Vitamin D receptor BsmI polymorphism modulates soy intake and 25-hydroxyvitamin D supplementation benefits in cardiovascular disease risk factors profile

Vitamin D receptor polymorphisms may predispose that not all individuals could have benefits from the nutritional supplementation of 25-hydroxyvitamin D. Furthermore, vitamin D-related cardiovascular effects may also be influenced by soy isoflavones considered endocrine regulators of cardiovascular homeostasis. To find possible gene–diet interactions by evaluating individualized lipid metabolism benefits from an increase in soy and 25-hydroxyvitamin D intake, 106 healthy individuals, genotyped for vitamin D receptor (VDR) gene polymorphism rs1544410 (BsmI) were randomly assigned to either no intake, to daily 250?mL or 500?mL of a 25-hydroxyvitamin D supplemented SB for 2 months. The soybean beverage induced differences in cardiovascular risk factors (lipid profile, blood pressure, TNFα and MCP-1), as well as vitamin D metabolites in a dose-gene-dependent relation. Thus, VDR BsmI polymorphism affected individual response being the GG genotype the ones that showed dose-dependent manner responsiveness in the reduction in total cholesterol, LDL and triglycerides in comparison with the AA/AG genotype. These differences were associated with increased plasma levels of 1α,25-dyhydroxyvitamin D3 in the carriers of the GG genotype. It was concluded that metabolic response to 25-hydroxyvitamin D and soybean supplementation is dependent on VDR BsmI GG genotype due to a higher conversion rate from vitamin D precursors.     

Formation of S-(carboxymethyl)-cysteine in rat liver mitochondrial proteins: effects of caloric and methionine restriction

Maillard reaction contributes to the chemical modification and cross-linking of proteins. This process plays a significant role in the aging process and determination of animal longevity. Oxidative conditions promote the Maillard reaction. Mitochondria are the primary site of oxidants due to the reactive molecular species production. Mitochondrial proteome cysteine residues are targets of oxidative attack due to their specific chemistry and localization. Their chemical, non-enzymatic modification leads to dysfunctional proteins, which entail cellular senescence and organismal aging. Previous studies have consistently shown that caloric and methionine restrictions, nutritional interventions that increase longevity, decrease the rate of mitochondrial oxidant production and the physiological steady-state levels of markers of oxidative damage to macromolecules. In this scenario, we have detected S-(carboxymethyl)-cysteine (CMC) as a new irreversible chemical modification in mitochondrial proteins. CMC content in mitochondrial proteins significantly correlated with that of the lysine-derived analog N ^ε-(carboxymethyl)-lysine. The concentration of CMC is, however, one order of magnitude lower compared with CML likely due in part to the lower content of cysteine with respect to lysine of the mitochondrial proteome. CMC concentrations decreases in liver mitochondrial proteins of rats subjected to 8.5 and 25 % caloric restriction, as well as in 40 and 80 % methionine restriction. This is associated with a concomitant and significant increase in the protein content of sulfhydryl groups. Data presented here evidence that CMC, a marker of Cys-AGE formation, could be candidate as a biomarker of mitochondrial damage during aging.     

T‐type calcium channel blockers inhibit autophagy and promote apoptosis of malignant melanoma cells

We have recently reported that human melanoma cells express a variety of voltage‐gated calcium (Ca²⁺) channel types, including low‐voltage‐activated T‐type channels that play a significant role in melanoma cell cycle progression. Here, we challenged melanoma metastatic cells with T‐type channel blockers of clinical use and found a dual effect on cell viability: (i) a reduction in the proliferation rate, through a halt in the progression to the G₁‐S phase; and (ii) a promotion of cell death that was partially dependent on the activation of caspases. An in‐depth analysis of the death process showed that the apoptotic pathway is preceded by endoplasmic reticulum stress and the subsequent inhibition of the basal macroautophagy which is active in these cells. The effects of pharmacological blockers on Ca²⁺ homeostasis, autophagy, and cell death were mimicked by T‐type channel gene silencing. These results provide the basis for a new pharmacological and/or gene silencing approach toward tackling melanoma metastasis.     

Aging increases Nepsilon-(carboxymethyl)lysine and caloric restriction decreases Nepsilon-(carboxyethyl)lysine and Nepsilon-(malondialdehyde)lysine in rat heart mitochondrial proteins

The present investigation studies the effect of aging, short-term and long-term caloric restriction on four different markers of oxidative, glycoxidative or lipoxidative damage to heart mitochondrial proteins: protein carbonyls (measured by ELISA); N epsilon -(carboxyethyl)lysine (CEL), N epsilon -(carboxymethyl)lysine (CML), and N epsilon -(malondialdehyde)lysine (MDA-lys) measured by gas chromatography/mass spectrometry. Aging increased the steady state level of CML in rat heart mitochondria without changing the levels of the other three markers of protein damage. Short-term caloric restriction (six weeks) did not change any of the parameters measured. However, long-term (one year) caloric restriction decreased CEL and MDA-lys in heart mitochondria and did not change protein carbonyls and CML levels. The decrease in MDA-lys was not due to changes in the sensitivity of mitochondrial lipids to peroxidation since the measurements of the fatty acid composition showed that the total number of fatty acid double bonds was not changed by caloric restriction. The decrease in CEL and MDA-lys in caloric restriction agrees with the previously and consistently described finding that caloric restriction agrees with the previously and consistently described finding that caloric restriction lowers the rate of generation of reactive oxygen species (ROS) in rodent heart mitochondria, although in the case of CEL a caloric restriction-induced lowering of glycaemia can also be involved. The CEL and MDA-lys results support the notion that caloric restriction decreases oxidative stress-derived damage to heart mitochondrial proteins.     

The buckling of spherical liposomes

In the classical "first approximation" theory of thin-shell structures, the constitutive relations for a generic shell element--i.e. the elastic relations between the bending moments and membrane stresses and the corresponding changes in curvature and strain, respectively-are written as if an element of the shell is flat, although in reality it is curved. In this theory it is believed that discrepancies on account of the use of "flat" constitutive relations will be negligible provided the ratio shell-radius/thickness is of sufficiently large order. In the study of drawing of narrow, cylindrical "tethers" from liposomes it has been known for many years that it is necessary to use instead a constitutive law which explicitly describes a curved element in order to make sense of the mechanics; and indeed such tethers are generally of "thick-walled" proportions. In this paper we show that the proper constitutive relations for a curved element must also be used in the study, by means of shell equations, of the buckling of initially spherical thin-walled giant liposomes under exterior pressure: these involve the inclusion of what we call the "Mkappa" terms, which are not present in the standard "first-approximation" theory. We obtain analytical expressions for both the bifurcation buckling pressure and the slope of the post-buckling path, in terms of the dimensions and elastic constants of the lipid bi-layer, and also the initial state of bending moment in the vesicle. We explain physically how the initial bending moment can affect the bifurcation pressure, whereas it cannot in "first-approximation" theory. We use these results to map the conditions under which the vesicle buckles into an oblate, as distinct from a prolate ("rugby-ball") shape. Some of our results were obtained long ago by the use of energy methods; but our aim here has been to identify precisely what is lacking in "first-approximation" theory in relation to liposomes, and so to put the "shell equations" approach onto a firm footing in mechanics.     

Proteins in human brain cortex are modified by oxidation, glycoxidation, and lipoxidation. Effects of Alzheimer disease and identification of lipoxidation targets

Diverse oxidative pathways, such as direct oxidation of amino acids, glycoxidation, and lipoxidation could contribute to Alzheimer disease pathogenesis. A global survey for the amount of structurally characterized probes for these reactions is lacking and could overcome the lack of specificity derived from measurement of 2,4-dinitrophenylhydrazine reactive carbonyls. Consequently we analyzed (i) the presence and concentrations of glutamic and aminoadipic semialdehydes, N(epsilon)-(carboxymethyl)-lysine, N(epsilon)-(carboxyethyl)-lysine, and N(epsilon)-(malondialdehyde)-lysine by means of gas chromatography/mass spectrometry, (ii) the biological response through expression of the receptor for advanced glycation end products, (iii) the fatty acid composition in brain samples from Alzheimer disease patients and age-matched controls, and (iv) the targets of N(epsilon)-(malondialdehyde)-lysine formation in brain cortex by proteomic techniques. Alzheimer disease was associated with significant, although heterogeneous, increases in the concentrations of all evaluated markers. Alzheimer disease samples presented increases in expression of the receptor for advanced glycation end products with high molecular heterogeneity. Samples from Alzheimer disease patients also showed content of docosahexaenoic acid, which increased lipid peroxidizability. In accordance, N(epsilon)-(malondialdehyde)-lysine formation targeted important proteins for both glial and neuronal homeostasis such as neurofilament L, alpha-tubulin, glial fibrillary acidic protein, ubiquinol-cytochrome c reductase complex protein I, and the beta chain of ATP synthase. These data support an important role for lipid peroxidation-derived protein modifications in Alzheimer disease pathogenesis.     

Effect of experimental and cold exposure induced hyperthyroidism on H2O2 production and susceptibility to oxidative stress of rat liver mitochondria

To investigate the iodothyronine role in liver responses to cold, we examined metabolic and oxidative mitochondrial changes in cold-exposed, T3-treated, and T4-treated rats, which exhibit different T4 serum levels. All treatments increased mitochondrial respiration which reached the highest and lowest values after T3 and cold treatment, respectively. The T3- and T4-induced changes agreed with the respective increases in Complex IV activities, while those elicited by cold were inconsistent with increased activities of respiratory complexes. Mitochondrial capacity to produce H2O2 was the highest in T3-treated rats, whereas it was similar in T4-treated and cold-exposed rats. The effects of respiratory inhibitors suggested that T3 and T4 mainly increase the mitochondrial content of autoxidizable electron carrier of Complex I and Complex III, respectively. The indices of oxidative modifications of proteins exhibited increases consistent with the treatment effects on H2O2 production. The increases in indices of lipid peroxidation were also dependent on changes in lipid composition. The increased protein damage in treatment groups was confirmed using immunoblotting analysis, which also showed oxidative damage in a 133 kDa fraction, which was not expressed in T3-treated rats. Antioxidant levels were not related to the extent of oxidative damage as only mitochondrial GSH levels decreased in T3-treated rats. Mitochondrial susceptibility to in vitro oxidative challenge and Ca2+-induced swelling was increased by all treatments, but was the highest in T3-treated rats. In the whole, our results indicate T3 as main responsible for the changes in the mitochondrial population associated with cold exposure. However, a significant role is also played by T4, which appears to acts mainly modulating T3 effects, but also inducing some effects different from the T3 ones.     

Do pollination syndromes cause modularity and predict interactions in a pollination network in tropical high‐altitude grasslands?

The concept of pollination syndromes has been widely questioned, since plant–pollinator interactions have proved to be more generalist than was previously thought. We examined whether the network of a tropical high‐altitude grassland contained groups of plants and pollinators that interact preferentially with each other. A general binary matrix was created. To assess the robustness of myophily, in all analyses we considered: 1) the whole network, 2) the network after the wasps were removed, and 3) the network after the flies were removed. For each network we evaluated whether: 1) the observed interactions were more related to syndromes than expected by chance, compared to an expected matrix; 2) there was a modular structure; 3) the modules found were more related to syndromes than expected by chance, compared to another expected matrix; 4) the syndromes were equally robust. For this analysis, the general matrix was subdivided into smaller matrices that included each pollination syndrome separately. To test the influence of the functional groups of pollinators and the phylogeny of plants, in addition to the general matrix, we also considered the first expected matrix, a quantitative functional group and a plant phylogeny matrix. The pollination syndromes determined the pattern of interactions in the network: 69% of the total interactions resulted from the functional group of pollinators predicted by the plant syndrome. The network showed greater modularity (13 modules) than expected by chance, mostly consisting of the expected functional groups of pollinators and plant syndromes. The modules were associated with pollination syndromes more than was predicted by chance. Most of the variation in interactions was explained by functional groups of pollinators or by plant syndromes. Plant phylogeny did not account for a significant amount of variation in the interactions. Our findings support the concept of pollination syndromes. However, the interactions were not equally predicted by different pollination syndromes, and the accuracy of the prediction was strongest for ornithophily and melittophily.     

Functional expression of voltage-gated calcium channels in human melanoma

The expression of voltage-gated calcium channels (VGCCs) has not been reported previously in melanoma cells in spite of increasing evidence of a role of VGCCs in tumorigenesis and tumour progression. To address this issue we have performed an extensive RT-PCR analysis of VGCC expression in human melanocytes and a range of melanoma cell lines and biopsies. In addition, we have tested the functional expression of these channels using Ca(2+) imaging techniques and examined their relevance for the viability and proliferation of the melanoma cells. Our results show that control melanocytes and melanoma cells express channel isoforms belonging to the Ca(v) 1 and Ca(v) 2 gene families. Importantly, the expression of low voltage-activated Ca(v) 3 (T-type) channels is restricted to melanoma. We have confirmed the function of T-type channels as mediators of constitutive Ca(2+) influx in melanoma cells. Finally, pharmacological and gene silencing approaches demonstrate a role for T-type channels in melanoma viability and proliferation. These results encourage the analysis of T-type VGCCs as targets for therapeutic intervention in melanoma tumorigenesis and/or tumour progression.     

HP1-β is required for development of the cerebral neocortex and neuromuscular junctions

HP1 proteins are thought to be modulators of chromatin organization in all mammals, yet their exact physiological function remains unknown. In a first attempt to elucidate the function of these proteins in vivo, we disrupted the murine Cbx1 gene, which encodes the HP1-β isotype, and show that the Cbx1^−/−-null mutation leads to perinatal lethality. The newborn mice succumbed to acute respiratory failure, whose likely cause is the defective development of neuromuscular junctions within the endplate of the diaphragm. We also observe aberrant cerebral cortex development in Cbx1^−/− mutant brains, which have reduced proliferation of neuronal precursors, widespread cell death, and edema. In vitro cultures of neurospheres from Cbx1^−/− mutant brains reveal a dramatic genomic instability. Our results demonstrate that HP1 proteins are not functionally redundant and that they are likely to regulate lineage-specific changes in heterochromatin organization.