The Rab4A effector protein Rabip4 is involved in migration of NIH 3T3 fibroblasts

The small GTP-binding protein Rab4 has been involved in the recycling of alphavbeta3 integrins in response to platelet-derived growth factor (PDGF) stimulation suggesting a role for Rab4 in cell adhesion and migration. In this study, we explored the role of Rabip4 and Rabip4', two Rab4 effector proteins, in migration of NIH 3T3 fibroblasts. In these cells, Rabip4 and Rabip4', collectively named Rabip4s, were partially co-localized with the early endosomal marker EEA1. PDGF treatment re-distributed endogenous Rabip4s toward the cell periphery where they colocalized with F-actin. In cells expressing green fluorescent protein (GFP)-Rabip4 or GFP-Rabip4', constitutive appearance of GFP-Rabip4s at the cell periphery was accompanied by local increase in cortical F-actin in membrane ruffles at the leading edge. The expression of GFP-Rabip4 induced an increased migration compared with control cells expressing GFP alone, even in the absence of PDGF stimulation. On the contrary, in cells expressing a mutated form of Rabip4s unable to interact with Rab4, lack of typical leading edge was observed. Furthermore, PDGF treatment did not stimulate the migration of these cells. Furthermore, down-regulation of the expression of Rabip4s inhibited PDGF-stimulated cell migration. Endogenous Rabip4s were localized with alphav integrins at the leading edge following PDGF treatment, whereas in cells expressing GFP-Rabip4s, alphav integrins, together with GFP-Rabip4s, were constitutively localized at the leading edge. In contrast, reduction in Rabip4s expression levels using small interfering RNA was associated with impaired PDGF-induced translocation of alphav integrins toward the leading edge. Taken together, our data provide evidence that Rabip4s, possibly via their interaction with Rab4, regulate integrin trafficking and are involved in the migration of NIH 3T3 fibroblasts.     

Combinations of Genetic Data Present in Bipolar Patients,but Absent in Control Persons

The main objective of the study was to find combinations of genetic variants significantly associated with bipolar disorder. In a previous study of bipolar disorder, combinations of three single nucleotide polymorphism (SNP) genotypes taken from 803 SNPs were analyzed, and four clusters of combinations were found to be significantly associated with bipolar disorder. In the present study, combinations of four SNP genotypes taken from the same 803 SNPs were analyzed, and one cluster of combinations was found to be significantly associated with bipolar disorder. Combinations from the new cluster and from the four previous clusters were identified in the genomes of 209 of the 607 patients in the study whereas none of the 1355 control participants had any of these combinations in their genome.     

Chronic Social Isolation Compromises the Activity of Both Glutathione Peroxidase and Catalase in Hippocampus of Male Wistar Rats

Chronic neuroendocrine stress usually leads to the elevation of the stress hormones and increased metabolic rate, which is frequently accompanied by oxidative damage to the CNS. In the present study we hypothesized that chronic psychosocial isolation (CPSI) of male Wistar rats, characterized by decreased serum corticosterone (CORT), unaltered catecholamines (CTs), and low blood glucose (GLU), may also promote oxidative imbalance in the CNS, by targeting antioxidant defense system. To test it, we have examined the relation between these input signals and protein expression/activity of antioxidant enzymes (AOEs): superoxide dismutases (SODs), catalase (CAT), glutathione peroxidase (GPx), and glutathione reductase (GLR) in the hippocampus (HIPPO) of CPSI animals. We found that CPSI did not affect SODs or CAT, but decreased activity of GPx and compromised GLR, an enzyme highly dependent on blood GLU for its substrate precursor. Further, we have tested whether the CPSI experience altered AOEs response to a novelty stress, and found that it attenuated peroxide-metabolizing enzymes, CAT and GPx, and decreased GLR activity, even though blood GLU was restored. The altered ratios of hippocampal AOEs in CPSI animals, which were worsened under the combined stress conditions, may lead to the accumulation of peroxide products and oxidative imbalance. The mechanism by which CPSI generate oxidative imbalance in the HIPPO is most likely based on poor systemic energy conditions set by this stress. Such conditions may cause functional decline of CNS structures, such as HIPPO, and are likely to promote state linked to onset of many mood disorders.     

Barks of Three Wild Pyrus Taxa: Phenolic Constituents,Antioxidant Activity,and in Vitro and in Silico Investigations of α-Amylase and α-Glucosidase Inhibition

Dry MeOH extracts of the twig barks of Pyrus communis subsp. pyraster, P. spinosa and their hybrid P.×jordanovii nothosubsp. velenovskyi, collected in wild in Serbia, were analyzed. By LC/MS, the contents of arbutin (99.9–131.0 mg/g), chlorogenic acid (2.2–6.3 mg/g), catechin (1.0–5.3 mg/g) and total dimeric and trimeric procyanidins (42.2–61.3 mg/g), including procyanidin B2 (8.9–17.2 mg/g), were determined. Colorimetrically, high contents of total phenolics (436.2–533.4 mg GAE/g) and tannins (339.4–425.7 mg GAE/g), as well as strong total antioxidant activities (FRAP values 4.5–5.9 mmol Fe²⁺/g), and DPPH (SC₅₀=6.6–7.1 μg/ml) and hydroxyl radical (SC₅₀=447.1–727.7 μg/ml) scavenging abilities were revealed. In vitro, all extracts exhibited notable inhibition of α-amylase (IC₅₀=310.8–617.7 μg/ml) and particularly strong inhibition of α-glucosidase (IC₅₀=2.1–3.7 μg/ml). Molecular docking predicted that among identified compounds procyanidin B2 is the best inhibitor of these carbohydrate-digesting enzymes. Obtained results showed that the barks of investigated Pyrus hybrid and its parent taxa have similar composition and bioactivity.     

Quality Assessment of Bee Pollen-Honey Mixtures Using Thin-Layer Chromatography in Combination with Chemometrics

The aim of this study is to develop a rapid, effect-directed screening method for quality assessment of bee pollen-honey mixtures. The comparative antioxidant potential and phenolic content of honey, bee pollen, and the bee pollen-honey mixtures, was performed using spectrophotometry. The total phenolic content and antioxidative activity of bee pollen-honey mixtures with 20 % bee pollen share were in the range 3.03–3.11 mg GAE/g, and 6.02–6.96 mmol TE/kg, respectively, while mixtures with 30 % bee pollen share contained 3.92–4.18 mg GAE/g, and 9.69–10.11 mmol TE/kg. Chromatographic fingerprint of bee pollen-honey mixtures was performed by high-performance thin-layer chromatography with conditions developed by authors and reported for the first time. Fingerprint analysis hyphenated with chemometrics enabled authenticity assessments of honey in mixtures. Results indicate that bee pollen-honey mixtures represent a food with highly, both, nutritious characteristics and health-promoting effect.     

A microsatellite-based genetic linkage map of the waterflea, Daphnia pulex: On the prospect of crustacean genomics

We describe the first genetic linkage map for Daphnia pulex using 185 microsatellite markers, including 115 new markers reported in this study. Our approach was to study the segregation of polymorphisms in 129 F2 progeny of one F1 hybrid obtained by crossing two genetically divergent lineages of Daphnia isolated from two Oregon populations. The map spanned 1206 Kosambi cM and had an average intermarker distance of 7 cM. Linkage groups ranged in size from 7 to 185 cM and contained 4 to 27 markers. The map revealed 12 linkage groups corresponding to the expected number of chromosomes and covers approximately 87% of the genome. Tests for random segregation of alleles at individual loci revealed that 21% of the markers showed significant transmission ratio distortion (primarily homozygote deficiency) likely due to markers being linked to deleterious recessive alleles. This map will become the anchor for the physical map of the Daphnia genome and will serve as a starting point for mapping single and quantitative trait loci affecting ecologically important phenotypes. By mapping 342 tentative orthologous gene pairs (Daphnia/Drosophila) into the Daphnia linkage map, we facilitate future comparative projects.     

Subspecies-specific distribution of intervening sequences in the Bacillus subtilis prophage ribonucleotide reductase genes

A collection of 212 gram-positive bacilli isolated from natural habitats was screened for the presence of intervening sequences (introns and intein-coding sequences) in the SPbeta prophage-related ribonucleotide reductase genes bnrdE and bnrdF. Three novel configurations were identified on the basis of the presence of (i) intervening sequences in bnrdE and bnrdF, and (ii) an ORF in the bnrdE-bnrdF spacer. Analysis of the cell wall genetic determinants as well as of the incorporation of radio-labelled glycerol into cell wall allowed newly and previously identified B. subtilis strains with different configurations of bnrdE/bnrdF intervening sequences to be assigned to one of two subspecies. Strains apparently belonging to the subsp. subtilis contain three intervening sequences many of which are associated with the putative homing endonuclease activity. Strains of the subsp. spizizenii contain only one or two ORF-less group I introns. Introns occupying bnrdF are confined to the subspecies subtilis.