On the mechanism of calcium-dependent activation of NADPH oxidase 5 (NOX5)

It is now accepted that reactive oxygen species (ROS) are not only dangerous oxidative agents but also chemical mediators of the redox cell signaling and innate immune response. A central role in ROS-controlled production is played by the NADPH oxidases (NOXs), a group of seven membrane-bound enzymes (NOX1-5 and DUOX1-2) whose unique function is to produce ROS. Here, we describe the regulation of NOX5, a widespread family member present in cyanobacteria, protists, plants, fungi, and the animal kingdom. We show that the calmodulin-like regulatory EF-domain of NOX5 is partially unfolded and detached from the rest of the protein in the absence of calcium. In the presence of calcium, the C-terminal lobe of the EF-domain acquires an ordered and more compact structure that enables its binding to the enzyme dehydrogenase (DH) domain. Our spectroscopic and mutagenesis studies further identified a set of conserved aspartate residues in the DH domain that are essential for NOX5 activation. Altogether, our work shows that calcium induces an unfolded-to-folded transition of the EF-domain that promotes direct interaction with a conserved regulatory region, resulting in NOX5 activation

Boceprevir is highly effective in treatment-experienced hepatitis C virus-positive genotype-1 menopausal women

AIM: To investigate the safety/efficacy of Boceprevirbased triple therapy in hepatitis C virus (HCV)-G1 menopausal women who were historic relapsers, partial-responders and null-responders. METHODS: In this single-assignment, unblinded study, we treated fifty-six menopausal women with HCV-G1, 46% F3-F4, and previous PEG-α/RBV failure (7% null, 41% non-responder, and 52% relapser) with 4 wk lead-in with PEG-IFNα2b/RBV followed by PEGIFNα2b/RBV+Boceprevir for 32 wk, with an additional 12 wk of PEG-IFN-α-2b/RBV if patients were HCV-RNA-positive by week 8. In previous null-responders, 44 wk of triple therapy was used. The primary objective of retreatment was to verify whether a sustained virological response (SVR) (HCV RNA undetectable at 24 wk of follow-up) rate of at least 20% could be obtained. The secondary objective was the evaluation of the percent of patients with negative HCV RNA at week 4 (RVR), 8 (RVR BOC), 12 (EVR), or at the end-of-treatment (ETR) that reached SVR. To assess the relationship between SVR and clinical and biochemical parameters, multiple logistic regression analysis was used. RESULTS: After lead-in, only two patients had RVR; HCV-RNA was unchanged in all but 62% who had ≤ 1 logio decrease. After Boceprevir, HCV RNA became undetectable at week 8 in 32/56 (57.1%) and at week 12 in 41/56 (73.2%). Of these, 53.8% and 52.0%, respectively, achieved SVR. Overall, SVR was obtained in 25/56 (44.6%). SVR was achieved in 55% previous relapsers vs. 41% non-responders (Ρ = 0.250), in 44% F0-F2 vs 54% F3-F4 (Ρ = 0.488), and in 11/19 (57.9%) of patients with cirrhosis. At univariate analysis for baseline predictors of SVR, only previous response to antiviral therapy (OR = 2.662, 95%CI: 0.957-6.881, Ρ= 0.043), was related with SVR. When considering "on treatment" factors, 1 log10 HCV RNA decline at week 4 (3.733, 95%CI: 1.676-12.658, Ρ= 0.034) and achievement of RVR BOC (7.347, 95%CI: 2.156-25.035, Ρ= 0.001) were significantly related with the SVR, al-though RVR BOC only (6.794, 95%CI: 1.596-21.644, Ρ = 0.010) maintained significance at multivariate logistic regression analysis. Anemia and neutropenia were managed with Erythropoietin and Filgrastim supplementation, respectively. Only six patients discontinued therapy. CONCLUSION: Boceprevir obtained high SVR response independent of previous response, RVR or baseline fibrosis or cirrhosis. RVR BOC was the only independent predictor of SVR

Association Study with 77 SNPs Confirms the Robust Role for the rs10830963/G of MTNR1B Variant and Identifies Two Novel Associations in Gestational Diabetes Mellitus Development

CONTEXT: Genetic variation in human maternal DNA contributes to the susceptibility for development of gestational diabetes mellitus (GDM). OBJECTIVE: We assessed 77 maternal single nucleotide gene polymorphisms (SNPs) for associations with GDM or plasma glucose levels at OGTT in pregnancy. METHODS: 960 pregnant women (after dropouts 820: case/control: m99'WHO: 303/517, IADPSG: 287/533) were enrolled in two countries into this case-control study. After genomic DNA isolation the 820 samples were collected in a GDM biobank and assessed using KASP (LGC Genomics) genotyping assay. Logistic regression risk models were used to calculate ORs according to IADPSG/m'99WHO criteria based on standard OGTT values. RESULTS: The most important risk alleles associated with GDM were rs10830963/G of MTNR1B (OR = 1.84/1.64 [IADPSG/m'99WHO], p = 0.0007/0.006), rs7754840/C (OR = 1.51/NS, p = 0.016) of CDKAL1 and rs1799884/T (OR = 1.4/1.56, p = 0.04/0.006) of GCK. The rs13266634/T (SLC30A8, OR = 0.74/0.71, p = 0.05/0.02) and rs7578326/G (LOC646736/IRS1, OR = 0.62/0.60, p = 0.001/0.006) variants were associated with lower risk to develop GDM. Carrying a minor allele of rs10830963 (MTNR1B); rs7903146 (TCF7L2); rs1799884 (GCK) SNPs were associated with increased plasma glucose levels at routine OGTT. CONCLUSIONS: We confirmed the robust association of MTNR1B rs10830963/G variant with GDM binary and glycemic traits in this Caucasian case-control study. As novel associations we report the minor, G allele of the rs7578326 SNP in the LOC646736/IRS1 region as a significant and the rs13266634/T SNP (SLC30A8) as a suggestive protective variant against GDM development. Genetic susceptibility appears to be more preponderant in individuals who meet both the modified 99'WHO and the IADPSG GDM diagnostic criteria

Intra-tumoural microvessel density in human solid tumours

Over the last decade assessment of angiogenesis has emerged as a potentially useful biological prognostic and predictive factor in human solid tumours. With the development of highly specific endothelial markers that can be assessed in histological archival specimens, several quantitative studies have been performed in various solid tumours. The majority of published studies have shown a positive correlation between intra-tumoural microvessel density, a measure of tumour angiogenesis, and prognosis in solid tumours. A minority of studies have not demonstrated an association and this may be attributed to significant differences in the methodologies employed for sample selection, immunostaining techniques, vessel counting and statistical analysis, although a number of biological differences may account for the discrepancy. In this review we evaluate the quantification of angiogenesis by immunohistochemistry, the relationship between tumour vascularity and metastasis, and the clinicopathological studies correlating intra-tumoral microvessel density with prognosis and response to anti-cancer therapy. In view of the extensive nature of this retrospective body of data, comparative studies are needed to identify the optimum technique and endothelial antigens (activated or pan-endothelial antigens) but subsequently prospective studies that allocate treatment on the basis of microvessel density are required