vitamin D exerts its activity through binding to the nuclear vitamin D receptor (VDR), most genetic studies have primarily focused on variation within this gene. Therefore, analysis of genetic variation in VDR and other vitamin D pathway genes may provide insight into the role of vitamin D in renal cell carcinoma (RCC) etiology. RCC cases (N = 777) and controls (N = 1,035) were genotyped to investigate the relationship between RCC risk and variation in eight target genes. Minimum-p-value permutation (Min-P) tests were used to identify genes associated with risk. A three single nucleotide polymorphism (SNP) sliding window was used to identify chromosomal regions with a False Discovery Rate of <10%, where subsequently, haplotype relative risks were computed in Haplostats. Min-P values showed that VDR (p-value = 0.02) and retinoid-X-receptor-alpha (RXRA) (p-value = 0.10) were associated with RCC risk. Within VDR, three haplotypes across two chromosomal regions of interest were identified. The first region, located within intron 2, contained two haplotypes that increased RCC risk by approximately 25%. The second region included a haplotype (rs2239179, rs12717991) across intron 4 that increased risk among participants with the TC (OR = 1.31, 95% CI = 1.09–1.57) haplotype compared to participants with the common haplotype, TT. Across RXRA, one haplotype located 3′ of the coding sequence (rs748964, rs3118523), increased RCC risk 35% among individuals with the variant haplotype compared to those with the most common haplotype. This study comprehensively evaluated genetic variation across eight vitamin D pathway genes in relation to RCC risk. We found increased risk associated with VDR and RXRA. Replication studies are warranted to confirm these findings.     

Reversal of IAA-induced inhibition of flowering by aminoethoxyvinylglycine inChenopodium

Aminoethoxyvinylglycine (AVG) applied as a droplet (3 l, 0.1 mM) to the plumule of seedlings of both the short-day plantChenopodium rubrum and the long-day plantChenopodium murale counteracted to a great extent or even canceled the inhibition of flowering due to exogenous indole-3-acetic acid (IAA). This effect was more pronounced with the two substances administered simultaneously than with later application of AVG alone. AVG by itself in some cases promoted the percentage of flowering in bothChenopodium species. Application of IAA to the shoot apex was shown to elevate ethylene production in both species, whereas application of AVG alone was shown to suppress it. Thus, ethylene may be considered an active agent of flowering inhibition brought about by IAA application.