Revealing New Analytical Insights into RNA Complexes: Divalent siRNA Characterization by Liquid Chromatography and Mass Spectrometry.

Accurate characterization of therapeutic RNA, including purity and identity, is critical in drug discovery and development. Here, we utilize denaturing and non-denaturing chromatography for the analysis of ∼25 kDa divalent small interfering RNA (di-siRNA), which comprises a complex 2:1 triplex structure. Ion pair reversed-phase (IPRP) liquid chromatography (LC) experiments with UV absorbance and mass spectrometry (MS) showcase a single denaturing LC method for identity confirmation, impurity profiling, and sequencing with automated MS data interpretation.

Para-aminopropiophenone toxicity in domestic dogs: a description of non-target toxicosis in Victoria, Australia: 13 dogs (2016-2023).

Background: PAPP is widely used in Australia as a potent vertebrate bait, with potential for off-target ingestion and poisoning in domestic dogs. Whilst toxicosis and resulting methaemoglobinaemia is anecdotally known to occur, this is the first description in the literature. This study reports thirteen clinical cases of suspected Para-aminopropiophenone (PAPP) toxicity in dogs, with the aim of describing clinical presentation and current management of toxicosis in this species.

Rational Design of Novel Allosteric EYA2 Inhibitors as Potential Therapeutics for Multiple Brain Cancers.

The Eyes Absent (EYA) family of developmental proteins, often in partnership with the sine oculis (SIX) homeobox proteins, promote cancer metastasis and recurrence in numerous tumor types. In addition to being a transcriptional coactivator, EYA2 is a Tyr phosphatase that dephosphorylates H2AX which leads to repair instead of apoptosis upon DNA damage and ERβ which inhibits the anti-tumor transcriptional activity of ERβ. The SIX members of the EYA-SIX complex are difficult to target, therefore, we targeted the EYA2 to promote cell death and prevent cancer progression.

Immunogenicity and Protective Efficacy of Psoralen-Inactivated SARS-CoV-2 Vaccine in Nonhuman Primates.

COVID-19 caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has significantly impacted public health and the economy worldwide. Most of the currently licensed COVID-19 vaccines act by inhibiting the receptor-binding function of the SARS-CoV-2 spike protein. The constant emergence of SARS-CoV-2 variants resulting from mutations in the receptor-binding domain (RBD) leads to vaccine immune evasion and underscores the importance of broadly acting COVID-19 vaccines.

Photochemical inactivation as an alternative method to produce a whole-cell vaccine for uropathogenic (UPEC).

Unlabelled: Uropathogenic (UPEC) is the primary causative agent of lower urinary tract infection (UTI). UTI presents a serious health risk and has considerable secondary implications including economic burden, recurring episodes, and overuse of antibiotics. A safe and effective vaccine would address this widespread health problem and emerging antibiotic resistance.

Characterization of antisense oligonucleotide and guide ribonucleic acid diastereomers by hydrophilic interaction liquid chromatography coupled to mass spectrometry.

Oligonucleotides have become an essential modality for a variety of therapeutic approaches, including cell and gene therapies. Rapid progress in the field has attracted significant research in designing novel oligonucleotide chemistries and structures. Beyond their polar nature, the length of large RNAs and presence of numerous diastereomers for phosphorothioate (PS)-modified RNAs pose heightened challenges for their characterization.

The Immunogenicity and Properties of a Whole-Cell ETEC Vaccine Inactivated with Psoralen and UVA Light in Comparison to Formalin.

Inactivated whole-cell vaccines present a full repertoire of antigens to the immune system. Formalin treatment, a standard method for microbial inactivation, can modify or destroy protein antigenic epitopes. We tested the hypothesis that photochemical inactivation with psoralen and UVA light (PUVA), which targets nucleic acid, would improve the immunogenicity of an Enterotoxigenic (ETEC) vaccine relative to a formalin-inactivated counterpart.

Frailty impacts immune responses to Moderna COVID-19 mRNA vaccine in older adults.

Background: Immune responses to COVID-19 mRNA vaccines have not been well characterized in frail older adults. We postulated that frailty is associated with impaired antibody and cellular mRNA vaccine responses.

Methods: We followed older adults in a retirement facility with longitudinal clinical and serological samples from the first Moderna mRNA-1273 vaccine dose starting in February 2021 through their 3rd (booster) vaccine dose.

Deeper Understanding of Solvent-Based Ambient Ionization Mass Spectrometry: Are Molecular Profiles Primarily Dictated by Extraction Mechanisms?

Solvent-based ambient ionization mass spectrometry (MS) techniques provide a powerful approach for direct chemical analysis and molecular profiling of biological tissues. While molecular profiling of tissues has been widely used for disease diagnosis, little is understood about how the interplay among solvent properties, matrix effects, and ion suppression can influence the detection of biological molecules. Here, we perform a systematic investigation of the extraction processes of lipids using an ambient ionization droplet microsampling platform to investigate how the physicochemical properties of the solvent systems and extraction time influence molecular extraction and detection.

Structural Characterization and Quantitation of Ether-Linked Glycerophospholipids in Peroxisome Biogenesis Disorder Tissue by Ultraviolet Photodissociation Mass Spectrometry.

The biological impact of ether glycerophospholipids (GP) in peroxisomal disorders and other diseases makes them significant targets as biomarkers for diagnostic assays or deciphering pathology of the disorders. Ether lipids include both plasmanyl and plasmenyl lipids, which each contain an ether or a vinyl ether bond at the -1 linkage position, respectively. This linkage, in contrast to traditional diacyl GPs, precludes their detailed characterization by mass spectrometry via traditional collisional-based MS/MS techniques.

Nanohydrophobic Interaction Chromatography Coupled to Ultraviolet Photodissociation Mass Spectrometry for the Analysis of Intact Proteins in Low Charge States.

The direct correlation between proteoforms and biological phenotype necessitates the exploration of mass spectrometry (MS)-based methods more suitable for proteoform detection and characterization. Here, we couple nano-hydrophobic interaction chromatography (nano-HIC) to ultraviolet photodissociation MS (UVPD-MS) for separation and characterization of intact proteins and proteoforms. High linearity, sensitivity, and sequence coverage are obtained with this method for a variety of proteins.

Frailty and Age Impact Immune Responses to Moderna COVID-19 mRNA Vaccine.

Background: Immune responses to COVID-19 mRNA vaccines have not been well characterized in frail older adults. We postulated that frailty is associated with impaired antibody and cellular mRNA vaccine responses.

Methods: We followed older adults in a retirement facility with longitudinal clinical and serological samples from the first Moderna mRNA-1273 vaccine dose starting in February 2021 through their 3rd (booster) vaccine dose.

Enhanced Ion Mobility Separation and Characterization of Isomeric Phosphatidylcholines Using Absorption Mode Fourier Transform Multiplexing and Ultraviolet Photodissociation Mass Spectrometry.

The structural diversity of phospholipids plays a critical role in cellular membrane dynamics, energy storage, and cellular signaling. Despite its importance, the extent of this diversity has only recently come into focus, largely owing to advances in separation science and mass spectrometry methodology and instrumentation. Characterization of glycerophospholipid (GP) isomers differing only in their acyl chain configurations and locations of carbon-carbon double bonds (C═C) remains challenging due to the need for both effective separation of isomers and advanced tandem mass spectrometry (MS/MS) technologies capable of double-bond localization.

Elevator Cabin Decontamination With ACTIVE Particle Control™ Technology.

Effectively reducing contamination and aerosolized bioburden may limit the risk of disease transmission in closed settings when social distancing is not possible. Unlike uncontrolled ionization and oxidation devices ACTIVE Particle Control™ conditions particles in a highly controlled fashion which provides effective air purification without the generation of ozone or other toxic by-products. The purpose of this study was to determine the impact of ACTIVE Particle Control™ on elevator cabin particle load compared to standard ventilation.

Characterization of the T4 gp32-ssDNA complex by native, cross-linking, and ultraviolet photodissociation mass spectrometry.

Protein-DNA interactions play crucial roles in DNA replication across all living organisms. Here, we apply a suite of mass spectrometry (MS) tools to characterize a protein-ssDNA complex, T4 gp32·ssDNA, with results that both support previous studies and simultaneously uncover novel insight into this non-covalent biological complex. Native mass spectrometry of the protein reveals the co-occurrence of Zn-bound monomers and homodimers, while addition of differing lengths of ssDNA generates a variety of protein:ssDNA complex stoichiometries (1 : 1, 2 : 1, 3 : 1), indicating sequential association of gp32 monomers with ssDNA.

Retrograde Parotid Gland Infusion through Stensen's Duct in a Non-Human Primate for Vectored Gene Delivery.

Salivary glands are an attractive tissue target for gene therapy with promising results already leading to human trials. They are inherently capable of secreting proteins into the bloodstream and are easily accessible, making them potentially superior tissue sites for replacement hormone production or vaccination by gene transfer. Suggested methods for gene delivery include transcutaneous injection and retrograde infusion through salivary ducts.

The time required to achieve endotracheal intubation in dogs: a comparison of veterinary students and qualified veterinary surgeons.

Objective: To determine whether final year veterinary students take longer to perform endotracheal intubation than qualified veterinary surgeons.

Study Design: Observational cohort study.

Animals: A total of 38 healthy mesocephalic dogs undergoing general anaesthesia for a clinical purpose unrelated to this study.

Inhibition of CtBP-Regulated Proinflammatory Gene Transcription Attenuates Psoriatic Skin Inflammation.

Psoriasis is a chronic immune-mediated disease characterized by excessive proliferation of epidermal keratinocytes and increased immune cell infiltration to the skin. Although it is well-known that psoriasis pathogenesis is driven by aberrant production of proinflammatory cytokines, the mechanisms underlying the imbalance between proinflammatory and anti-inflammatory cytokine expression are incompletely understood. In this study, we report that the transcriptional coregulators CtBP1 and 2 can transactivate a common set of proinflammatory genes both in the skin of imiquimod-induced mouse psoriasis model and in human keratinocytes and macrophages stimulated by imiquimod.

The COVID-19 pandemic and weight management: Effective behaviors and pandemic-specific risk factors.

Background: Obesity and comorbid conditions are associated with worse outcomes related to COVID-19. Moreover, social distancing adherence during the COVID-19 pandemic may predict weight gain due to decreased physical activity, increased emotional eating, and social isolation. While early studies suggest that many individuals struggled with weight management during the pandemic, less is known about healthy eating and weight control behaviors among those enrolled in weight loss programs.

Detection of Environmental Spread of SARS-CoV-2 and Associated Patient Characteristics.

Understanding transmission of severe acute respiratory syndrome coronavirus 2 informs infection prevention practices. Air sampling devices were placed in patient hospital rooms for consecutive collections with and without masks. With patient mask use, no virus was detected in the room.

Field-Portable Microplastic Sensing in Aqueous Environments: A Perspective on Emerging Techniques.

Microplastics (MPs) have been found in aqueous environments ranging from rural ponds and lakes to the deep ocean. Despite the ubiquity of MPs, our ability to characterize MPs in the environment is limited by the lack of technologies for rapidly and accurately identifying and quantifying MPs. Although standards exist for MP sample collection and preparation, methods of MP analysis vary considerably and produce data with a broad range of data content and quality.

High contrast cleavage detection.

Photonic biosensors that use optical resonances to amplify signals from refractive index changes offer high sensitivity, real-time readout, and scalable, low-cost fabrication. However, when used with classic affinity assays, they struggle with noise from nonspecific binding and are limited by the low refractive index and small size of target biological molecules. In this Letter, we evaluate the performance of an integrated microring photonic biosensor using the high contrast cleavage detection (HCCD) mechanism, which we recently introduced.

An Entrustable Professional Activities Model for Assessment of Undergraduate Competence in Anesthesia and Surgery: Performance of the Assessment Scheme and the Impact of Assessment Timing and Variation in Structure of Teaching Activities on Student Outcomes.

An entrustable professional activity (EPA) model was used to assess the anesthesia and surgery competence of year 4 students during elective neutering procedures over 3 academic years (cohort A, cohort B, and cohort C). Two competence thresholds were defined by an expert panel, the minimum acceptable standard (MAS) and the standard expected at the start of final-year rotations (SFR). The assessment scheme performed as expected, and the median level of supervision achieved by students either matched or exceeded the SFR for all EPAs except one, which matched the MAS.

Directed Evolution of an Improved Aminoacyl-tRNA Synthetase for Incorporation of L-3,4-Dihydroxyphenylalanine (L-DOPA).

The catechol group of 3,4-dihydroxyphenylalanine (L-DOPA) derived from L-tyrosine oxidation is a key post-translational modification (PTM) in many protein biomaterials and has potential as a bioorthogonal handle for precision protein conjugation applications such as antibody-drug conjugates. Despite this potential, indiscriminate enzymatic modification of exposed tyrosine residues or complete replacement of tyrosine using auxotrophic hosts remains the preferred method of introducing the catechol moiety into proteins, which precludes many protein engineering applications. We have developed new orthogonal translation machinery to site-specifically incorporate L-DOPA into recombinant proteins and a new fluorescent biosensor to selectively monitor L-DOPA incorporation in vivo.

Atypical Ulcerative Necrobiosis Lipoidica Diabeticorum: A Case Study.

Necrobiosis lipoidica is a granulomatous skin condition commonly associated with diabetes. When associated with diabetes mellitus, the name expands to necrobiosis lipoidica diabeticorum (NLD). In these patients, this rare condition has an incidence rate of 0.