Polycarbonate Ultracentrifuge Tube Re-use in Proteomic Analyses of Extracellular Vesicles.

Single-use laboratory plastics exacerbate the pollution crisis and contribute to consumable costs. In extracellular vesicle (EV) isolation, polycarbonate ultracentrifuge (UC) tubes are used to endure the associated high centrifugal forces. EV proteomics is an advancing field and validated re-use protocols for these tubes are lacking. Re-using consumables for low-yield protein isolation protocols and downstream proteomics requires reagent compatibility with mass spectroscopy acquisitions, such as the absence of centrifuge tube-derived synthetic polymer contamination, and sufficient removal of residual proteins. This protocol describes and validates a method for cleaning polycarbonate UC tubes for re-use in EV proteomics experiments. The cleaning process involves immediate submersion of UC tubes in H2O to prevent protein drying, washing in 0.1% sodium dodecyl sulfate (SDS) detergent, rinsing in hot tap water, demineralized water, and 70% ethanol. To validate the UC tube re-use protocol for downstream EV proteomics, used tubes were obtained following an experiment isolating EVs from cardiovascular tissue using differential UC and density gradient separation. Tubes were cleaned and the experimental process was repeated without EV samples comparing blank never-used UC tubes to cleaned UC tubes. The pseudo-EV pellets obtained from the isolation procedures were lysed and prepared for liquid chromatography-tandem mass spectrometry using a commercial protein sample preparation kit with modifications for low-abundance protein samples. Following cleaning, the number of identified proteins was reduced by 98% in the pseudo-pellet versus the previous EV isolation sample from the same tube. Comparing a cleaned tube against a blank tube, both samples contained a very small number of proteins (≤20) with 86% similarity. The absence of polymer peaks in the chromatograms of the cleaned tubes was confirmed. Ultimately, the validation of a UC tube cleaning protocol suitable for the enrichment of EVs will reduce the waste produced by EV laboratories and lower the experimental costs.

Genetic mechanisms underlying arrhythmogenic mitral valve prolapse: Current and future perspectives.

Mitral valve prolapse (MVP) is a heart valve disease that is often familial, affecting 2%-3% of the general population. MVP with or without mitral regurgitation can be associated with an increased risk of ventricular arrhythmias and sudden cardiac death (SCD). Research on familial MVP has specifically focused on genetic factors, which may explain the heritable component of the disease estimated to be present in 20%-35%. Furthermore, the structural and electrophysiological substrates underlying SCD/ventricular arrhythmia risk in MVP have been studied postmortem and in the electrophysiology laboratory, respectively. Understanding how familial MVP and rhythm disorders are related may help patients with MVP by individualizing risk and working to develop effective management strategies. This contemporary, state-of-the-art, expert review focuses on genetic factors and familial components that underlie MVP and arrhythmia and encapsulates clinical, genetic, and electrophysiological issues that should be the objectives of future research.

An Unexpected Turn of Events in a Patient With Mitral Valve Endocarditis.

A variety of gram-positive infections can be treated with daptomycin. Daptomycin-induced acute eosinophilic pneumonia (DEP) is a rare adverse drug reaction with nonspecific clinical findings of dyspnea, dry cough, and fever. Although diagnostic criteria exist, prompt recognition is important to prevent rapid progression and respiratory failure. In this case, a 69-year-old female was initially admitted due to a prosthetic joint infection; however, her case was complicated by DEP.

A Case of Bilateral Inflammatory Breast Cancer.

Inflammatory breast cancer (IBC) is an aggressive form of breast carcinoma. Bilateral occurrences of IBC within a short time frame are rare, particularly without significant surgical intervention. This case presents a patient with contralateral recurrence of IBC less than a year after the initial diagnosis. A 39-year-old female was diagnosed with stage IV inflammatory breast cancer in her left breast. Less than a year later, extensive disease was found in her right breast. The patient had received incomplete treatment for the left IBC due to barriers to accessing care. Imaging confirmed the presence of inflammatory breast cancer in the contralateral breast, along with regional adenopathy and metastases. The patient began a chemotherapy regimen similar to her previous treatment. This case highlights the uncommon occurrence of contralateral recurrence of IBC and the hypothesized mechanism of lymphatic spread, suggesting local metastasis rather than a new primary tumor. The patient's incomplete treatment and lack of surgical intervention likely contributed to the development of contralateral IBC. The case underscores the importance of magnetic resonance imaging (MRI) in evaluating soft tissue and lymphatic changes in IBC. Barriers to care negatively impact prognosis, emphasizing the need for timely follow-up, diagnostic imaging, and oncologic therapy for successful treatment.

Ophthalmia nodosa secondary to intraocular, white-marked tussock caterpillar setae (Orgyia leucostigma) in a 15-year-old.

A 15-year-old female presented to the emergency department with a 1-day history of pain and swelling of her right eye following ocular contact with a caterpillar. Caterpillars of the white-marked tussock moth and other related species possess hairlike structures called setae, with angled barbs along their length, allowing them to travel linearly upon penetration of an enemy, resisting backward movement and becoming very difficult to extract once imbedded. When these fine, pointed hairs contact the surface of the eye, they can easily migrate in, eliciting movement of the globe, blinking, and rubbing of the eye in an attempt to remove the offending agent, potentially leading to ophthalmia nodosa. One of the most important aspects of ophthalmia nodosa diagnosis is a thorough history and prompt slit-lamp examination to identify the presence of foreign bodies and where they are located; this will help guide clinical management decisions. This case demonstrates that, depending on the number and location, more than one attempt may be required to remove all of the barbed setae. If ophthalmia nodosa is suspected, it is important to promptly refer to ophthalmology for a thorough eye exam, keep the eye clean, prescribe prophylactic topical antibiotics and/or steroids to reduce the potential for infection and inflammation, and emphasize the importance of keeping the eye protected during the healing process with an eye shield.

Shotgun metagenomic analysis of microbial communities from the Loxahatchee nature preserve in the Florida Everglades.

BACKGROUND: Currently, much is unknown about the taxonomic diversity and the mechanisms of methane metabolism in the Florida Everglades ecosystem. The Loxahatchee National Wildlife Refuge is a section of the Florida Everglades that is almost entirely unstudied in regard to taxonomic profiling. This short report analyzes the metagenome of soil samples from this Refuge to investigate the predominant taxa, as well as the abundance of genes involved in environmentally significant metabolic pathways related to methane production (nitrogen fixation and dissimilatory sulfite reduction). METHODS: Shotgun metagenomic sequencing using the Illumina platform was performed on 17 soil samples from four different sites within the Loxahatchee National Wildlife Refuge, and underwent quality control, assembly, and annotation. The soil from each sample was tested for water content and concentrations of organic carbon and nitrogen. RESULTS: The three most common phyla of bacteria for every site were Actinobacteria, Acidobacteria, and Proteobacteria; however, there was variation in relative phylum composition. The most common phylum of Archaea was Euryarchaeota for all sites. Alpha and beta diversity analyses indicated significant congruity in taxonomic diversity in most samples from Sites 1, 3, and 4 and negligible congruity between Site 2 and the other sites. Shotgun metagenomic sequencing revealed the presence of biogeochemical biomarkers of particular interest (e.g., mrcA, nifH, and dsrB) within the samples. The normalized abundances of mcrA, nifH, and dsrB exhibited a positive correlation with nitrogen concentration and water content, and a negative correlation with organic carbon concentration. CONCLUSION: This Everglades soil metagenomic study allowed examination of wetlands biological processes and showed expected correlations between measured organic constituents and prokaryotic gene frequency. Additionally, the taxonomic profile generated gives a basis for the diversity of prokaryotic microbial life throughout the Everglades.