Establishment of Bovine 3D Enteroid-derived 2D monolayers

Three-dimensional (3D) intestinal enteroids are powerful in vitro models for studying intestinal biology. However, due to their closed structure direct access to the apical surface is impeded, limiting high-throughput applications of exogenous compounds and pathogens. In this study, we describe a method for generating confluent 2D enteroids from single-cell suspensions of enzymatically-dissociated ileum-derived bovine 3D enteroids. Confluent monolayers were first achieved using IntestiCult media but to establish a defined, cost-effective culture media, we also developed a bovine enteroid monolayer (BEM) medium. The monolayers cultured in BEM media proliferated extensively and formed confluent cell layers on both Matrigel-coated plastic plates and transwell inserts by day 3 of culture. The 2D enteroids maintained the epithelial cell lineages found in 3D enteroids and ileum tissue. In addition, the monolayers formed a functional epithelial barrier based on the presence of the adherens and tight junction proteins, E-cadherin and ZO-1, and electrical resistance across the monolayer was measured from day 3 and maintained for up to 7 days in culture. The method described here will provide a useful model to study bovine epithelial cell biology with ease of access to the apical surface of epithelial cells and has potential to investigate host–pathogen interactions and screen bioactive compounds

Practitioners’ perspectives on quality of life in aphasia rehabilitation in Denmark

Objective: This study reports on Danish speech and language therapists’ knowledge and understanding of quality of life (QOL) in aphasia, including therapists’ views on education and training in relation to preparedness for working on QOL, use of measures, and barriers to applying QOL in practice. Methods: 14 Danish clinicians completed a 48-item online questionnaire regarding their views, perspectives and practices that included multiple-choice questions, rating scales, and boxes permitting free text responses. Descriptive statistics were used to characterize the numerical data, and content analysis was applied to text responses. Results: The clinicians interpreted QOL as subjective wellbeing and participation, and explored it with most clients and relatives using informal methods, primarily conversation, for the purposes of identifying relevant goals to direct treatment. Clinicians perceived a need for greater theoretical, practical and experiential knowledge regarding QOL. They also identified a need for translated QOL instruments and training in these measures in practice. Conclusion: Despite a reported lack of knowledge about and tools for measuring QOL, Danish clinicians are applying QOL issues in their practice and perceive these issues as valuable and important in assessment and therapy. The findings have clear implications for tool development, and workforce education

Visualizing insulin vesicle neighborhoods in β cells by cryo-electron tomography

Subcellular neighborhoods, comprising specific ratios of organelles and proteins, serve a multitude of biological functions and are of particular importance in secretory cells. However, the role of subcellular neighborhoods in insulin vesicle maturation is poorly understood. Here, we present single-cell multiple distinct tomogram acquisitions of β cells for in situ visualization of distinct subcellular neighborhoods that are involved in the insulin vesicle secretory pathway. We propose that these neighborhoods play an essential role in the specific function of cellular material. In the regions where we observed insulin vesicles, a measurable increase in both the fraction of cellular volume occupied by vesicles and the average size (diameter) of the vesicles was apparent as sampling moved from the area near the nucleus toward the plasma membrane. These findings describe the important role of the nanometer-scale organization of subcellular neighborhoods on insulin vesicle maturation

Visualizing insulin vesicle neighborhoods in β cells by cryo-electron tomography

Subcellular neighborhoods, comprising specific ratios of organelles and proteins, serve a multitude of biological functions and are of particular importance in secretory cells. However, the role of subcellular neighborhoods in insulin vesicle maturation is poorly understood. Here, we present single-cell multiple distinct tomogram acquisitions of β cells for in situ visualization of distinct subcellular neighborhoods that are involved in the insulin vesicle secretory pathway. We propose that these neighborhoods play an essential role in the specific function of cellular material. In the regions where we observed insulin vesicles, a measurable increase in both the fraction of cellular volume occupied by vesicles and the average size (diameter) of the vesicles was apparent as sampling moved from the area near the nucleus toward the plasma membrane. These findings describe the important role of the nanometer-scale organization of subcellular neighborhoods on insulin vesicle maturation

Cardiorespiratory fitness assessment using risk-stratified exercise testing and dose-response relationships with disease outcomes.

Cardiorespiratory fitness (CRF) is associated with mortality and cardiovascular disease, but assessing CRF in the population is challenging. Here we develop and validate a novel framework to estimate CRF (as maximal oxygen consumption, VO2max) from heart rate response to low-risk personalised exercise tests. We apply the method to examine associations between CRF and health outcomes in the UK Biobank study, one of the world's largest and most inclusive studies of CRF, showing that risk of all-cause mortality is 8% lower (95%CI 5-11%, 2670 deaths among 79,981 participants) and cardiovascular mortality is 9% lower (95%CI 4-14%, 854 deaths) per 1-metabolic equivalent difference in CRF. Associations obtained with the novel validated CRF estimation method are stronger than those obtained using previous methodology, suggesting previous methods may have underestimated the importance of fitness for human health

Topical Administration of a Soluble TNF Inhibitor Reduces Infarct Volume After Focal Cerebral Ischemia in Mice

Tumor necrosis factor, which exists both as a soluble (solTNF) and a transmembrane (tmTNF) protein, plays an important role in post-stroke inflammation. The objective of the present study was to test the effect of topical versus intracerebroventricular administration of XPro1595 (a solTNF inhibitor) and etanercept (a solTNF and tmTNF inhibitor) compared to saline on output measures such as infarct volume and post-stroke inflammation in mice. Adult male C57BL/6 mice were treated topically (2.5 mg/ml/1μl/h for 3 consecutive days) or intracerebroventricularly (1.25 mg/kg/0.5 ml, once) with saline, XPro1595, or etanercept immediately after permanent middle cerebral artery occlusion (pMCAO). Mice were allowed to survive 1 or 3 days. Infarct volume, microglial and leukocyte profiles, and inflammatory markers were evaluated. We found that topical, and not intracerebroventricular, administration of XPro1595 reduced infarct volume at both 1 and 3 days after pMCAO. Etanercept showed no effect. We observed no changes in microglial or leukocyte populations. XPro1595 increased gene expression of at 1 day and at 1 and 3 days, while decreasing expression at 1 and 3 days after pMCAO, suggesting a change in microglial activation toward a phagocytic phenotype. Our data demonstrate that topical administration of XPro1595 for 3 consecutive days decreases infarct volumes after ischemic stroke, while modifying microglial activation and the inflammatory response post-stroke. This suggests that inhibitors of solTNF hold great promise for future neuroprotective treatment in ischemic stroke

Возникновение и развитие еврейской прессы Крыма

В статье выделяются основные этапы процесса возникновения и развития еврейской прессы Крыма, вводится в научный оборот ряд еврейских изданий.У статті виділяються основні етапи процесу виникнення і розвитку єврейської преси Криму, вводиться в науковий обіг ряд єврейських видань.The article researches the Jewish Crimean mass-media