TGF-β mediates early angiogenesis and latent fibrosis in an Emilin1-deficient mouse model of aortic valve disease

Aortic valve disease (AVD) is characterized by elastic fiber fragmentation (EFF), fibrosis and aberrant angiogenesis. Emilin1 is an elastin-binding glycoprotein that regulates elastogenesis and inhibits TGF-β signaling, but the role of Emilin1 in valve tissue is unknown. We tested the hypothesis that Emilin1 deficiency results in AVD, mediated by non-canonical (MAPK/phosphorylated Erk1 and Erk2) TGF-β dysregulation. Using histology, immunohistochemistry, electron microscopy, quantitative gene expression analysis, immunoblotting and echocardiography, we examined the effects of Emilin1 deficiency (Emilin1^−/−) in mouse aortic valve tissue. Emilin1 deficiency results in early postnatal cell-matrix defects in aortic valve tissue, including EFF, that progress to latent AVD and premature death. The Emilin1^−/− aortic valve displays early aberrant provisional angiogenesis and late neovascularization. In addition, Emilin1^−/− aortic valves are characterized by early valve interstitial cell activation and proliferation and late myofibroblast-like cell activation and fibrosis. Interestingly, canonical TGF-β signaling (phosphorylated Smad2 and Smad3) is upregulated constitutively from birth to senescence, whereas non-canonical TGF-β signaling (phosphorylated Erk1 and Erk2) progressively increases over time. Emilin1 deficiency recapitulates human fibrotic AVD, and advanced disease is mediated by non-canonical (MAPK/phosphorylated Erk1 and Erk2) TGF-β activation. The early manifestation of EFF and aberrant angiogenesis suggests that these processes are crucial intermediate factors involved in disease progression and therefore might provide new therapeutic targets for human AVD.KEY WORDS: Elastic fibers, Extracellular matrix, Aortic valve, Fibrosis, Angiogenesis     

Profile of toxigenicity of Clostridium difficile strains isolated from paediatric patients with clinical diagnosis of antibiotic associated diarrhea (AAD)

This study was performed to determine profile of toxigenicity of 18 Clostridium difficile strains isolated from paeditric patients suffering from antibiotic associated diarrhea (AAD). Toxigenicity of C. difficile strains was tested for detection toxin A and toxin B by phenotypic methods and for detection of the tcdA and tcdB genes using of PCR. Changes in the repeating regions of the tcdA genes were detected with the NK9/NKV011 primer pairs. For detection of binary toxin (CDT) cdtA and cdtB genes, cdtApos/cdtArev i cdtBpos/cdtBrev two pair primers in PCR was used. Among C. difficile strains was detected three profiles of toxigenicity: C. difficile strains possesing of tcdA and tcdB genes but not possesing cdtA and cdtB genes of binary toxin (A+B+CDT-), strains possesing tcdA and tcdB and cdtA and cdtB genes (A+B+CDT+), strains with deletion of toxin A gene (A-B+CDT-). This is the first report on the occurence of binary positive C. difficile strains isolated from paediatric patients.     

Polar lipid and fatty acid profiles – Re-vitalizing old approaches as a modern tool for the classification of mycoplasmas?

A set of 20 Mollicutes strains representing different lines of descent, including the type species of the genus Mycoplasma, Mycoplasma mycoides, Acholeplasma laidlawii and a strain of Mesoplasma, were subjected to polar lipid and fatty acid analyses in order to evaluate their suitability for classification purposes within members of this group. Complex polar lipid and fatty acid profiles were detected for each examined strain. All strains contained the polar lipids phosphocholine-6'-alpha-glucopyranosyl-(1'-3)-1, 2-diacyl-glycerol (MfGL-I), 1-O-alkyl/alkenyl-2-O-acyl-glycero-3-phosphocholine (MfEL), sphingomyelin (SphM), 1-O-alkyl/alkenyl-glycero-3-phosphocholine (lysoMfEL), the unknown aminophospholipid APL1 and the cholesterol Chol2. A total of 19 strains revealed the presence of phosphatidylethanolamine (PE) and/or phosphatidylglycerol (PG), and the presence of diphosphatidylglycerol (DPG) was detected in 13 strains. The unknown aminolipid AL1 was found in the extracts of 17 strains. Unbranched saturated and unsaturated compounds predominated in the fatty acid profiles. Major fatty acids were usually C16:0, C18:0, C18:1 omega9c and 'Summed feature 5' (C18:2 omega6, 9c/C18:0 anteiso). Our results demonstrated that members of the M. mycoides cluster showed rather homogenous polar lipid and fatty acid profiles. In contrast, each of the other strains was characterized by a unique polar lipid profile and significant quantitative differences in the presence of certain fatty acids. These results indicate that analyses of both polar lipid and fatty acid profiles could be a useful tool for classification of mycoplasmas.     

Effects of prey mite species on life history of the phytoseiid predators Typhlodromalus manihoti and Typhlodromalus aripo

The effects of prey mite suitability on several demographic characteristics of phytoseiid predators and the relationship of these effects to the potential of phytoseiid predators to control herbivorous mite populations are well documented. Evidence has also accumulated in the last 20 years demonstrating that phytoseiid predators utilize herbivorous prey mite-induced plant volatiles as olfactory cues in locating their herbivorous mite prey, but less well established is the predictability of reproductive success from the ability of the predators to utilize olfactory cues to locate their prey, and how these processes are related to the success of the predators as biological control agents of the herbivorous mite. In this study, we determined in laboratory no choice experiments, the development, survivorship and fecundity of the two neotropical phytoseiid predators Typhlodromalus manihoti Moraes and T. aripo DeLeon when feeding on three herbivorous mites, including the key prey species Mononychellus tanajoa (Bondar), and the two alternative prey species Oligonychus gossypii (Zacher) and Tetranychus urticae (Koch). Intrinsic rate of increase (r_m) of T. aripo was 2.1 fold higher on M. tanajoa as prey compared with T. urticae as prey, while it was almost nil on O. gossypii. For T. manihoti, r_m was 2.3 fold higher on M. tanajoa as prey compared with O. gossypii as prey, while reproduction was nil on T. urticae. An independent experiment on odor-related prey preference of the two predator species showed that T. manihoti and T. aripo preferred odors from M. tanajoa-infested leaves to odors from O. gossypii-infested leaves. Moreover, both predator species preferred odors from M. tanajoa-infested leaves over those from T. urticae-infested leaves. As reported here, life history of the two predatory mites matches odor-related prey preference if the key prey species is compared to the two inferior prey species. The implications of our findings for the persistence of T. manihoti and T. aripo and biological control of M. tanajoa in the cassava agroecosystem in Africa are discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

Insights into virus evolution and membrane biogenesis from the structure of the marine lipid-containing bacteriophage PM2

Recent, primarily structural observations indicate that related viruses, harboring no sequence similarity, infect hosts of different domains of life. One such clade of viruses, defined by common capsid architecture and coat protein fold, is the so-called PRD1-adenovirus lineage. Here we report the structure of the marine lipid-containing bacteriophage PM2 determined by crystallographic analyses of the entire approximately 45 MDa virion and of the outer coat proteins P1 and P2, revealing PM2 to be a primeval member of the PRD1-adenovirus lineage with an icosahedral shell and canonical double beta barrel major coat protein. The view of the lipid bilayer, richly decorated with membrane proteins, constitutes a rare visualization of an in vivo membrane. The viral membrane proteins P3 and P6 are organized into a lattice, suggesting a possible assembly pathway to produce the mature virus.     

CYP7B1-mediated metabolism of 5α-androstane-3α,17β-diol (3α-Adiol): A novel pathway for potential regulation of the cellular levels of androgens and neurosteroids

The current study presents data indicating that 5α-androstane-3α,17β-diol (3α-Adiol) undergoes a previously unknown metabolism into hydroxymetabolites, catalyzed by CYP7B1. 3α-Adiol is an androgenic steroid which serves as a source for the potent androgen dihydrotestosterone and also can modulate gamma-amino butyric acid A (GABA_A) receptor function in the brain. The steroid hydroxylase CYP7B1 is known to metabolize cholesterol derivatives, sex hormone precursors and certain estrogens, but has previously not been thought to act on androgens or 3α-hydroxylated steroids. 3α-Adiol was found to undergo NADPH-dependent metabolism into 6- and 7-hydroxymetabolites in incubations with porcine microsomes and human kidney-derived HEK293 cells, which are high in CYP7B1 content. This metabolism was suppressed by addition of steroids known to be metabolized by CYP7B1. In addition, 3α-Adiol significantly suppressed CYP7B1-mediated catalytic reactions, in a way as would be expected for substrates that compete for the same enzyme. Recombinant expression of human CYP7B1 in HEK293 cells significantly increased the rate of 3α-Adiol hydroxylation. Furthermore, the observed hydroxylase activity towards 3α-Adiol was very low or undetectable in livers of Cyp7b1(?/?) knockout mice. The present results indicate that CYP7B1-mediated catalysis may play a role for control of the cellular levels of androgens, not only of estrogens. These findings suggest a previously unknown mechanism for metabolic elimination of 3α-Adiol which may impact intracellular levels of dihydrotestosterone and GABA_A-modulating steroids.     

5-(2-Pyrimidinyl)-imidazo[1,2-a]pyridines are antibacterial agents targeting the ATPase domains of DNA gyrase and topoisomerase IV

Dual inhibitors of bacterial gyrB and parE based on a 5-(2-pyrimidinyl)-imidazo[1,2-a]pyridine template exhibited MICs (μg/mL) of 0.06–64 (Sau), 0.25–64 (MRSA), 0.06–64 (Spy), 0.06–64 (Spn), and 0.03–64 (FQR Spn). Selected examples were efficacious in mouse sepsis and lung infection models at <50 mg/kg (PO dosing).