Characterisation of extracellular vesicles (EVs) from dysfunctional endothelial cells following exposure to alcohol

Endothelial cell (EC) dysfunction is classically associated with subclinical atherosclerosis and the development of atheroprone lesions. Extracellular vesicles (EVs) are biological nanoparticles secreted by most cell types that contain a complex mix of cell-type specific signalling proteins and nucleic acids. At high concentrations, alcohol is a known risk factor for atherosclerosis but may be protective at lower concentrations. In this study, we first investigated whether alcohol (EtOH) at varying concentrations can alter the number and/or phenotype of EVs released from endothelial cells before the proteomic profile of endothelial cell-derived EVs was assessed. Human aortic endothelial (HAEC) and smooth aortic vascular muscle cells (HuSMCs) were first characterised for lineage specific markers before endothelial cell dysfunction was induced following treatment of HAECs with low and high doses of EtOH. EVs released from HAEC were isolated from conditioned cell media, enumerated and characterised using dynamic light scattering (DLS), immunoblot and AmnisTM Cell Stream FACS analysis for EV biomarkers before the proteomic profile of the EVs was evaluated using Liquid chromatography massspectrometry (LC-MS) analysis. The data generated suggest that the number of endothelial derived-EVs released following endothelial dysfunction increases following exposure of cells to different concentrations of EtOH. Moreover, the protein cargo within these EVs varies, even though the EVs originated from the same cell source. Proteomic and bioinformatic analysis revealed that protein metabolism and cell growth and/or maintenance are the key pathways and biological processes associated with these EVs and provides further insight into elucidating their putative role following endothelial dysfunction. Lastly, EVs from dysfunctional HAECs had little effect on the growth and migration of HuSMCs but did promote HuSMC de-differentiation following treatment with 25-EXO. These data highlight the potential important role for EVs released following endothelial dysfunction on the underlying smooth muscle cell population that may contribute to arteriosclerotic lesion formatio

Mentalisation Amongst Maternal and Child Health Nurses Using the Newborn Behavioural Observations With Infant-mother dyads: A Qualitative Study

© 2020 Elsevier Inc. Purpose: This study explored Maternal and Child Health Nurses\u27 (MCHN) mentalisation processes towards infant-mother dyads when using the Newborn Behavioural Observations (NBO) system in practice. Design and methods: Ten Australian MCHNs (female; aged 31–66 years), who had used the NBO clinically within the last 12 months, were recruited from a database of NBO-trained practitioners. Interpretative phenomenological analysis of one-on-one semi-structured interviews explored MCHNs experiential meaning-making. Results: Analysis of the data produced four main themes: reflections regarding the dyad, personal reflections, reflection into action, and professional identity and future practice. MCHNs reported that the NBO\u27s focus on the pre-verbal infant provided them with an added dynamic to consultations outside of the practitioner-caregiver relationship. Thus, they were able to provide holistic and collaborative relationship support to infant-mother dyads. Emotional satisfaction and pride in profession were also reported; in current literature, these factors have been found to reduce burnout in primary care providers. Conclusions: The NBO appears to promote practitioner mentalisation, offering MCHNs a framework and confidence to apply infant mental health theory practically. Practice implications: The NBO offers potential benefits to child and family health nursing practice, and other primary care providers, who offer infant mental health and relationship support as part of their work with families in the first three months. The NBO also provides a means to shift from prescriptive to mentalisation-based, infant-inclusive, and individualised practice

Synergistic activity of endochitinase and exochitinase from Trichoderma atroviride (T. harzianum) against the pathogenic fungus (Venturia inaequalis) in transgenic apple plants

Genes from the biocontrol fungus Trichoderma atroviride encoding the antifungal proteins endochitinase or exochitinase (N-acetyl-β-D-hexosaminidase) were inserted into ‘Marshall McIntosh' apple singly and in combination. The genes were driven by a modified CaMV35S promoter. The resulting plants were screened for resistance to Venturia inaequalis, the causal agent of apple scab, and for effects of enzyme expression on growth. Disease resistance was correlated with the level of expression of either enzyme when expressed alone but exochitinase was less effective than endochitinase. The level of expression of endochitinase was negatively correlated with plant growth while exochitinase had no consistent effect on this character. Plants expressing both enzymes simultaneously were more resistant than plants expressing either single enzyme at the same level; analyses indicated that the two enzymes acted synergistically to reduce disease. Selected lines, especially one expressing low levels of endochitinase activity and moderate levels of exochitinase activity, were highly resistant in growth chamber trials and had negligible reduction in vigor relative to control plants. We believe that this is the first report of resistance in plants induced by expression of an N-acetylhexosaminidase and is the first report of in planta synergy between an exochitinase and an endochitinas

Insect-Nematode-Red Pine Association in Western Maryland with Major Emphasis on Bursaphelenchus xylophilus and Monochamus spp.

Western Maryland red pines, Pinus resinosa Ait. Were examined over three years, 1982-1984, to determine the distribution of Bursaphelenchus xylophilus, the pinewood nematode. Ca. 47-year-old (older) and ca. 25-year-old (younger) trees were subdivided into the following categories: (1) trees with mostly green needles; (2) trees with mostly reddish-brown needles; (3) trees with no needles but with bark intact; (4) trees with no bark; and (5) trees with chlorotic, bleached-green needles. The pinewood. nematode infected 76.5% of older red pines and 68% of younger red pines. The nematode was not evenly distributed in trees in any tree decadence category or by tree age. Bursaphelenchus xylophilus infected 15.1% of the samples from trunk, primary and secondary branches in older red pines and 18.5% in younger red pines. By tree decadence category older trees had the highest infection (25.2%) in green needled trees (category 1) whereas younger trees had the highest infection (28.7%) in bleached-green needled trees (category 5). Trees collected in late May-early June 1982 lacked Monochamus rearings and belonged to categories 1, 2, 3 and 4. Trees from these categories had emergences of Ips spp., Tetropium schwarzianum Casey, Pissodes approximatus Hopkins and Otiorhynchus spp. in summer 1982. From bleached-green needled red pines (category 5) collected in late July-early August 1983 two Monochamus species emerged in 1984, M. carolinensis (Olivier) and M. scutellatus (Say), as well as other insects including Neacanthocinus pusillus (Kirby) and Amniscus collaris Haldemann. Chrysobothris scabripennis Cast. and Gory emerged from trees cut in 1983, but which remained uncaged in the field until 1984. Bursaphelenchus xylophilus was present in 94% of tracheal systems of both M. scutellatus and M. carolinensis. Pinewood nematode was found infesting 4.2% of N. pusillus specimens. One specimen each off. approximatus, Ips spp. and C. scabripennis were positive for B. xylophilus. Dolichomitus tuberculatue tuberculatus (Geoff.), an ichneumonid parasite, was reared from Monochamus spp. larvae. Two deutonymph mites, Dendrolaelap isodentatus (Hurlbutt) and Trichouropoda hirsuta Hirschmann, were located externally on Monochamus spp. beetles

Human and Non-Human Primate Intestinal FcRn Expression and Immunoglobulin G Transcytosis

PURPOSE: To evaluate transcytosis of immunoglobulin G (IgG) by the neonatal Fc receptor (FcRn) in adult primate intestine to determine whether this is a means for oral delivery of monoclonal antibodies (mAbs). METHODS: Relative regional expression of FcRn and localization in human intestinal mucosa by RT-PCR, ELISA & immunohistochemistry. Transcytosis of full-length mAbs (sandwich ELISA-based detection) across human intestinal segments mounted in Ussing-type chambers, human intestinal (caco-2) cell monolayers grown in transwells, and serum levels after regional intestinal delivery in isoflurane-anesthetized cynomolgus monkeys. RESULTS: In human intestine, there was an increasing proximal-distal gradient of mucosal FcRn mRNA and protein expression. In cynomolgus, serum mAb levels were greater after ileum-proximal colon infusion than after administration to stomach or proximal small intestine (1–5 mg/kg). Serum levels of wild-type mAb dosed into ileum/proximal colon (2 mg/kg) were 124 ± 104 ng/ml (n = 3) compared to 48 ± 48 ng/ml (n = 2) after a non-FcRn binding variant. In vitro, mAb transcytosis in polarized caco-2 cell monolayers and was not enhanced by increased apical cell surface IgG binding to FcRn. An unexpected finding in primate small intestine, was intense FcRn expression in enteroendocrine cells (chromagranin A, GLP-1 and GLP-2 containing). CONCLUSIONS: In adult primates, FcRn is expressed more highly in distal intestinal epithelial cells. However, mAb delivery to that region results in low serum levels, in part because apical surface FcRn binding does not influence mAb transcytosis. High FcRn expression in enteroendocrine cells could provide a novel means to target mAbs for metabolic diseases after systemic administration

Decreased Mitochondrial DNA Mutagenesis in Human Colorectal Cancer

Genome instability is regarded as a hallmark of cancer. Human tumors frequently carry clonally expanded mutations in their mitochondrial DNA (mtDNA), some of which may drive cancer progression and metastasis. The high prevalence of clonal mutations in tumor mtDNA has commonly led to the assumption that the mitochondrial genome in cancer is genetically unstable, yet this hypothesis has not been experimentally tested. In this study, we directly measured the frequency of non-clonal (random) de novo single base substitutions in the mtDNA of human colorectal cancers. Remarkably, tumor tissue exhibited a decreased prevalence of these mutations relative to adjacent non-tumor tissue. The difference in mutation burden was attributable to a reduction in C∶G to T∶A transitions, which are associated with oxidative damage. We demonstrate that the lower random mutation frequency in tumor tissue was also coupled with a shift in glucose metabolism from oxidative phosphorylation to anaerobic glycolysis, as compared to non-neoplastic colon. Together these findings raise the intriguing possibility that fidelity of mitochondrial genome is, in fact, increased in cancer as a result of a decrease in reactive oxygen species-mediated mtDNA damage

Development of a National Pain Management Competency Profile to Guide Entry-Level Physiotherapy Education in Canada

Background National strategies from North America call for substantive improvements in entry-level pain management education to help reduce the burden of chronic pain. Past work has generated a valuable set of interprofessional pain management competencies to guide the education of future health professionals. However, there has been very limited work that has explored the development of such competencies for individual professions in different regions. Developing profession-specific competencies tailored to the local context is a necessary first step to integrate them within local regulatory systems. Our group is working toward this goal within the context of entry-level physiotherapy (PT) programs across Canada. Aims This study aimed to create a consensus-based competency profile for pain management, specific to the Canadian PT context. Methods A modified Delphi design was used to achieve consensus across Canadian university-based and clinical pain educators. Results Representatives from 14 entry-level PT programs (93% of Canadian programs) and six clinical educators were recruited. After two rounds, a total of 15 competencies reached the predetermined endorsement threshold (75%). Most participants (85%) reported being “very satisfied” with the process. Conclusions This process achieved consensus on a novel pain management competency profile specific to the Canadian PT context. The resulting profile delineates the necessary abilities required by physiotherapists to manage pain upon entry to practice. Participants were very satisfied with the process. This study also contributes to the emerging literature on integrated research in pain management by profiling research methodology that can be used to inform related work in other health professions and regions

The calcium binding protein S100β marks hedgehog-responsive resident vascular stem cells within vascular lesions

A hallmark of subclinical atherosclerosis is the accumulation of vascular smooth muscle cell (SMC)-like cells leading to intimal thickening. While medial SMCs contribute, the participation of hedgehog-responsive resident vascular stem cells (vSCs) to lesion formation remains unclear. Using transgenic eGFP mice and genetic lineage tracing of S100β vSCs in vivo, we identified S100β/ Sca1 cells derived from a S100β non-SMC parent population within lesions that co-localise with smooth muscle α-actin (SMA) cells following iatrogenic flow restriction, an effect attenuated following hedgehog inhibition with the smoothened inhibitor, cyclopamine. In vitro, S100β/Sca1 cells isolated from atheroprone regions of the mouse aorta expressed hedgehog signalling components, acquired the di-methylation of histone 3 lysine 4 (H3K4me2) stable SMC epigenetic mark at the Myh11 locus and underwent myogenic differentiation in response to recombinant sonic hedgehog (SHh). Both S100β and PTCH1 cells were present in human vessels while S100β cells were enriched in arteriosclerotic lesions. Recombinant SHh promoted myogenic differentiation of human induced pluripotent stem cell-derived S100β neuroectoderm progenitors in vitro. We conclude that hedgehog-responsive S100β vSCs contribute to lesion formation and support targeting hedgehog signalling to treat subclinical arteriosclerosis

Genomic, Pathway Network, and Immunologic Features Distinguishing Squamous Carcinomas

This integrated, multiplatform PanCancer Atlas study co-mapped and identified distinguishing molecular features of squamous cell carcinomas (SCCs) from five sites associated with smokin