Isolation of dengue serotype 3 virus from the cerebrospinal fluid of an encephalitis patient in Hai Phong, Vietnam in 2013.

Dengue encephalitis (DE) is characterized as unusual presentation of dengue infection. Despite the reports that DE accounts for only 1-5% of dengue cases, this disease tends to be increasingly reported to threaten global human health throughout dengue endemic areas particularly in Southeast Asia. The molecular information of clinically characterized, neurotropic dengue virus (DENV) in human beings is extremely scarce despite it playing an important role in deciphering the pathogenesis of dengue-related neurological cases. Here we report a case of DE caused by DENV3 genotype III in a male patient with atypical symptoms of DENV infection in Hai Phong, Vietnam in 2013. The virus isolated from the cerebrospinal fluid of this case-patient was closely related to DENV3 genotype III strains isolated from serum of two other patients, who manifested classical dengue in the same year and residing in the same area as the case-patient. It is noteworthy to mention that in 2013, DENV3 genotype III was detected for the first time in Vietnam.

17ß-Estradiol-induced interaction of ERα with NPPA regulates gene expression in cardiomyocytes.

AIMS: 17ß-Oestradiol (E2) and its receptors (ERα and ERß) are important regulators of physiological and pathological processes in the cardiovascular system. ER act in concert with other regulatory factors mediating oestrogenic effects. However, the underlying mechanisms modulating ER transcriptional activity are not fully elucidated. To gain better understanding of E2-induced ERα action in the human heart, we aimed to identify and functionally analyse interaction partners of ERα. METHODS AND RESULTS: Using yeast two-hybrid assays with a human heart cDNA library, we identified atrial natriuretic peptide precursor A (NPPA), a well-known cardiac hypertrophy marker, as a novel ERα interaction partner interacting in an E2-dependent manner. Mutation analyses and immunofluorescence data indicated that the LXXLL motif within NPPA is necessary for its E2-induced interaction with ERα, its action as a co-repressor of ERα, and its translocation into the nucleus of human and rat cardiomyocytes. Expression analysis and chromatin immunoprecipitation assays in a human left ventricular cardiomyocyte cell line, AC16, showed that NPPA interacts with E2/ERα, suppressing the transcriptional activity of ERα on E2-target genes, such as NPPA, connexin43, αactinin-2, nuclear factor of activated T-cells, and collagens I and III. CONCLUSION: We characterize for the first time an E2-regulated interaction of NPPA with ERα in cardiomyocytes, that may be crucial in physiological and/or pathological cardiac processes, thereby representing a potential therapeutic target.

17beta-Estradiol inhibits matrix metalloproteinase-2 transcription via MAP kinase in fibroblasts.

AIMS: Female sex and sex hormones contribute to cardiac remodelling. 17beta-estradiol (E2) is involved in the modulation of extracellular matrix composition and function. Here, we analysed the effect of E2 on matrix metalloproteinase (MMP)-2 gene expression and studied the underlying molecular mechanisms in rat cardiac fibroblasts and in a human fibroblast cell line. METHODS AND RESULTS: In adult rat cardiac fibroblasts, E2 significantly decreased MMP-2 gene expression in an estrogen receptor (ER)-dependent manner. Transient transfection experiments of human MMP-2 (hMMP-2) promoter deletion constructs in a human fibroblast cell line revealed a regulatory region between -324 and -260 bp that is involved in E2/ERalpha-mediated repression of hMMP-2 gene transcription. Electrophoretic mobility shift assays (EMSA) and supershift analysis demonstrated the binding of transcription factor Elk-1 within this promoter region. Elk-1 was phosphorylated by E2 via the mitogen-activated protein kinase (MAPK) signalling pathway as shown by western blotting. Treatment of cells with the MAPK inhibitor PD98059 blocked the E2-dependent repression of hMMP-2 promoter activity as well as the endogenous MMP-2 mRNA levels in both human fibroblast cells and rat cardiac fibroblasts. CONCLUSION: E2 inhibits MMP-2 expression via the ER and the MAPK pathway in rat cardiac fibroblasts and in a human fibroblast cell line. These mechanisms may contribute to sex-specific differences in fibrotic processes that are observed in human heart and other diseases.

The role of polyhydroxyalkanoate biosynthesis by Pseudomonas aeruginosa in rhamnolipid and alginate production as well as stress tolerance and biofilm formation.

Pseudomonas aeruginosa is capable of synthesizing polyhydroxyalkanoic acids (PHAs) and rhamnolipids, both of which are composed of 3-hydroxydecanoic acids connected by ester bonds, as well as synthesizing the biofilm matrix polymer alginate. In order to study the influence of PHA biosynthesis on rhamnolipid and alginate biosynthesis, as well as stress tolerance and biofilm formation, isogenic knock-out mutants deficient in PHA biosynthesis were generated for P. aeruginosa PAO1 and the alginate-overproducing P. aeruginosa FRD1. A gentamicin-resistance cassette was inserted replacing the 3' region of phaC1, the whole of phaZ and the 5' region of phaC2. Gas chromatography/mass spectrometry analysis showed that PHA accumulation was completely abolished in both strains. Interestingly, this gene replacement did not abolish rhamnolipid production. Thus, as previously suggested, the PHA synthase is not directly involved in rhamnolipid biosynthesis. In the PHA-negative mutant of mucoid FRD1 alginate biosynthesis was not affected, whereas in the PHA-negative PAO1 mutant an almost threefold increase in biosynthesis was observed compared to the wild-type. Consistently, PHA accumulation in FRD1 contributed only 4.7 % of cell dry weight, which is fourfold less than in PAO1. These data suggest that PHA biosynthesis and alginate biosynthesis are in competition with respect to a common precursor. The surface attachment and biofilm development of the PHA-negative mutants were also compared to those of wild-type strains in glass flow-cell reactors. PHA-negative mutants of P. aeruginosa PAO1 and FRD1 showed reduced attachment to glass. However, the PAO1 PHA-negative mutant, in contrast to the wild-type, formed a stable biofilm with large, distinct and differentiated microcolonies characteristic of alginate-overproducing strains of P. aeruginosa. The stress tolerance of PHA-negative mutants with respect to elevated temperature was strongly impaired. These data indicated a functional role for PHA in stress response and tolerance.