Shear stress-induced up-regulation of the intermediate-conductance Ca(2+)-activated K(+) channel in human endothelium

OBJECTIVE: Wall shear stress associated with blood flow is a major stimuli for generation of endothelial vasodilating and antithrombotic factors and it also regulates endothelial gene expression. Activation of endothelial intermediate-conductance Ca(2+)-activated K(+) channels (IK(Ca)) is important for the control of endothelial function by inducing cell hyperpolarization and thus generation of the endothelium-derived hyperpolarizing factor. In the present study we tested whether the IK(Ca) encoding IKCa1 gene is regulated by laminar shear stress (LSS). METHODS: Human umbilical vein endothelial cells (HUVEC) were subjected to LSS with a magnitude of 0.5-15 dyn/cm(2) and time intervals of 2-24 h in a flow cone apparatus. Expression of the IKCa1 gene and IK(Ca)-functions were determined by using real time RT-PCR and patch-clamp techniques. RESULTS: A short 2-4 h-or long 24 h-exposure to a LSS with a low (venous) magnitude of 0.5 dyn/cm(2) had no effect on IKCa1 expression levels. An exposure for 2 and 4 h to LSS with an intermediate magnitude of 5 dyn/cm(2) was also ineffective, whereas an exposure for 24 h induced a significant threefold up-regulation of IKCa1 expression levels. An exposure to LSS with a higher (arterial) magnitude of 15 dyn/cm(2), resulted in an eightfold up-regulation of IKCa1 expression levels after a 4 h-exposure and a fourfold increase of IKCa1 expression levels at 24 h. The increased IKCa1 expression levels following exposure to high levels of LSS resulted in enhanced IK(Ca) whole-cell currents and in an increased hyperpolarization of the endothelium in response to ATP and the IK(Ca) opener 1-EBIO. Inhibition of the mitogen-activated protein kinase/extracellular-signal-regulated kinase (ERK) kinase 1/2 (MEK/ERK) pathway by PD98059 prevented the LSS-induced up-regulation of IKCa1 expression levels and IK(Ca) whole-cell currents indicating that augmentation of IKCa1 expression levels is mediated by the LSS-induced activation of the MEK/ERK pathway. CONCLUSION: Long term exposure to LSS up-regulates expression and function of endothelial IK(Ca). This increase might represent a new important mechanism in endothelial adaptation to altered hemodynamics.     

NAD(P)H oxidase-dependent platelet superoxide anion release increases platelet recruitment

Platelets, although not phagocytotic, have been suggested to release O. Since O-producing reduced nicotinamide adenine dinucleotide (phosphate) (NAD(P)H) oxidases can be specifically activated by certain agonists and are found in several nonphagocytotic tissues, we investigated whether such an enzyme is the source of platelet-derived O. We further studied which agonists cause platelet O release and whether platelet-derived O influences thrombus formation in vitro. Collagen, but not adenosine 5'-diphosphate (ADP) or thrombin, increased O formation in washed human platelets. This was a reduced nicotinamide adenine dinucleotide (NADH)-dependent process, as shown in platelet lysates. Consistent with a role of a platelet, NAD(P)H oxidase expression of its subunits p47(phox) and p67(phox) and inhibition of platelet O formation by diphenylene-iodoniumchloride (DPI) and by the specific peptide-antagonist gp91ds-tat were observed. Whereas platelet-derived O did not influence initial aggregation, platelet recruitment to a preformed thrombus following collagen stimulation was significantly attenuated by superoxide dismutase (SOD) or DPI. It was also inhibited when ADP released during aggregation was cleaved by the ectonucleotidase apyrase. ADP in supernatants of collagen-activated platelets was decreased in the presence of SOD, resulting in lower ADP concentrations available for recruitment of further platelets. Exogenous O increased ADP- concentrations in supernatants of collagen-stimulated platelets and induced irreversible aggregation when platelets were stimulated with otherwise subthreshold concentrations of ADP. These results strongly suggest that collagen activation induces NAD(P)H oxidase-dependent O release in platelets, which in turn enhances availability of released ADP, resulting in increased platelet recruitment.     

Phenotype-Genotype Correlations and Estimated Carrier Frequencies of Primary Hyperoxaluria

Primary hyperoxaluria (PH) is a rare autosomal recessive disease characterized by oxalate accumulation in the kidneys and other organs. Three loci have been identified: AGXT (PH1), GRHPR (PH2), and HOGA1 (PH3). Here, we compared genotype to phenotype in 355 patients in the Rare Kidney Stone Consortium PH registry and calculated prevalence using publicly available whole-exome data. PH1 (68.4% of families) was the most severe PH type, whereas PH3 (11.0% of families) showed the slowest decline in renal function but the earliest symptoms. A group of patients with disease progression similar to that of PH3, but for whom no mutation was detected (11.3% of families), suggested further genetic heterogeneity. We confirmed that the AGXT p.G170R mistargeting allele resulted in a milder PH1 phenotype; however, other potential AGXT mistargeting alleles caused more severe (fully penetrant) disease. We identified the first PH3 patient with ESRD; a homozygote for two linked, novel missense mutations. Population analysis suggested that PH is an order of magnitude more common than determined from clinical cohorts (prevalence, approximately 1:58,000; carrier frequency, approximately 1:70). We estimated PH to be approximately three times less prevalent among African Americans than among European Americans because of a limited number of common European origin alleles. PH3 was predicted to be as prevalent as PH1 and twice as common as PH2, indicating that PH3 (and PH2) cases are underdiagnosed and/or incompletely penetrant. These results highlight a role for molecular analyses in PH diagnostics and prognostics and suggest that wider analysis of the idiopathic stone-forming population may be beneficial.     

Pulmonary Hypertension in Patients With Severe Aortic Stenosis: Prognostic Impact After Transcatheter Aortic Valve Replacement: Pulmonary Hypertension in Patients Undergoing TAVR

Objectives

The authors investigated the development of pulmonary hypertension (PH), predictors of PH regression, and its prognostic impact on short, mid-, and long-term outcomes in patients undergoing transcatheter aortic valve replacement (TAVR) for severe aortic stenosis (AS).

Genomic and Functional Portrait of a Highly Virulent,CTX-M-15-Producing H30-Rx Subclone of Escherichia coli Sequence Type 131

Escherichia coli sequence type 131 (ST131) is a pandemic clone associated with multidrug-resistant, extraintestinal infections, attributable to the presence of the CTX-M-15 extended-spectrum β-lactamase gene and mutations entailing fluoroquinolone resistance. Studies on subclones within E. coli ST131 are critically required for targeting and implementation of successful control efforts. Our study comprehensively analyzed the genomic and functional attributes of the H30-Rx subclonal strains NA097 and NA114, belonging to the ST131 lineage. We carried out whole-genome sequencing, comparative analysis, phenotypic virulence assays, and profiling of the antibacterial responses of THP1 cells infected with these subclones. Phylogenomic analysis suggested that the strains were clonal in nature and confined entirely to a single clade. Comparative genomic analysis revealed that the virulence and resistance repertoires were comparable among the H30-Rx ST131 strains except for the commensal ST131 strain SE15. Similarly, seven phage-specific regions were found to be strongly associated with the H30-Rx strains but were largely absent in the genome of SE15. Phenotypic analysis confirmed the virulence and resistance similarities between the two strains. However, NA097 was found to be more robust than NA114 in terms of virulence gene carriage (dra operon), invasion ability (P < 0.05), and antimicrobial resistance (streptomycin resistance). RT² gene expression profiling revealed generic upregulation of key proinflammatory responses in THP1 cells, irrespective of ST131 lineage status. In conclusion, our study provides comprehensive, genome-inferred insights into the biology and immunological properties of ST131 strains and suggests clonal diversification of genomic and phenotypic features within the H30-Rx subclone of E. coli ST131.     

Unique method of tooth replacement in durophagous placodont marine reptiles,with new data on the dentition of Chinese taxa

The placodonts of the Triassic period (~252–201 mya) represent one of the earliest and most extreme specialisations to a durophagous diet of any known reptile group. Exceptionally enlarged crushing tooth plates on the maxilla, dentary and palatine cooperated to form functional crushing areas in the buccal cavity. However, the extreme size of these teeth, combined with the unusual way they occluded, constrained how replacement occurred. Using an extensive micro‐computed tomographic dataset of 11 specimens that span all geographic regions and placodont morphotypes, tooth replacement patterns were investigated. In addition, the previously undescribed dental morphologies and formulae of Chinese taxa are described for the first time and incorporated into the analysis. Placodonts have a unique tooth replacement pattern and results follow a phylogenetic trend. The plesiomorphic Placodus species show many replacement teeth at various stages of growth, with little or no discernible pattern. On the other hand, the more derived cyamodontoids tend to have fewer replacement teeth growing at any one time, replacing teeth unilaterally and/or in functional units, thus maintaining at least one functional crushing area at all times. The highly derived placochelyids have fewer teeth and, as a result, only have one or two replacement teeth in the upper jaw. This supports previous suggestions that these taxa had an alternative diet to other placodonts. Importantly, all specimens show at least one replacement tooth growing at the most posterior palatine tooth plates, indicating increased wear at this point and thus the most efficient functional crushing area.     

Imaging Neuroinflammation? A Perspective from MR Spectroscopy

Neuroinflammatory mechanisms contribute to the brain pathology resulting from human immunodeficiency virus (HIV) infection. Magnetic resonance spectroscopy (MRS) has been touted as a suitable method for discriminating in vivo markers of neuroinflammation. The present MRS study was conducted in four groups: alcohol dependent (A, n = 37), HIV‐infected (H, n = 33), alcohol dependent + HIV infected (HA, n = 38) and healthy control (C, n = 62) individuals to determine whether metabolites would change in a pattern reflecting neuroinflammation. Significant four‐group comparisons were evident only for striatal choline‐containing compounds (Cho) and myo‐inositol (mI), which follow‐up analysis demonstrated were due to higher levels in HA compared with C individuals. To explore the potential relevance of elevated Cho and mI, correlations between blood markers, medication status and alcohol consumption were evaluated in H + HA subjects. Having an acquired immune deficiency syndrome (AIDS)‐defining event or hepatitis C was associated with higher Cho; lower Cho levels, however, were associated with low thiamine levels and with highly active antiretroviral HIV treatment (HAART). Higher levels of mI were related to greater lifetime alcohol consumed, whereas HAART was associated with lower mI levels. The current results suggest that competing mechanisms can influence in vivo Cho and mI levels, and that elevations in these metabolites cannot necessarily be interpreted as reflecting a single underlying mechanism, including neuroinflammation.