Simultaneous determination of natural and synthetic steroid estrogens and their conjugates in aqueous matrices by liquid chromatography / mass spectrometry

An analytical method for the simultaneous determination of nine free and conjugated steroid estrogens was developed with application to environmental aqueous matrices. Solid phase extraction (SPE) was employed for isolation and concentration, with detection by liquid chromatography/mass spectrometry (LC/MS) using electrospray ionisation (ESI) in the negative mode. Method recoveries for various aqueous matrices (wastewater, lake and drinking water) were determined, recoveries proving to be sample dependent. When spiked at 50 ng/l concentrations in sewage influent, recoveries ranged from 62-89 % with relative standard deviations (RSD) < 8.1 %. In comparison, drinking water spiked at the same concentrations had recoveries between 82-100 % with an RSD < 5%. Ion suppression is a known phenomenon when using ESI; hence its impact on method recovery was elucidated for raw sewage. Both ion suppression from matrix interferences and the extraction procedure has bearing on the overall method recovery. Analysis of municipal raw sewage identified several of the analytes of interest at ng/l concentrations, estriol (E3) being the most abundant. Only one conjugate, estrone 3-sulphate (E1-3S) was observe

ESTs, cDNA microarrays, and gene expression profiling : tools for dissecting plant physiology and development

Gene expression profiling holds tremendous promise for dissecting the regulatory mechanisms and transcriptional networks that underlie biological processes. Here we provide details of approaches used by others and ourselves for gene expression profiling in plants with emphasis on cDNA microarrays and discussion of both experimental design and downstream analysis. We focus on methods and techniques emphasizing fabrication of cDNA microarrays, fluorescent labeling, cDNA hybridization, experimental design, and data processing. We include specific examples that demonstrate how this technology can be used to further our understanding of plant physiology and development (specifically fruit development and ripening) and for comparative genomics by comparing transcriptome activity in tomato and pepper fruit

PeerGAD: a peer-review-based and community-centric web application for viewing and annotating prokaryotic genome sequences

PeerGAD is a web-based database-driven application that allows community-wide peer-reviewed annotation of prokaryotic genome sequences. The application was developed to support the annotation of the Pseudomonas syringae pv. tomato strain DC3000 genome sequence and is easily portable to other genome sequence annotation projects. PeerGAD incorporates several innovative design and operation features and accepts annotations pertaining to gene naming, role classification, gene translation and annotation derivation. The annotator tool in PeerGAD is built around a genome browser that offers users the ability to search and navigate the genome sequence. Because the application encourages annotation of the genome sequence directly by researchers and relies on peer review, it circumvents the need for an annotation curator while providing added value to the annotation data. Support for the Gene Ontology™ vocabulary, a structured and controlled vocabulary used in classification of gene roles, is emphasized throughout the system. Here we present the underlying concepts integral to the functionality of PeerGAD

Outcomes of Transcatheter Edge-to-Edge Repair in Degenerative vs. Functional Mitral Regurgitation

Current guidelines support the use of transcatheter edge-to-edge repair (TEER) for patients with both primary and secondary mitral regurgitation. We aimed to compare the prognoses of TEER in degenerative mitral regurgitation (DMR) vs. functional mitral regurgitation (FMR). A total of 208 consecutive patients who underwent TEER over a ten-year period were analyzed. Primary endpoints included rates of all-cause death and major adverse cardiac events (MACE: composite of all-cause death, hospitalizations for heart failure, mitral valve surgery, or TEER re-intervention). A total of 148 (71%) patients were identified with FMR, while 60 (29%) were identified with DMR. Patients in the FMR group were younger (77.2 ± 8.4 vs. 80.2 ± 7.2, p = 0.02), suffered more frequently from coronary artery disease (54.1% vs. 10.0%, p = 0.02), and atrial fibrillation/flutter (70.9% vs. 38.3%, p = 0.02). Rates of 1-year death (21.6% vs. 10.0%, p = 0.03) and MACE (41.2% vs. 21.7%, p = 0.02) were higher for the FMR group, as compared to the DMR group. After correcting for variables, FMR independently predicted rates of MACE (HR-1.78, 95% CI 1.23–2.48, p = 0.04) and had a non-significant effect on one-year mortality (HR-1.67, 95%CI 0.98–3.74, p = 0.07). In our experience, worse overall 1-year composite MACE outcomes were observed after TEER in patients with FMR as compared to patients with DMR

An Integrated Tool to Study MHC Region: Accurate SNV Detection and HLA Genes Typing in Human MHC Region Using Targeted High-Throughput Sequencing

<div><p>The major histocompatibility complex (MHC) is one of the most variable and gene-dense regions of the human genome. Most studies of the MHC, and associated regions, focus on minor variants and HLA typing, many of which have been demonstrated to be associated with human disease susceptibility and metabolic pathways. However, the detection of variants in the MHC region, and diagnostic HLA typing, still lacks a coherent, standardized, cost effective and high coverage protocol of clinical quality and reliability. In this paper, we presented such a method for the accurate detection of minor variants and HLA types in the human MHC region, using high-throughput, high-coverage sequencing of target regions. A probe set was designed to template upon the 8 annotated human MHC haplotypes, and to encompass the 5 megabases (Mb) of the extended MHC region. We deployed our probes upon three, genetically diverse human samples for probe set evaluation, and sequencing data show that ∼97% of the MHC region, and over 99% of the genes in MHC region, are covered with sufficient depth and good evenness. 98% of genotypes called by this capture sequencing prove consistent with established HapMap genotypes. We have concurrently developed a one-step pipeline for calling any HLA type referenced in the IMGT/HLA database from this target capture sequencing data, which shows over 96% typing accuracy when deployed at 4 digital resolution. This cost-effective and highly accurate approach for variant detection and HLA typing in the MHC region may lend further insight into immune-mediated diseases studies, and may find clinical utility in transplantation medicine research. This one-step pipeline is released for general evaluation and use by the scientific community.</p></div

Acute Reduction in Left Ventricular Function Following Transcatheter Mitral Edge‐to‐Edge Repair

Background Little is known about the impact of transcatheter mitral valve edge‐to‐edge repair on changes in left ventricular ejection fraction (LVEF) and the effect of an acute reduction in LVEF on prognosis. We aimed to assess changes in LVEF after transcatheter mitral valve edge‐to‐edge repair for both primary and secondary mitral regurgitation (PMR and SMR, respectively), identify rates and predictors of LVEF reduction, and estimate its impact on prognosis. Methods and Results In this international multicenter registry, patients with both PMR and SMR undergoing transcatheter mitral valve edge‐to‐edge repair were included. We assessed rates of acute LVEF reduction (LVEFR), defined as an acute relative decrease of >15% in LVEF, its impact on all‐cause mortality, major adverse cardiac event (composite end point of all‐cause death, mitral valve surgery, and residual mitral regurgitation grade ≥2), and LVEF at 12 months, as well as predictors for LVEFR. Of 2534 patients included (727 with PMR, and 1807 with SMR), 469 (18.5%) developed LVEFR. Patients with PMR were older (79.0±9.2 versus 71.8±8.9 years; P<0.001) and had higher mean LVEF (54.8±14.0% versus 32.7±10.4%; P<0.001) at baseline. After 6 to 12 months (median, 9.9 months; interquartile range, 7.8–11.9 months), LVEF was significantly lower in patients with PMR (53.0% versus 56.0%; P<0.001) but not in patients with SMR. The 1‐year mortality was higher in patients with PMR with LVEFR (16.9% versus 9.7%; P<0.001) but not in those with SMR (P=0.236). LVEF at baseline (odds ratio, 1.03 [95% CI, 1.01–1.05]; P=0.002) was predictive of LVEFR for patients with PMR, but not those with SMR (P=0.092). Conclusions Reduction in LVEF is not uncommon after transcatheter mitral valve edge‐to‐edge repair and is correlated with worsened prognosis in patients with PMR but not patients with SMR. Registration URL: https://www.clinicaltrials.gov; Unique identifier: NCT05311163