Associations of Whole Blood n-3 and n-6 Polyunsaturated Fatty Acids with Blood Pressure in Children and Adolescents – Results from the IDEFICS/I.Family Cohort

BackgroundPolyunsaturated n-3 and n-6 polyunsaturated fatty acids (PUFA) are precursors of biologically active metabolites that affect blood pressure (BP) regulation. This study investigated the association of n-3 and n-6 PUFA and BP in children and adolescents.MethodsIn a subsample of 1267 children aged 2–9 years at baseline of the European IDEFICS (Identification and prevention of dietary- and lifestyle-induced health effects in children and infants) cohort whole blood fatty acids were measured by a validated gas chromatographic method. Systolic and diastolic BP was measured at baseline and after two and six years. Mixed-effects models were used to assess the associations between fatty acids at baseline and BP z-scores over time adjusting for relevant covariables. Models were further estimated stratified by sex and weight status.ResultsThe baseline level of arachidonic acid was positively associated with subsequent systolic BP (β = 0.08, P = 0.002) and diastolic BP (β = 0.07, P<0.001). In thin/normal weight children, baseline alpha-linolenic (β = -1.13, P = 0.003) and eicosapentaenoic acid (β = -0.85, P = 0.003) levels were inversely related to baseline and also to subsequent systolic BP and alpha-linolenic acid to subsequent diastolic BP. In overweight/obese children, baseline eicosapentaenoic acid level was positively associated with baseline diastolic BP (β = 0.54, P = 0.005).ConclusionsLow blood arachidonic acid levels in the whole sample and high n-3 PUFA levels in thin/normal weight children are associated with lower and therefore healthier BP. The beneficial effects of high n-3 PUFA on BP were not observed in overweight/obese children, suggesting that they may have been overlaid by the unfavorable effects of excess weight.     

Plasmid Characterization and Chromosome Analysis of Two netF+ Clostridium perfringens Isolates Associated with Foal and Canine Necrotizing Enteritis

The recent discovery of a novel beta-pore-forming toxin, NetF, which is strongly associated with canine and foal necrotizing enteritis should improve our understanding of the role of type A Clostridium perfringens associated disease in these animals. The current study presents the complete genome sequence of two netF-positive strains, JFP55 and JFP838, which were recovered from cases of foal necrotizing enteritis and canine hemorrhagic gastroenteritis, respectively. Genome sequencing was done using Single Molecule, Real-Time (SMRT) technology-PacBio and Illumina Hiseq2000. The JFP55 and JFP838 genomes include a single 3.34 Mb and 3.53 Mb chromosome, respectively, and both genomes include five circular plasmids. Plasmid annotation revealed that three plasmids were shared by the two newly sequenced genomes, including a NetF/NetE toxins-encoding tcp-conjugative plasmid, a CPE/CPB2 toxins-encoding tcp-conjugative plasmid and a putative bacteriocin-encoding plasmid. The putative beta-pore-forming toxin genes, netF, netE and netG, were located in unique pathogenicity loci on tcp-conjugative plasmids. The C. perfringens JFP55 chromosome carries 2,825 protein-coding genes whereas the chromosome of JFP838 contains 3,014 protein-encoding genes. Comparison of these two chromosomes with three available reference C. perfringens chromosome sequences identified 48 (~247 kb) and 81 (~430 kb) regions unique to JFP55 and JFP838, respectively. Some of these divergent genomic regions in both chromosomes are phage- and plasmid-related segments. Sixteen of these unique chromosomal regions (~69 kb) were shared between the two isolates. Five of these shared regions formed a mosaic of plasmid-integrated segments, suggesting that these elements were acquired early in a clonal lineage of netF-positive C. perfringens strains. These results provide significant insight into the basis of canine and foal necrotizing enteritis and are the first to demonstrate that netF resides on a large and unique plasmid-encoded locus.     

Inhibiting the urokinase‐type plasminogen activator receptor system recovers STZ‐induced diabetic nephropathy

The urokinase‐type plasminogen activator (uPA) receptor (uPAR) participates to the mechanisms causing renal damage in response to hyperglycaemia. The main function of uPAR in podocytes (as well as soluble uPAR ‐(s)uPAR‐ from circulation) is to regulate podocyte function through αvβ3 integrin/Rac‐1. We addressed the question of whether blocking the uPAR pathway with the small peptide UPARANT, which inhibits uPAR binding to the formyl peptide receptors (FPRs) can improve kidney lesions in a rat model of streptozotocin (STZ)‐induced diabetes. The concentration of systemically administered UPARANT was measured in the plasma, in kidney and liver extracts and UPARANT effects on dysregulated uPAR pathway, αvβ3 integrin/Rac‐1 activity, renal fibrosis and kidney morphology were determined. UPARANT was found to revert STZ‐induced up‐regulation of uPA levels and activity, while uPAR on podocytes and (s)uPAR were unaffected. In glomeruli, UPARANT inhibited FPR2 expression suggesting that the drug may act downstream uPAR, and recovered the increased activity of the αvβ3 integrin/Rac‐1 pathway indicating a major role of uPAR in regulating podocyte function. At the functional level, UPARANT was shown to ameliorate: (a) the standard renal parameters, (b) the vascular permeability, (c) the renal inflammation, (d) the renal fibrosis including dysregulated plasminogen‐plasmin system, extracellular matrix accumulation and glomerular fibrotic areas and (e) morphological alterations of the glomerulus including diseased filtration barrier. These results provide the first demonstration that blocking the uPAR pathway can improve diabetic kidney lesion in the STZ model, thus suggesting the uPA/uPAR system as a promising target for the development of novel uPAR‐targeting approaches.     

Clinanthus microstephium,an Amaryllidaceae Species with Cholinesterase Inhibitor Alkaloids: Structure−Activity Analysis of Haemanthamine Skeleton Derivatives

Plants of the Amaryllidaceae family are well‐known (not only) for their ornamental value but also for the alkaloids that they produce. In this report, the first phytochemical study of Clinanthus genus was carried out. The chemical composition of alkaloid fractions from Clinanthus microstephium was analyzed by GC/MS and NMR. Seven known compounds belonging to three structural types of Amaryllidaceae alkaloids were identified. An epimeric mixture of a haemanthamine‐type compound (6‐hydroxymaritidine) was tested as an inhibitor against acetyl‐ and butyrylcholinesterase enzymes (AChE and BChE, respectively), two enzymes relevant in the treatment of Alzheimer's disease, with good results. Structure–activity relationships through molecular docking studies with this alkaloid and other structurally related compounds were discussed.     

Efecto inhibitorio del dietilestilbestrol sobre aislamientos clínicos de Candida albicans sensibles y resistentes a los azoles

BackgroundCandida albicans is a microorganism frequently involved in several infections; the patient's oral cavity, caries niches or periodontal disease can sometimes be the reservoir.. The fungal resistance to the available treatments, among other reasons, has led to the search for new antifungal alternatives.AimsTo carry out a comparative study of the in vitro effects of diethylstilboestrol (DES) and fluconazole (FLZ) on the growth of clinical strains of C. albicans.MethodsSeven strains of C. albicans were used: a) one FLZ-sensitive culture collection strain, ATCC 90028 (ATCC); b) four oral isolates from four oncological patients with periodontal disease (period 8, 9, 10, and 11); and c) two oral isolates from an AIDS patient with oropharyngeal candidiasis: one FLZ- sensitive (2-76), and another FLZ- resistant (12-99). The MIC was evaluated by standard spectrophotometric techniques using the CLSI (M27-A3) guidelines. The inhibitory concentration 50% (IC₅₀) was calculated using functional analysis with the Graph Pad software.ResultsDES inhibited the growth of all C. albicans strains, whether sensitive or resistant to FLZ. Experimental data fitted non-linear functions of inhibitor concentration versus response. Minimum inhibitory concentrations (MIC) for DES and FLZ were as follows: 28.18 µg/ml and 4.90 µg/ml (ATCC); 17.16 µg/ml and 3.14 µg/ml (period); 27.64 µg/ml and 4.22 µg/ml (2-76); 6.16 µg/ml and 438.19 µg/ml (12-99), respectively.ConclusionsDES showed antifungal activity on all clinical C. albicans strains isolated from patients with dental and medical diseases. It showed the highest potency on the FLZ-resistant isolate.     

Human MICAL1: Activation by the small GTPase Rab8 and small‐angle X‐ray scattering studies on the oligomerization state of MICAL1 and its complex with Rab8

Human MICAL1 is a member of a recently discovered family of multidomain proteins that couple a FAD‐containing monooxygenase‐like domain to typical protein interaction domains. Growing evidence implicates the NADPH oxidase reaction catalyzed by the flavoprotein domain in generation of hydrogen peroxide as a second messenger in an increasing number of cell types and as a specific modulator of actin filaments stability. Several proteins of the Rab families of small GTPases are emerging as regulators of MICAL activity by binding to its C‐terminal helical domain presumably shifting the equilibrium from the free – auto‐inhibited – conformation to the active one. We here extend the characterization of the MICAL1–Rab8 interaction and show that indeed Rab8, in the active GTP‐bound state, stabilizes the active MICAL1 conformation causing a specific four‐fold increase of k_cat of the NADPH oxidase reaction. Kinetic data and small‐angle X‐ray scattering (SAXS) measurements support the formation of a 1:1 complex between full‐length MICAL1 and Rab8 with an apparent dissociation constant of approximately 8 μM. This finding supports the hypothesis that Rab8 is a physiological regulator of MICAL1 activity and shows how the protein region preceding the C‐terminal Rab‐binding domain may mask one of the Rab‐binding sites detected with the isolated C‐terminal fragment. SAXS‐based modeling allowed us to propose the first model of the free full‐length MICAL1, which is consistent with an auto‐inhibited conformation in which the C‐terminal region prevents catalysis by interfering with the conformational changes that are predicted to occur during the catalytic cycle.     

The histone-like protein HupB influences biofilm formation and virulence in Xanthomonas citri ssp. citri through the regulation of flagellar biosynthesis

Citrus canker is an important disease of citrus, whose causal agent is the bacterium Xanthomonas citri ssp. citri (Xcc). In previous studies, we found a group of Xcc mutants, generated by the insertion of the Tn5 transposon, which showed impaired ability to attach to an abiotic substrate. One of these mutants carries the Tn5 insertion in hupB, a gene encoding a bacterial histone-like protein, homologue to the β-subunit of the Heat-Unstable (HU) nucleoid protein of Escherichia coli. These types of protein are necessary to maintain the bacterial nucleoid organization and the global regulation of gene expression. Here, we characterized the influence of the mutation in hupB regarding Xcc biofilm formation and virulence. The mutant strain hupB was incapable of swimming in soft agar, whereas its complemented strain partially recovered this phenotype. Electron microscope imaging revealed that impaired motility of hupB was a consequence of the absence of the flagellum. Comparison of the expression of flagellar genes between the wild-type strain and hupB showed that the mutant exhibited decreased expression of fliC (encoding flagellin). The hupB mutant also displayed reduced virulence compared with the wild-type strain when they were used to infect Citrus lemon plants using different infection methods. Our results therefore show that the histone-like protein HupB plays an essential role in the pathogenesis of Xcc through the regulation of biofilm formation and biosynthesis of the flagellum.