Forensic profiling of smokeless powders (SLPs) by gas chromatography-mass spectrometry (GC-MS): a systematic investigation into injector conditions and their effect on the characterisation of samples.

Smokeless powders (SLPs) are composed of a combination of thermolabile and non-thermolabile compounds. When analysed by GC-MS, injection conditions may therefore play a fundamental role on the characterisation of forensic samples. However, no systematic investigations have ever been carried out. This casts doubt on the optimal conditions that should be adopted in advanced profiling applications (e.g. class attribution and source association), especially when a traditional split/splitless (S/SL) injector is used. Herein, a study is reported that specifically focused on the evaluation of the liner type (Ltype) and inlet temperature (Tinj). Results showed that both could affect the exhaustiveness and repeatability of the observed chemical profiles, with Ltype being particularly sensitive despite typically not being clarified in published works. Perhaps as expected, degradation effects were observed for the most thermolabile compounds (e.g. nitroglycerin) at conditions maximising the heat transfer rates (Ltype = packed and Tinj ≥ 200 °C). However, these did not seem to be as influential as, perhaps, suggested in previous studies. Indeed, the harshest injection conditions in terms of heat transfer rate (Ltype = packed and Tinj = 260 °C) were found to lead to better performances (including better overall %RSDs and LODs) compared to the mildest ones. This suggested that implementing conditions minimising heat-induced breakdowns during injection was not necessarily a good strategy for comparison purposes. The reported findings represent a concrete step forward in the field, providing a robust body of data for the development of the next generation of SLP profiling methods.

Transport of acylated anthocyanins by the Arabidopsis ATP-binding cassette transporters AtABCC1, AtABCC2, and AtABCC14.

Anthocyanins are a group of pigments that have various roles in plants including attracting pollinators and seed dispersers and protecting against various types of stress. In vegetative tissue, these anthocyanins are sequestered in the vacuole following biosynthesis in the cytoplasm, though there remain questions as to the events leading to the vacuolar sequestration. In this study, we were able to show that the uptake of acylated anthocyanins by vacuolar membrane-enriched vesicles isolated from Arabidopsis was stimulated by the addition of MgATP and was inhibited by both vanadate and glybenclamide, but not by gramicidin D or bafilomycin A1 , suggesting that uptake involves an ATP-binding cassette (ABC) transporter and not an H+ -antiporter. Membrane vesicles isolated from yeast expressing the ABC transporters designated AtABCC1, AtABCC2, and AtABCC14 are capable of MgATP-dependent uptake of acylated anthocyanins. This uptake was not dependent on glutathione as seen previously for anthocyanidin 3-O-monoglucosides. Compared to the wild-type, the transport of acylated anthocyanins was lower in vacuolar membrane-enriched vesicles isolated from atabcc1 cell cultures providing evidence that AtABCC1 may be the predominant transporter of these compounds in vivo. In addition, the pattern of anthocyanin accumulation differed between the atabcc1, atabcc2, and atabcc14 mutants and the wild-type seedlings under anthocyanin inductive conditions. We suggest that AtABCC1, AtABCC2, and AtABCC14 are involved in the vacuolar transport of acylated anthocyanins produced in the vegetative tissue of Arabidopsis and that the pattern of anthocyanin accumulation can be altered depending on the presence or absence of a specific vacuolar ABC transporter.

Checkpoint Inhibition Reduces the Threshold for Drug-Specific T-Cell Priming and Increases the Incidence of Sulfasalazine Hypersensitivity.

An emerging clinical issue associated with immune-oncology agents is the collateral effects on the tolerability of concomitant medications. One report of this phenomenon was the increased incidence of hypersensitivity reactions observed in patients receiving concurrent immune checkpoint inhibitors (ICIs) and sulfasalazine (SLZ). Thus, the aim of this study was to characterize the T cells involved in the pathogenesis of such reactions, and recapitulate the effects of inhibitory checkpoint blockade on de-novo priming responses to compounds within in vitro platforms. A regulatory competent human dendritic cell/T-cell coculture assay was used to model the effects of ICIs on de novo nitroso sulfamethoxazole- and sulfapyridine (SP) (the sulfonamide component of SLZ) hydroxylamine-specific priming responses. The role of T cells in the pathogenesis of the observed reactions was explored in 3 patients through phenotypic characterization of SP/sulfapyridine hydroxylamine (SPHA)-responsive T-cell clones (TCC), and assessment of cross-reactivity and pathways of T-cell activation. Augmentation of the frequency of responding drug-specific T cells and intensity of the T-cell response was observed with PD-1/PD-L1 blockade. Monoclonal populations of SP- and SPHA-responsive T cells were isolated from all 3 patients. A core secretory effector molecule profile (IFN-γ, IL-13, granzyme B, and perforin) was identified for SP and SPHA-responsive TCC, which proceeded through Pi and hapten mechanisms, respectively. Data presented herein provides evidence that drug-responsive T cells are effectors of hypersensitivity reactions observed in oncology patients administered ICIs and SLZ. Perturbation of drug-specific T-cell priming is a plausible explanation for clinical observations of how an increased incidence of these adverse events is occurring.

Metastatic Cardiac Tumor Presenting as an Anteroseptal ST-Segment Elevation Myocardial Infarction in a Young Male.

In the appropriate clinical context, ST-segment elevation on electrocardiogram (ECG) necessitates prompt evaluation for coronary artery occlusion requiring reperfusion with percutaneous coronary intervention. Conversely, the etiology of ST-segment elevation may be representative of an alternative diagnosis other than myocardial infarction. We report the case of a patient with a history of primary bone sarcoma who presented for further evaluation of a large pericardial effusion identified on an outpatient echocardiogram and was found to have ST-segment elevation on ECG in the absence of any cardiopulmonary symptoms. The ECG abnormalities were attributed to a likely persistent current of injury resulting from a mass in the interventricular septum, likely representative of a metastatic lesion of his known malignancy. This case highlights the importance of maintaining a broad differential for ST-segment elevation, particularly in patients without symptoms of angina and those with a history of aggressive or relapsing cancer to minimize the morbidity and mortality of invasive procedures.

Deletion of Exon 1 in AMER1 in Osteopathia Striata with Cranial Sclerosis.

Osteopathia striata with cranial sclerosis (OSCS) is an X-linked dominant condition characterised by metaphyseal striations, macrocephaly, cleft palate, and developmental delay in affected females. Males have a more severe phenotype with multi-organ malformations, and rarely survive. To date, only frameshift and nonsense variants in exon 2, the single coding exon of AMER1, or whole gene deletions have been reported to cause OSCS. In this study, we describe two families with phenotypic features typical of OSCS. Exome sequencing and multiplex ligation-dependent probe amplification (MLPA) did not identify pathogenic variants in AMER1. Therefore, genome sequencing was employed which identified two deletions containing the non-coding exon 1 of AMER1 in the families. These families highlight the importance of considering variants or deletions of upstream non-coding exons in conditions such as OSCS, noting that often such exons are not captured on probe or enrichment-based platforms because of their high G/C content.

Histone H3.3 beyond cancer: Germline mutations in Histone 3 Family 3A and 3B cause a previously unidentified neurodegenerative disorder in 46 patients.

Although somatic mutations in Histone 3.3 (H3.3) are well-studied drivers of oncogenesis, the role of germline mutations remains unreported. We analyze 46 patients bearing de novo germline mutations in histone 3 family 3A (H3F3A) or H3F3B with progressive neurologic dysfunction and congenital anomalies without malignancies. Molecular modeling of all 37 variants demonstrated clear disruptions in interactions with DNA, other histones, and histone chaperone proteins. Patient histone posttranslational modifications (PTMs) analysis revealed notably aberrant local PTM patterns distinct from the somatic lysine mutations that cause global PTM dysregulation. RNA sequencing on patient cells demonstrated up-regulated gene expression related to mitosis and cell division, and cellular assays confirmed an increased proliferative capacity. A zebrafish model showed craniofacial anomalies and a defect in Foxd3-derived glia. These data suggest that the mechanism of germline mutations are distinct from cancer-associated somatic histone mutations but may converge on control of cell proliferation.

The occurrence and distribution of polycyclic aromatic hydrocarbons, bisphenol A and organophosphate flame retardants in indoor dust and soils from public open spaces: Implications for human exposure.

Global concern exists regarding human exposure to organic pollutants derived from public open spaces and indoor dust. This study has evaluated the occurrence of 18 polycyclic aromatic hydrocarbons (PAHs), 11 organophosphorus flame retardants (OPFRs) and bisphenol A (BPA). To achieve this, a new simple, efficient and fast multi-residue analytical method based on a fully automated pressurised liquid extraction (PLE) and subsequent quantification by gas chromatography coupled to electron ionization-mass spectrometry (GC-EI-MS) in selected ion monitoring (SIM) mode was developed. The developed method was applied to indoor dust (12 sampling households) and soil derived from two public open spaces (POSs). Among all compounds studied, PAHs were the most ubiquitous contaminants detected in POS soils and indoor dust although some OPFRs and BPA were detected in lower concentrations. An assessment of the incremental lifetime cancer risk (ILCR) was done and indicated a high potential cancer risk from the POS sites and some of the indoor dust sampled sites. However, key variables, such as the actual exposure duration, frequency of contact and indoor cleaning protocols will significantly reduce the potential risk. Finally, the ingestion of soils and indoor dust contaminated with OPFRs and BPA was investigated and noted in almost all cases to be below the USEPA reference doses.

Guidance for the prevention and emergency management of adult patients with adrenal insufficiency.

Adrenal insufficiency (AI) is an often-unrecognised endocrine disorder, which can lead to adrenal crisis and death if not identified and treated. Omission of steroids in patients with AI, particularly during physiological stress such as an intercurrent illness or surgery, can also lead to an adrenal crisis. The National Reporting and Learning System (NRLS) identified 78 incidents including two deaths and six incidents of severe harm to patients in a recent 4-year period. This guidance will go through causes of adrenal insufficiency, groups at risk of an adrenal crisis, emergency management and management for surgical procedures. A new NHS Steroid Emergency Card has been developed to be carried by patients at risk of adrenal crisis. We hope the new emergency card and this guidance will increase awareness of the need to start steroids promptly in patients at risk of an adrenal crisis, particularly those presenting in the emergency department or to acute medicine teams and those undergoing surgery or invasive procedures.

Detection of ß-alanyl aminopeptidase as a biomarker for Pseudomonas aeruginosa in the sputum of patients with cystic fibrosis using exogenous volatile organic compound evolution.

A novel, rapid and sensitive analytical method has been developed and applied to 105 sputum samples from patients with cystic fibrosis, including 5 samples from post-lung transplant patients. This new method is specifically targeted to measure ß-alanyl aminopeptidase activity which is characteristic of some important Gram-negative pathogens. Of relevance to this study are Pseudomonas aeruginosa and pathogens of the Burkholderia cepacia complex both of which are commonly associated with respiratory infections as well as increased morbidity and mortality in adult cystic fibrosis patients. The analytical method involves the addition of a novel enzyme substrate (i.e. 3-amino-N-(3-fluorophenyl)propanamide) that interacts with ß-alanyl aminopeptidase to generate an exogenous volatile organic compound 3-fluoroaniline (LOD 0.02 µg mL-1; LOQ 0.06 µg mL-1). 3-Fluoroaniline was determined at 20 times above its calculated limit of quantification in the sputum samples by HS-SPME-GC-MS and then the results compared with standard culture methods and bacterial identification using MALDI-TOF-MS. Detection of 3-fluoroaniline was possible after only 8 h incubation of the sputum samples with a 95% success rate; this increased to 100% at 24 h which was well within the typical routine timeframe of 48 h. To our knowledge, this is the first demonstration of detection of P. aeruginosa by use of a custom-designed substrate to liberate a detectable and unique VOC. The very high negative predictive value (100% in this study) means such an assay could be appropriate as a screening technique for patients who are not yet colonized by this pathogen.

Amino/Amido Conjugates Form to Nanoscale Cobalt Physiometacomposite (PMC) Materials Functionally Delivering Nucleic Acid Therapeutic to Nucleus Enhancing Anticancer Activity via Ras-Targeted Protein Interference.

Aberrant splicing and protein interaction of Ras binding domain (RBD) are associated with melanoma drug resistance. Here, cobalt or nickel doped zinc oxide (ZnO) physiometacomposite (PMC) materials bind to RNA and peptide shown by Ninhydrin staining, UV-vis, Fourier transform infrared, and circular dichroism spectroscopy. PMCs deliver splice switching oligomer (SSO) into melanoma cells or 3-D tumor spheroids shown by flow cytometry, fluorescence microscopy, and bioluminescence. Stability in serum, liver, or tumor homogenate up to 48 h and B16F10 melanoma inhibition ≥98-99% is shown. These data suggest preclinical potential of PMC for delivery of SSO, RBD, or other nucleic acid therapeutic and anticancer peptides.

The CHD8 overgrowth syndrome: A detailed evaluation of an emerging overgrowth phenotype in 27 patients.

CHD8 has been reported as an autism susceptibility/intellectual disability gene but emerging evidence suggests that it additionally causes an overgrowth phenotype. This study reports 27 unrelated patients with pathogenic or likely pathogenic CHD8 variants (25 null variants, two missense variants) and a male:female ratio of 21:6 (3.5:1, p < .01). All patients presented with intellectual disability, with 85% in the mild or moderate range, and 85% had a height and/or head circumference ≥2 standard deviations above the mean, meeting our clinical criteria for overgrowth. Behavioral problems were reported in the majority of patients (78%), with over half (56%) either formally diagnosed with an autistic spectrum disorder or described as having autistic traits. Additional clinical features included neonatal hypotonia (33%), and less frequently seizures, pes planus, scoliosis, fifth finger clinodactyly, umbilical hernia, and glabellar hemangioma (≤15% each). These results suggest that, in addition to its established link with autism and intellectual disability, CHD8 causes an overgrowth phenotype, and should be considered in the differential diagnosis of patients presenting with increased height and/or head circumference in association with intellectual disability.

Secondary Abdominoplasty: Management of the Umbilicus after Prior Stalk Transection.

Patients with prior umbilical stalk detachment, secondary to limited abdominoplasty or prior umbilical hernia repair, may in the future be candidates for full abdominoplasty. In these patients, a circumferential incision around the umbilicus detaches the remaining cutaneous blood supply, prompting concern for umbilical viability. Minimal literature exists to guide clinical decision-making for these patients. Inquiries were made to Louisiana Society of Plastic Surgery members and the Plastic Surgery Education Network online forum of the American Society of Plastic Surgeons. Metrics obtained included patient age, sex, nature of primary surgery, time between umbilical stalk detachment and secondary full abdominoplasty, complications, and whether rectus plication was performed at the time of secondary surgery. Eleven physicians provided complete documentation for 18 cases. All patients healed without evidence of umbilical ischemia or necrosis. Average patient age was 40.6 years. Average interval between procedures was 3.6 years. Sixty-seven percent of patients had the umbilicus delayed before the secondary procedure, with the median delay time being 18 days. Rectus fascia was plicated during secondary surgery in 72 percent of patients. This is the first reported series of patients undergoing full abdominoplasty after prior umbilical stalk detachment. It is also the first time the Plastic Surgery Education Network online forum has been used to collect research data, highlighting its potential as a valuable research tool. The data set was obtained from a wide range of practices, which allows for consideration of various technical solutions when this or other diverse clinical scenarios are encountered. CLINICAL QUESTION/LEVEL OF EVIDENCE:: Therapeutic, V.

Transport of Anthocyanins and other Flavonoids by the Arabidopsis ATP-Binding Cassette Transporter AtABCC2.

Flavonoids have important developmental, physiological, and ecological roles in plants and are primarily stored in the large central vacuole. Here we show that both an ATP-binding cassette (ABC) transporter(s) and an H+-antiporter(s) are involved in the uptake of cyanidin 3-O-glucoside (C3G) by Arabidopsis vacuolar membrane-enriched vesicles. We also demonstrate that vesicles isolated from yeast expressing the ABC protein AtABCC2 are capable of MgATP-dependent uptake of C3G and other anthocyanins. The uptake of C3G by AtABCC2 depended on the co-transport of glutathione (GSH). C3G was not altered during transport and a GSH conjugate was not formed. Vesicles from yeast expressing AtABCC2 also transported flavone and flavonol glucosides. We performed ligand docking studies to a homology model of AtABCC2 and probed the putative binding sites of C3G and GSH through site-directed mutagenesis and functional studies. These studies identified residues important for substrate recognition and transport activity in AtABCC2, and suggest that C3G and GSH bind closely, mutually enhancing each other's binding. In conclusion, we suggest that AtABCC2 along with possibly other ABCC proteins are involved in the vacuolar transport of anthocyanins and other flavonoids in the vegetative tissue of Arabidopsis.

Clinical exome sequencing reveals locus heterogeneity and phenotypic variability of cohesinopathies.

PURPOSE: Defects in the cohesin pathway are associated with cohesinopathies, notably Cornelia de Lange syndrome (CdLS). We aimed to delineate pathogenic variants in known and candidate cohesinopathy genes from a clinical exome perspective. METHODS: We retrospectively studied patients referred for clinical exome sequencing (CES, N = 10,698). Patients with causative variants in novel or recently described cohesinopathy genes were enrolled for phenotypic characterization. RESULTS: Pathogenic or likely pathogenic single-nucleotide and insertion/deletion variants (SNVs/indels) were identified in established disease genes including NIPBL (N = 5), SMC1A (N = 14), SMC3 (N = 4), RAD21 (N = 2), and HDAC8 (N = 8). The phenotypes in this genetically defined cohort skew towards the mild end of CdLS spectrum as compared with phenotype-driven cohorts. Candidate or recently reported cohesinopathy genes were supported by de novo SNVs/indels in STAG1 (N = 3), STAG2 (N = 5), PDS5A (N = 1), and WAPL (N = 1), and one inherited SNV in PDS5A. We also identified copy-number deletions affecting STAG1 (two de novo, one of unknown inheritance) and STAG2 (one of unknown inheritance). Patients with STAG1 and STAG2 variants presented with overlapping features yet without characteristic facial features of CdLS. CONCLUSION: CES effectively identified disease-causing alleles at the mild end of the cohensinopathy spectrum and enabled characterization of candidate disease genes.

Structural and functional roles of ether lipids.

Ether lipids, such as plasmalogens, are peroxisome-derived glycerophospholipids in which the hydrocarbon chain at the sn-1 position of the glycerol backbone is attached by an ether bond, as opposed to an ester bond in the more common diacyl phospholipids. This seemingly simple biochemical change has profound structural and functional implications. Notably, the tendency of ether lipids to form non-lamellar inverted hexagonal structures in model membranes suggests that they have a role in facilitating membrane fusion processes. Ether lipids are also important for the organization and stability of lipid raft microdomains, cholesterol-rich membrane regions involved in cellular signaling. In addition to their structural roles, a subset of ether lipids are thought to function as endogenous antioxidants, and emerging studies suggest that they are involved in cell differentiation and signaling pathways. Here, we review the biology of ether lipids and their potential significance in human disorders, including neurological diseases, cancer, and metabolic disorders.

Mechanistic differences in the uptake of salicylic acid glucose conjugates by vacuolar membrane-enriched vesicles isolated from Arabidopsis thaliana.

Salicylic acid (SA) is a plant hormone involved in a number of physiological responses including both local and systemic resistance of plants to pathogens. In Arabidopsis, SA is glucosylated to form either SA 2-O-ß-d-glucose (SAG) or SA glucose ester (SGE). In this study, we show that SAG accumulates in the vacuole of Arabidopsis, while the majority of SGE was located outside the vacuole. The uptake of SAG by vacuolar membrane-enriched vesicles isolated from Arabidopsis was stimulated by the addition of MgATP and was inhibited by both vanadate (ABC transporter inhibitor) and bafilomycin A1 (vacuolar H+ -ATPase inhibitor), suggesting that SAG uptake involves both an ABC transporter and H+ -antiporter. Despite its absence in the vacuole, we observed the MgATP-dependent uptake of SGE by Arabidopsis vacuolar membrane-enriched vesicles. SGE uptake was not inhibited by vanadate but was inhibited by bafilomycin A1 and gramicidin D providing evidence that uptake was dependent on an H+ -antiporter. The uptake of both SAG and SGE was also inhibited by quercetin and verapamil (two known inhibitors of multidrug efflux pumps) and salicin and arbutin. MgATP-dependent SAG and SGE uptake exhibited Michaelis-Menten-type saturation kinetics. The vacuolar enriched-membrane vesicles had a 46-fold greater affinity and a 10-fold greater transport activity with SGE than with SAG. We propose that in Arabidopsis, SAG is transported into the vacuole to serve as a long-term storage form of SA while SGE, although also transported into the vacuole, is easily hydrolyzed to release the active hormone which can then be remobilized to other cellular locations.

Analysis of Listeria using exogenous volatile organic compound metabolites and their detection by static headspace-multi-capillary column-gas chromatography-ion mobility spectrometry (SHS-MCC-GC-IMS).

Listeria monocytogenes is a Gram-positive bacterium and an opportunistic food-borne pathogen which poses significant risk to the immune-compromised and pregnant due to the increased likelihood of acquiring infection and potential transmission of infection to the unborn child. Conventional methods of analysis suffer from either long turn-around times or lack the ability to discriminate between Listeria spp. reliably. This paper investigates an alternative method of detecting Listeria spp. using two novel enzyme substrates that liberate exogenous volatile organic compounds in the presence of α-mannosidase and D-alanyl aminopeptidase. The discriminating capabilities of this approach for identifying L. monocytogenes from other species of Listeria are investigated. The liberated volatile organic compounds (VOCs) are detected using an automated analytical technique based on static headspace-multi-capillary column-gas chromatography-ion mobility spectrometry (SHS-MCC-GC-IMS). The results obtained by SHS-MCC-GC-IMS are compared with those obtained by the more conventional analytical technique of headspace-solid phase microextraction-gas chromatography-mass spectrometry (HS-SPME-GC-MS). The results found that it was possible to differentiate between L. monocytogenes and L. ivanovii, based on their VOC response from α-mannosidase activity.

Existing and Future Educational Needs in Graduate and Postgraduate Education.

INTRODUCTION: This review was designed to make recommendations on future educational needs, principles of curricular development, and how the International Society for Sexual Medicine (ISSM) should address the need to enhance and promote human sexuality education around the world. AIM: To explore the ways in which graduate and postgraduate medical education in human sexuality has evolved and is currently delivered. METHODS: We reviewed existing literature concerning sexuality education, curriculum development, learning strategies, educational formats, evaluation of programs, evaluation of students, and faculty development. We reviewed literature relating to four main areas: (i) the current status of the international regulation of training in sexual medicine; (ii) the current delivery of education and training in sexual medicine; (iii) resident and postgraduate education in sexual medicine surgery; and (iv) education and training for allied health professionals. RESULTS: The main findings in these four areas are as follows. Sexual medicine has grown considerably as a specialty during the past 20 years, with many drivers being identified. However, the regulatory aspects of training, assessment, and certification are currently in the early stages of development and are in many ways lagging behind the scientific and clinical knowledge in the field. However, there are examples of the development of curricula with accompanying assessments that have attempted to set standards of education and training that might underlie the delivery of high-quality care to patients in sexual medicine. The development of competence assessment has been applied to surgical training in sexual medicine, and there is increasing interest in simulation as a means of enhancing technical skills training. Although the focus of curriculum development has largely been the medical profession, there is early interest in the development of standards for training and education of allied health professionals. CONCLUSION: Organizations of professionals in sexual health, such as the ISSM, have an opportunity, and indeed a responsibility, to provide and disseminate learning opportunities, curricula, and standards of training for doctors and allied health professionals in sexual medicine. Eardley I, Reisman Y, Goldstein S, et al. Existing and Future Educational Needs in Graduate and Postgraduate Education. J Sex Med 2017;14:475-485.