HSV-mediated expression of interleukin-4 in dorsal root ganglion neurons reduces neuropathic pain

BACKGROUND: To examine the role of inflammatory mediators in neuropathic pain, we used a replication-defective genomic herpes simplex virus (HSV)-based vector containing the coding sequence for the anti-inflammatory peptide interleukin (IL)-4 under the transcriptional control of the HSV ICP4 immediate early promoter, vector S4IL4, to express IL-4 in dorsal root ganglion (DRG) neurons in vivo. RESULTS: Subcutaneous inoculation of S4IL4 in the foot transduced lumbar DRG to produce IL-4. Transgene-mediated expression of IL-4 did not alter thermal latency or tactile threshold in normal animals, but inoculation of S4IL4 1 week after spinal nerve ligation (SNL) reduced mechanical allodynia and reversed thermal hyperalgesia resulting from SNL. Inoculation of S4IL4 1 week before SNL delayed the development of thermal hyperalgesia and tactile allodynia, but did not prevent the ultimate development of these manifestations of neuropathic pain. S4IL4 inoculation suppressed non-noxious-induced expression of c-Fos immunoreactivity in dorsal horn of spinal cord and reversed the upregulation of spinal IL-1β, PGE2, and phosphorylated-p38 MAP kinase, characteristic of neuropathic pain. CONCLUSION: HSV-mediated expression of IL-4 effectively reduces the behavioral manifestations of neuropathic pain, and reverses some of the biochemical and histologic correlates of neuropathic pain at the spinal level

An expedited screening platform for the discovery of anti-ageing compounds in vitro and in vivo.

BACKGROUND: Restraining or slowing ageing hallmarks at the cellular level have been proposed as a route to increased organismal lifespan and healthspan. Consequently, there is great interest in anti-ageing drug discovery. However, this currently requires laborious and lengthy longevity analysis. Here, we present a novel screening readout for the expedited discovery of compounds that restrain ageing of cell populations in vitro and enable extension of in vivo lifespan. METHODS: Using Illumina methylation arrays, we monitored DNA methylation changes accompanying long-term passaging of adult primary human cells in culture. This enabled us to develop, test, and validate the CellPopAge Clock, an epigenetic clock with underlying algorithm, unique among existing epigenetic clocks for its design to detect anti-ageing compounds in vitro. Additionally, we measured markers of senescence and performed longevity experiments in vivo in Drosophila, to further validate our approach to discover novel anti-ageing compounds. Finally, we bench mark our epigenetic clock with other available epigenetic clocks to consolidate its usefulness and specialisation for primary cells in culture. RESULTS: We developed a novel epigenetic clock, the CellPopAge Clock, to accurately monitor the age of a population of adult human primary cells. We find that the CellPopAge Clock can detect decelerated passage-based ageing of human primary cells treated with rapamycin or trametinib, well-established longevity drugs. We then utilise the CellPopAge Clock as a screening tool for the identification of compounds which decelerate ageing of cell populations, uncovering novel anti-ageing drugs, torin2 and dactolisib (BEZ-235). We demonstrate that delayed epigenetic ageing in human primary cells treated with anti-ageing compounds is accompanied by a reduction in senescence and ageing biomarkers. Finally, we extend our screening platform in vivo by taking advantage of a specially formulated holidic medium for increased drug bioavailability in Drosophila. We show that the novel anti-ageing drugs, torin2 and dactolisib (BEZ-235), increase longevity in vivo. CONCLUSIONS: Our method expands the scope of CpG methylation profiling to accurately and rapidly detecting anti-ageing potential of drugs using human cells in vitro, and in vivo, providing a novel accelerated discovery platform to test sought after anti-ageing compounds and geroprotectors

Identifying SARS-CoV-2 antiviral compounds by screening for small molecule inhibitors of nsp13 helicase

The coronavirus disease 2019 (COVID-19) pandemic, which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is a global public health challenge. While the efficacy of vaccines against emerging and future virus variants remains unclear, there is a need for therapeutics. Repurposing existing drugs represents a promising and potentially rapid opportunity to find novel antivirals against SARS-CoV-2. The virus encodes at least nine enzymatic activities that are potential drug targets. Here, we have expressed, purified and developed enzymatic assays for SARS-CoV-2 nsp13 helicase, a viral replication protein that is essential for the coronavirus life cycle. We screened a custom chemical library of over 5000 previously characterized pharmaceuticals for nsp13 inhibitors using a fluorescence resonance energy transfer-based high-throughput screening approach. From this, we have identified FPA-124 and several suramin-related compounds as novel inhibitors of nsp13 helicase activity in vitro. We describe the efficacy of these drugs using assays we developed to monitor SARS-CoV-2 growth in Vero E6 cells

Investigating diagnosis, treatment, and burden of disease in patients with ankylosing spondylitis in Central Eastern Europe and the United States: a real-world study.

INTRODUCTION/OBJECTIVES: Ankylosing spondylitis (AS) is a chronic inflammatory immune-mediated condition. We compared AS diagnosis, treatment, and burden in Central Eastern European countries (CEE), where this has been less researched, and the United States (US) from a real-world perspective. METHODS: Point-in-time survey of rheumatologists and their AS patients was conducted in the US (Apr-Oct 2018) and CEE (Aug-Nov 2019) via physician- and patient-completed record forms, including clinical and patient-reported outcomes. Statistical analysis included descriptive statistics, t-tests, Fisher's exact tests, and generalized linear models. RESULTS: In total, 487 patients were recruited from 88 rheumatologists in the US and 922 patients from 126 rheumatologists in CEE. Time from onset of symptoms to final AS diagnosis was longer in CEE than the US (4.2 vs 2.7 years, p < 0.05). At diagnosis, a greater use of conventional synthetic disease-modifying antirheumatic drugs (DMARDs) and injected steroids was reported in CEE vs the US (43.7% vs 27.6%, p < 0.05; 19.3% vs 8.7%, p < 0.05). 22.9% of US patients received a biologic DMARD at diagnosis vs 10% of CEE patients (p < 0.05). At current consultation, biologic DMARD use in CEE was lower vs the US (27.9% vs 71.0%, p < 0.05). CEE vs US patients had greater disease activity (mean Bath Ankylosing Spondylitis Disease Activity Index 4.2 vs 3.1, p < 0.05) and worse quality of life (QoL; mean Ankylosing Spondylitis Quality of Life Questionnaire score 6.2 vs 8.4, p < 0.05). CONCLUSIONS: AS patients in CEE vs the US faced slower diagnosis and worse access to biologics, disease activity, and QoL. Whether early access to biologics can improve symptoms, QoL, and daily activities in AS patients in CEE remains to be seen. Key Points • The study provided evidence on the real-world approach to the diagnosis, treatment, and burden of axSpA (axial spondyloarthritis) in CEE compared with the US. • The study reported patients in CEE experienced longer delays in diagnosis and poorer access to biologics than in the US. • This may have resulted in higher disease activity, greater levels of pain, and poorer outcomes, as reported by patients with axSpA in CEE

Modulation of adipocyte G-protein expression in cancer cachexia by a lipid-mobilizing factor (LMF)

Adipocytes isolated from cachectic mice bearing the MAC 16 tumour showed over a 3-fold increase in lipolytic response to both low concentrations of isoprenaline and a tumour-derived lipid mobilizing factor (LMF). This was reflected by an enhanced stimulation of adenylate cyclase in plasma membrane fractions of adipocytes in the presence of both factors. There was no up-regulation of adenylate cyclase in response to forskolin, suggesting that the effect arose from a change in receptor number or G-protein expression. Immunoblotting of adipocyte membranes from mice bearing the MAC16 tumour showed an increased expression of Gαs up to 10% weight loss and a reciprocal decrease in Gα. There was also an increased expression of Gαs and a decrease in Gα in adipose tissue from a patient with cancer-associated weight loss compared with a non-cachectic cancer patient. The changes in G-protein expression were also seen in adipose tissue of normal mice administered pure LMF as well as in 3T3L1 adipocytes in vitro. The changes in G-protein expression induced by LMF were attenuated by the polyunsaturated fatty acid, eicosapentaenoic acid (EPA). This suggests that this tumour-derived lipolytic factor acts to sensitize adipose tissue to lipolytic stimuli, and that this effect is attenuated by EPA, which is known to preserve adipose tissue in cancer cachexia. © 2001 Cancer Research Campaig

Using molecular data for epidemiological inference: assessing the prevalence of Trypanosoma brucei rhodesiense in Tsetse in Serengeti, Tanzania

Background: Measuring the prevalence of transmissible Trypanosoma brucei rhodesiense in tsetse populations is essential for understanding transmission dynamics, assessing human disease risk and monitoring spatio-temporal trends and the impact of control interventions. Although an important epidemiological variable, identifying flies which carry transmissible infections is difficult, with challenges including low prevalence, presence of other trypanosome species in the same fly, and concurrent detection of immature non-transmissible infections. Diagnostic tests to measure the prevalence of T. b. rhodesiense in tsetse are applied and interpreted inconsistently, and discrepancies between studies suggest this value is not consistently estimated even to within an order of magnitude. Methodology/Principal Findings: Three approaches were used to estimate the prevalence of transmissible Trypanosoma brucei s.l. and T. b. rhodesiense in Glossina swynnertoni and G. pallidipes in Serengeti National Park, Tanzania: (i) dissection/microscopy; (ii) PCR on infected tsetse midguts; and (iii) inference from a mathematical model. Using dissection/microscopy the prevalence of transmissible T. brucei s.l. was 0% (95% CI 0–0.085) for G. swynnertoni and 0% (0–0.18) G. pallidipes; using PCR the prevalence of transmissible T. b. rhodesiense was 0.010% (0–0.054) and 0.0089% (0–0.059) respectively, and by model inference 0.0064% and 0.00085% respectively. Conclusions/Significance: The zero prevalence result by dissection/microscopy (likely really greater than zero given the results of other approaches) is not unusual by this technique, often ascribed to poor sensitivity. The application of additional techniques confirmed the very low prevalence of T. brucei suggesting the zero prevalence result was attributable to insufficient sample size (despite examination of 6000 tsetse). Given the prohibitively high sample sizes required to obtain meaningful results by dissection/microscopy, PCR-based approaches offer the current best option for assessing trypanosome prevalence in tsetse but inconsistencies in relating PCR results to transmissibility highlight the need for a consensus approach to generate meaningful and comparable data

Examining the Moderating Role of Rational-Versus Emotional-Focused Websites: The Case of Boutique Hotels

This article examines whether emotional or rational advertising appeals are more effective for website hospitality services. Specifically, it considers how attitudes towards websites, service expectations and attitudes towards boutique hotels may influence the two different types of advertising appeals and purchase intention. Results show that websites using emotional appeals tend to show a positive relationship between attitude towards hotels and attitude towards websites and purchase intention. It also indicates that emotional advertisements are better at gaining and retaining interest and attention, and as such can be highly beneficial for small boutique hotels. This article provides several marketing and business policy implications to aid practitioners and strategists in making better decisions

Modified Vaccinia Virus Ankara Preferentially Targets Antigen Presenting Cells In Vitro, Ex Vivo and In Vivo

Modified Vaccinia virus Ankara (MVA) is a promising vaccine vector with an excellent safety profile. However, despite extensive pre-clinical and clinical testing, surprisingly little is known about the cellular tropism of MVA, especially in relevant animal species. Here, we performed in vitro, ex vivo and in vivo experiments with recombinant MVA expressing green fluorescent protein (rMVA-GFP). In both human peripheral blood mononuclear cells and mouse lung explants, rMVA-GFP predominantly infected antigen presenting cells. Subsequent in vivo experiments performed in mice, ferrets and non-human primates indicated that preferential targeting of dendritic cells and alveolar macrophages was observed after respiratory administration, although subtle differences were observed between the respective animal species. Following intramuscular injection, rMVA-GFP was detected in interdigitating cells between myocytes, but also in myocytes themselves. These data are important in advancing our understanding of the basis for the immunogenicity of MVA-based vaccines and aid rational vaccine design and delivery strategies

Chemical characterization of oligosaccharides in the milk of six species of New and Old world monkeys

Human and great ape milks contain a diverse array of milk oligosaccharides, but little is known about the milk oligosaccharides of other primates, and how they differ among taxa. Neutral and acidic oligosaccharides were isolated from the milk of three species of Old World or catarrhine monkeys (Cercopithecidae: rhesus macaque (Macaca mulatta), toque macaque (Macaca sinica) and Hamadryas baboon (Papio hamadryas)) and three of New World or platyrrhine monkeys (Cebidae: tufted capuchin (Cebus apella) and Bolivian squirrel monkey (Saimiri boliviensis); Atelidae: mantled howler (Alouatta palliata)). The milks of these species contained 6–8% total sugar, most of which was lactose: the estimated ratio of oligosaccharides to lactose in Old World monkeys (1:4 to 1:6) was greater than in New World monkeys (1:12 to 1:23). The chemical structures of the oligosaccharides were determined mainly by 1H-NMR spectroscopy. Oligosaccharides containing the type II unit (Gal(β1-4)GlcNAc) were found in the milk of the rhesus macaque, toque macaque, Hamadryas baboon and tufted capuchin, but oligosaccharides containing the type I unit (Gal(β1-3)GlcNAc), which have been found in human and many great ape milks, were absent from the milk of all species studied. Oligosaccharides containing Lewis x (Gal(β1-4)[Fuc(α1-3)]GlcNAc) and 3-fucosyl lactose (3-FL, Gal(β1-4)[Fuc(α1-3)]Glc) were found in the milk of the three cercopithecid monkey species, while 2-fucosyl lactose (5'-FL, Fuc(α1-2)Gal(β1-4)Glc) was absent from all species studied. All of these milks contained acidic oligosaccharides that had N-acetylneuraminic acid as part of their structures, but did not contain oligosaccharides that had N-glycolylneuraminic acid, in contrast to the milk or colostrum of great apes which contain both types of acidic oligosaccharides. Two GalNAc-containing oligosaccharides, lactose 3′-O-sulfate and lacto-N-novopentaose I (Gal(β1-3)[Gal(β1-4)GlcNAc(β1-6)]Gal(β1-4)Glc) were found only in the milk of rhesus macaque, hamadryas baboon and tufted capuchin, respectively. Further research is needed to determine the extent to which the milk oligosaccharide patterns observed among these taxa represent wider phylogenetic trends among primates and how much variation occurs among individuals or species

An Anatomy Massive Open Online Course as a Continuing Professional Development Tool for Healthcare Professionals

Massive open online courses (MOOCs) remain a novel and under-evaluated learning tool within anatomical and medical education. This study aimed to provide valuable information by using an anatomy MOOC to investigate the demographic profile, patterns of engagement and self-perceived benefits to healthcare professionals. A 21-item survey aimed at healthcare professionals was embedded into the Exploring Anatomy: The Human Abdomen MOOC, in April 2016. The course attracted 2711 individual learners with 94 of these completing the survey, and 79 of those confirming they worked full- or part-time as healthcare professionals. Variations in use across healthcare profession (allied healthcare professional, nurse or doctor) were explored using a Fisher’s exact test to calculate significance across demographic, motivation and engagement items; one-way ANOVA was used to compare self-perceived benefits. Survey data revealed that 53.2% were allied healthcare professionals, 35.4% nurses and 11.4% doctors. Across all professions, the main motivation for enrolling was to learn new things in relation to their clinical practice, with a majority following the prescribed course pathway and utilising core, and clinically relevant, material. The main benefits were in relation to improving anatomy knowledge, which enabled better support for patients. This exploratory study assessing engagement and self-perceived benefits of an anatomy MOOC has shown a high level of ordered involvement, with some indicators suggesting possible benefits to patients by enhancing the subject knowledge of those enrolled. It is suggested that this type of learning tool should be further explored as an approach to continuing professional, and interprofessional, education