Glucosylceramide synthase inhibition alleviates aberrations in synucleinopathy models.

Mutations in the glucocerebrosidase gene (GBA) confer a heightened risk of developing Parkinson's disease (PD) and other synucleinopathies, resulting in a lower age of onset and exacerbating disease progression. However, the precise mechanisms by which mutations in GBA increase PD risk and accelerate its progression remain unclear. Here, we investigated the merits of glucosylceramide synthase (GCS) inhibition as a potential treatment for synucleinopathies. Two murine models of synucleinopathy (a Gaucher-related synucleinopathy model, GbaD409V/D409V and a A53T-α-synuclein overexpressing model harboring wild-type alleles of GBA, A53T-SNCA mouse model) were exposed to a brain-penetrant GCS inhibitor, GZ667161. Treatment of GbaD409V/D409V mice with the GCS inhibitor reduced levels of glucosylceramide and glucosylsphingosine in the central nervous system (CNS), demonstrating target engagement. Remarkably, treatment with GZ667161 slowed the accumulation of hippocampal aggregates of α-synuclein, ubiquitin, and tau, and improved the associated memory deficits. Similarly, prolonged treatment of A53T-SNCA mice with GZ667161 reduced membrane-associated α-synuclein in the CNS and ameliorated cognitive deficits. The data support the contention that prolonged antagonism of GCS in the CNS can affect α-synuclein processing and improve behavioral outcomes. Hence, inhibition of GCS represents a disease-modifying therapeutic strategy for GBA-related synucleinopathies and conceivably for certain forms of sporadic disease.

The Staphylococcus aureus superantigen SElX is a bifunctional toxin that inhibits neutrophil function.

Bacterial superantigens (SAgs) cause Vß-dependent T-cell proliferation leading to immune dysregulation associated with the pathogenesis of life-threatening infections such as toxic shock syndrome, and necrotizing pneumonia. Previously, we demonstrated that staphylococcal enterotoxin-like toxin X (SElX) from Staphylococcus aureus is a classical superantigen that exhibits T-cell activation in a Vß-specific manner, and contributes to the pathogenesis of necrotizing pneumonia. Here, we discovered that SElX can also bind to neutrophils from human and other mammalian species and disrupt IgG-mediated phagocytosis. Site-directed mutagenesis of the conserved sialic acid-binding motif of SElX abolished neutrophil binding and phagocytic killing, and revealed multiple glycosylated neutrophil receptors for SElX binding. Furthermore, the neutrophil binding-deficient mutant of SElX retained its capacity for T-cell activation demonstrating that SElX exhibits mechanistically independent activities on distinct cell populations associated with acquired and innate immunity, respectively. Finally, we demonstrated that the neutrophil-binding activity rather than superantigenicity is responsible for the SElX-dependent virulence observed in a necrotizing pneumonia rabbit model of infection. Taken together, we report the first example of a SAg, that can manipulate both the innate and adaptive arms of the human immune system during S. aureus pathogenesis.

Staphylococcus pseudintermedius Surface Protein L (SpsL) Is Required for Abscess Formation in a Murine Model of Cutaneous Infection.

Staphylococcus pseudintermedius is the leading cause of pyoderma in dogs and is often associated with recurrent skin infections that require prolonged antibiotic therapy. High levels of antibiotic use have led to multidrug resistance, including the emergence of epidemic methicillin-resistant clones. Our understanding of the pathogenesis of S. pseudintermedius skin infection is very limited, and the identification of the key host-pathogen interactions underpinning infection could lead to the design of novel therapeutic or vaccine-based approaches for controlling disease. Here, we employ a novel murine cutaneous-infection model of S. pseudintermedius and investigate the role of the two cell wall-associated proteins (SpsD and SpsL) in skin disease pathogenesis. Experimental infection with wild-type S. pseudintermedius strain ED99 or a gene-deletion derivative deficient in expression of SpsD led to a focal accumulation of neutrophils and necrotic debris in the dermis and deeper tissues of the skin characteristic of a classical cutaneous abscess. In contrast, mice infected with mutants deficient in SpsL or both SpsD and SpsL developed larger cutaneous lesions with distinct histopathological features of regionally extensive cellulitis rather than focal abscessation. Furthermore, comparison of the bacterial loads in S. pseudintermedius-induced cutaneous lesions revealed a significantly increased burden of bacteria in the mice infected with SpsL-deficient mutants. These findings reveal a key role for SpsL in murine skin abscess formation and highlight a novel function for a bacterial surface protein in determining the clinical outcome and pathology of infection caused by a major canine pathogen.

Primary cervico-uterine anastomosis in a patient with agenesis of the uterine isthmus: A case report and review.

Arrested caudal extension of the fused Müllerian ducts, either complete or segmental, causes obstruction of the outflow tract. We describe a case of agenesis of the uterine isthmus and the procedure to establish the continuity of the outflow tract. A 15-year-old girl with cyclic pelvic pain and amenorrhea was found to have a 3-cm gap between the uterine body and the cervix. After cervico-uterine anastomosis via laparotomy she menstruated normally and was pain-free. Selected cases of arrested Müllerian duct extension can be corrected by primary anastomosis, which establishes anatomical continuity and restores menstruation and, possibly, future fertility.

Fibronectin Binding Proteins SpsD and SpsL Both Support Invasion of Canine Epithelial Cells by Staphylococcus pseudintermedius.

In this study, we investigated the cell wall-anchored fibronectin-binding proteins SpsD and SpsL from the canine commensal and pathogen Staphylococcus pseudintermedius for their role in promoting bacterial invasion of canine progenitor epidermal keratinocytes (CPEK). Invasion was examined by the gentamicin protection assay and fluorescence microscopy. An ΔspsD ΔspsL mutant of strain ED99 had a dramatically reduced capacity to invade CPEK monolayers, while no difference in the invasion level was observed with single mutants. Lactococcus lactis transformed with plasmids expressing SpsD and SpsL promoted invasion, showing that both proteins are important. Soluble fibronectin was required for invasion, and an RGD-containing peptide or antibodies recognizing the integrin α5ß1 markedly reduced invasion, suggesting an important role for the integrin in this process. Src kinase inhibitors effectively blocked internalization, suggesting a functional role for the kinase in invasion. In order to identify the minimal fibronectin-binding region of SpsD and SpsL involved in the internalization process, recombinant fragments of both proteins were produced. The SpsD520-846 and SpsL538-823 regions harboring the major fibronectin-binding sites inhibited S. pseudintermedius internalization. Finally, the effects of staphylococcal invasion on the integrity of different cell lines were examined. Because SpsD and SpsL are critical factors for adhesion and invasion, blocking these processes could provide a strategy for future approaches to treating infections.

Evaluation of a student-led interprofessional innovative health promotion model for an underserved population with diabetes: a pilot project.

Interprofessional education seeks to encourage different health professions to interact and learn together during their training process which will eventually lead to collaborative healthcare practices and improved care for patients. This study determined whether student understanding of diabetes management and the role of health professionals in diabetes care improved after the implementation of an interprofessional health promotion program. Sixty-three students from five health professions led six educational sessions concentrating on critical components of diabetes management. The longitudinal program covered topics within the Alphabet Strategy (A-G). Students were surveyed to determine their understanding of diabetes management. Data were gathered at the beginning of the study and its conclusion. Forty-seven students completed the program and the pre- and post-survey. There were significant improvements in students' knowledge of diabetes care, understanding of the roles of healthcare professionals and ability to work with other healthcare professionals. Nineteen patients completed the study. There were no significant differences in patients' diabetes knowledge, understanding of diabetes care and clinical outcomes. This study acknowledged the potential value of an interprofessional team approach to care. This innovative model could be applied to other practice settings and used for the management of other chronic diseases.

Epidemiology of compassion: A literature review.

Psychology and neuroscience have contributed significantly to advances in understanding compassion. In contrast, little attention has been given to the epidemiology of compassion. The human experience of compassion is heterogeneous with respect to time, place, and person. Therefore, compassion has an epidemiology, although little is known about the factors that account for spatial or temporal clustering of compassion or how these factors might be harnessed to promote and realize a more compassionate world. We reviewed the scientific literature to describe what is known about "risk factors" for compassion towards others. Studies were included if they used quantitative methods, treated compassion as an outcome, and used measures of compassion that included elements of empathy and action to alleviate suffering. Eighty-two studies met the inclusion criteria; 89 potential risk factors were tested 418 times for association with compassion. Significant associations with compassion were found for individual demographic factors (e.g., gender, religious faith); personal characteristics (e.g., emotional intelligence, perspective-taking, secure attachment); personal experience (e.g., previous adversity); behaviors (e.g., church attendance); circumstantial factors during the compassion encounter (e.g., perceptions of suffering severity, relational proximity of the compassion-giver and -receiver, emotional state of the compassion-giver); and organizational features. Few studies explored the capacity to receive, rather than give, compassion. Definitions and measures of compassion varied widely across disciplines; 87% of studies used self-report measures and 39% used a cross-sectional design. Ten randomized clinical trials documented the effectiveness of compassion training. From an epidemiologic perspective, most studies treated compassion as an individual host factor rather than as transmissible or influenced by time or the environment. The causal pathways leading from suffering to a compassionate response appear to be non-linear and complex. A variety of factors (acting as effect modifiers) appear to be permissive of-or essential for-the arising of compassion in certain settings or specific populations. Future epidemiologic research on compassion should take into account contextual and environmental factors and should elucidate compassion-related dynamics within organizations and human systems. Such research should be informed by a range of epidemiologic tools and methods, as well as insights from other scientific disciplines and spiritual and religious traditions.

Preclinical pharmacology of glucosylceramide synthase inhibitor venglustat in a GBA-related synucleinopathy model.

Mutations in GBA, the gene encoding the lysosomal enzyme glucocerebrosidase (GCase), represent the greatest genetic risk factor for developing synucleinopathies including Parkinson's disease (PD). Additionally, PD patients harboring a mutant GBA allele present with an earlier disease onset and an accelerated disease progression of both motor and non-motor symptoms. Preclinical studies in mouse models of synucleinopathy suggest that modulation of the sphingolipid metabolism pathway via inhibition of glucosylceramide synthase (GCS) using a CNS-penetrant small molecule may be a potential treatment for synucleinopathies. Here, we aim to alleviate the lipid storage burden by inhibiting the de novo synthesis of the primary glycosphingolipid substrate of GCase, glucosylceramide (GlcCer). We have previously shown that systemic GCS inhibition reduced GlcCer and glucosylsphingosine (GlcSph) accumulation, slowed α-synuclein buildup in the hippocampus, and improved cognitive deficits. Here, we studied the efficacy of a brain-penetrant clinical candidate GCS inhibitor, venglustat, in mouse models of GBA-related synucleinopathy, including a heterozygous Gba mouse model which more closely replicates the typical GBA-PD patient genotype. Collectively, these data support the rationale for modulation of GCase-related sphingolipid metabolism as a therapeutic strategy for treating GBA-related synucleinopathies.

The BYPASS-CTCA Study: the value of Computed Tomography Cardiac Angiography (CTCA) in improving patient-related outcomes in patients with previous bypass operation undergoing invasive coronary angiography: Study Protocol of a Randomised Controlled Trial.

BACKGROUND: Patients with ischaemic heart disease and previous coronary artery bypass grafting (CABG) often need coronary evaluation by means of invasive coronary angiography (ICA). ICA in such patients is technically more challenging and carries a higher risk of complications including kidney damage, myocardial infarction, stroke and death. Improvements in Computed Tomography Cardiac Angiography (CTCA) technology have ensured its emergence as a useful clinical tool in CABG assessment, allowing for its potential use in planning interventional procedures in this patient group. METHODS: The BYPASS-CTCA study is a prospective, single centre, randomised controlled trial assessing the value of upfront CTCA in patients with previous surgical revascularisation undergoing ICA procedures. A total of 688 patients with previous CABG, requiring ICA for standard indications, will be recruited and randomised to receive ICA alone, or CTCA prior to angiography. Subjects will be followed up over a 12-month period post procedure. The primary endpoints are ICA procedural duration, incidence of contrast-induced nephropathy (CIN) and patient satisfaction scores post ICA. Secondary endpoints include contrast dose (mL) and radiation dose (mSv) during ICA, number of catheters used, angiography-related complications and cost-effectiveness of CTCA (QALY) over 12 months. DISCUSSION: The study will investigate the hypothesis that CTCA prior to ICA in patients with previous CABG can reduce procedural duration, post-procedural kidney damage and improve patient satisfaction, therefore strengthening its role in this group of patients. TRIAL REGISTRATION: The study is registered on ClinicalTrials.gov which is a resource maintained by the U.S. National Library of Medicine. Registration number NCT03736018.

Evaluating hours of sleep and perceived stress on dietary cognitive restraint in a survey of college students.

OBJECTIVE: The objective of this study was to examine associations of sleep quality and quantity, food security, and physical activity with eating behaviors that may be associated with college weight gain. Participants: College students enrolled in multiple sections of a general education class completed an online survey in January 2016 (n = 153; 18-52 years of age). Methods: A cross-sectional study was conducted. Outcome variables included emotional eating (EE), uncontrolled eating (UE), and cognitive restraint (CR) as measured by The Three Factor Eating Questionnaire Revised. Bivariate analyses, ANOVA, and multiple linear regression were completed with significance at p≤.05. Results: Higher EE was associated with higher stress levels and female sex (p <.001 and p=.02) and higher UE scores were associated with higher perceived stress (p<.001) while lower UE scores were associated with tobacco use (p=.03 regression, p=.098, bivariate). Higher CR was associated with higher parental education and use of relaxation methods. Higher CR also was associated with perceived stress, but this relationship differed depending upon freshmen status and amount of physical activity, and a relationship with sleep was observed that differed depending upon freshmen status. Conclusion: Interventions to help college students reduce stress and improve sleep may improve eating behaviors.

Sodium Nitrite-Mediated Cardioprotection in Primary Percutaneous Coronary Intervention for ST-Segment Elevation Myocardial Infarction: A Cost-Effectiveness Analysis.

OBJECTIVES: In the follow-up of patients in a trial of intracoronary sodium nitrite given during primary percutaneous coronary intervention (PCI) after acute myocardial infarction (AMI), we found a reduction in the incidence of major adverse cardiac events (MACEs). Specifically, MACE rates were 5.2% versus 25.0% with placebo at 3 years ( P = .013). Such MACE reductions should also be associated with economic benefit. Thus, we assessed the cost utility of sodium nitrite therapy versus standard primary PCI only. METHODS AND RESULTS: We developed a model to simulate costs and quality-adjusted life years (QALYs) over the first 36 months after ST-Segment Elevation Myocardial Infarction (STEMI). Decision tree analysis was used to assess different potential cardiovascular outcomes after STEMI for patients in both treatment groups. Model inputs were derived from the NITRITE-AMI study. Cost of comparative treatments and follow-up in relation to cardiovascular events was calculated from the United Kingdom National Health Service perspective. Higher procedural costs for nitrite treatment were offset by lower costs for repeat revascularization, myocardial infarction, and hospitalization for heart failure compared to primary PCI plus placebo. Nitrite treatment was associated with higher utility values (0.91 ± 0.19 vs 0.82 ± 0.30, P = .041). The calculated incremental cost-effectiveness ratio of £2177 per QALY indicates a cost-effective strategy. Furthermore, positive results were maintained when input parameters varied, indicating the robustness of our model. In fact, based on the difference in utility values, the cost of nitrite could increase by 4-fold (£2006 per vial) and remain cost-effective. CONCLUSION: This first analysis of sodium nitrite as a cardioprotective treatment demonstrates cost-effectiveness. Although more comparative analysis and assessment of longer follow-up times are required, our data indicate the considerable potential of nitrite-mediated cardioprotection.

Motivating 18- to 24-year-olds to increase their fruit and vegetable consumption.

OBJECTIVE: This study assessed the effectiveness of a 4-month intervention using stage-based newsletters, computer-based communication, and motivational interviewing to increase fruit and vegetable consumption by college students aged 18 to 24 years. DESIGN: Participants were stratified by stage of change for fruit and vegetable consumption and randomized to an intervention or control group. Participants completed the staging algorithm for fruit and vegetable intake, which included a one-item food frequency question, a 26-item food frequency questionnaire (FFQ), an 18-item decisional balance questionnaire, and a five-item self-efficacy questionnaire at baseline and completion of study. SUBJECTS: A convenience sample of 437 college students enrolled in a rural, land grant university was enrolled in the study. Only nondietetics majors between ages 18 to 24 years were included in the study. A total of 314 students finished the study for a completion rate of 72%. INTERVENTION: After baseline staging and randomization, the intervention group participants received four stage-based newsletters, one motivational interview, and an individually tailored e-mail follow-up over a 4-month period. Control group participants only received assessment at baseline and at completion. MAIN OUTCOME MEASURES: Two fruit and vegetable instruments, a one-item food frequency question, and a 26-item FFQ measured daily consumption of fruits and vegetables at baseline and postintervention. STATISTICAL ANALYSES PERFORMED: The SAS system for Windows, version 8 (1999, SAS Institute, Inc, Cary, NC), was used for analysis, including the following tests: PROC GLM, PROC FREQ, and PROC NPAR1WAY, Kruskal-Wallis, Fisher, Wilcoxon rank sum, and chi(2). RESULTS: Fruit and vegetable consumption increased significantly more for the intervention group than the control group. Consumption increased in the intervention group by one serving a day for both instruments compared with 0.4 servings a day in the control group for a one-item instrument and no change in the control group for a 26-item FFQ. CONCLUSIONS: This intervention is an effective way to increase fruit and vegetable consumption by young adults.

Recombination-mediated remodelling of host-pathogen interactions during Staphylococcus aureus niche adaptation.

Large-scale recombination events have led to the emergence of epidemic clones of several major bacterial pathogens. However, the functional impact of the recombination on clonal success is not understood. Here, we identified a novel widespread hybrid clone (ST71) of livestock-associated Staphylococcus aureus that evolved from an ancestor belonging to the major bovine lineage CC97, through multiple large-scale recombination events with other S. aureus lineages occupying the same ruminant niche. The recombination events, affecting a 329 kb region of the chromosome spanning the origin of replication, resulted in allele replacement and loss or gain of an array of genes influencing host-pathogen interactions. Of note, molecular functional analyses revealed that the ST71 hybrid clone has acquired multiple novel pathogenic traits associated with acquired and innate immune evasion and bovine extracellular matrix adherence. These findings provide a paradigm for the impact of large-scale recombination events on the rapid evolution of bacterial pathogens within defined ecological niches.

Silencing mutant huntingtin by adeno-associated virus-mediated RNA interference ameliorates disease manifestations in the YAC128 mouse model of Huntington's disease.

Huntington's disease (HD) is a fatal autosomal dominant neurodegenerative disease caused by an increase in the number of polyglutamine residues in the huntingtin (Htt) protein. With the identification of the underlying basis of HD, therapies are being developed that reduce expression of the causative mutant Htt. RNA interference (RNAi) that seeks to selectively reduce the expression of such disease-causing agents is emerging as a potential therapeutic strategy for this and similar disorders. This study examines the merits of administering a recombinant adeno-associated viral (AAV) vector designed to deliver small interfering RNA (siRNA) that targets the degradation of the Htt transcript. The aim was to lower Htt levels and to correct the behavioral, biochemical, and neuropathological deficits shown to be associated with the YAC128 mouse model of HD. Our data demonstrate that AAV-mediated RNAi is effective at transducing greater than 80% of the cells in the striatum and partially reducing the levels (~40%) of both wild-type and mutant Htt in this region. Concomitant with these reductions are significant improvements in behavioral deficits, reduction of striatal Htt aggregates, and partial correction of the aberrant striatal transcriptional profile observed in YAC128 mice. Importantly, a partial reduction of both the mutant and wild-type Htt levels is not associated with any notable overt neurotoxicity. Collectively, these results support the continued development of AAV-mediated RNAi as a therapeutic strategy for HD.

Translational fidelity of intrathecal delivery of self-complementary AAV9-survival motor neuron 1 for spinal muscular atrophy.

Spinal muscular atrophy (SMA) is a neuromuscular disease caused by mutations in survival motor neuron 1 (SMN1). Previously, we showed that central nervous system (CNS) delivery of an adeno-associated viral (AAV) vector encoding SMN1 produced significant improvements in survival in a mouse model of SMA. Here, we performed a dose-response study in SMA mice to determine the levels of SMN in the spinal cord necessary for efficacy, and measured the efficiency of motor neuron transduction in the spinal cord after intrathecal delivery in pigs and nonhuman primates (NHPs). CNS injections of 5e10, 1e10, and 1e9 genome copies (gc) of self-complementary AAV9 (scAAV9)-hSMN1 into SMA mice extended their survival from 17 to 153, 70, and 18 days, respectively. Spinal cords treated with 5e10, 1e10, and 1e9 gc showed that 70-170%, 30-100%, and 10-20% of wild-type levels of SMN were attained, respectively. Furthermore, detectable SMN expression in a minimum of 30% motor neurons correlated with efficacy. A comprehensive analysis showed that intrathecal delivery of 2.5e13 gc of scAAV9-GFP transduced 25-75% of the spinal cord motor neurons in NHPs. Thus, the extent of gene expression in motor neurons necessary to confer efficacy in SMA mice could be obtained in large-animal models, justifying the continual development of gene therapy for SMA.

Antisense oligonucleotides delivered to the mouse CNS ameliorate symptoms of severe spinal muscular atrophy.

Spinal muscular atrophy (SMA) is an autosomal recessive neuromuscular disorder caused by mutations in the SMN1 gene that result in a deficiency of SMN protein. One approach to treat SMA is to use antisense oligonucleotides (ASOs) to redirect the splicing of a paralogous gene, SMN2, to boost production of functional SMN. Injection of a 2'-O-2-methoxyethyl-modified ASO (ASO-10-27) into the cerebral lateral ventricles of mice with a severe form of SMA resulted in splice-mediated increases in SMN protein and in the number of motor neurons in the spinal cord, which led to improvements in muscle physiology, motor function and survival. Intrathecal infusion of ASO-10-27 into cynomolgus monkeys delivered putative therapeutic levels of the oligonucleotide to all regions of the spinal cord. These data demonstrate that central nervous system-directed ASO therapy is efficacious and that intrathecal infusion may represent a practical route for delivering this therapeutic in the clinic.

CNS-targeted gene therapy improves survival and motor function in a mouse model of spinal muscular atrophy.

Spinal muscular atrophy (SMA) is a neuromuscular disease caused by a deficiency of survival motor neuron (SMN) due to mutations in the SMN1 gene. In this study, an adeno-associated virus (AAV) vector expressing human SMN (AAV8-hSMN) was injected at birth into the CNS of mice modeling SMA. Western blot analysis showed that these injections resulted in widespread expression of SMN throughout the spinal cord, and this translated into robust improvement in skeletal muscle physiology, including increased myofiber size and improved neuromuscular junction architecture. Treated mice also displayed substantial improvements on behavioral tests of muscle strength, coordination, and locomotion, indicating that the neuromuscular junction was functional. Treatment with AAV8-hSMN increased the median life span of mice with SMA-like disease to 50 days compared with 15 days for untreated controls. Moreover, injecting mice with SMA-like disease with a human SMN-expressing self-complementary AAV vector - a vector that leads to earlier onset of gene expression compared with standard AAV vectors - led to improved efficacy of gene therapy, including a substantial extension in median survival to 157 days. These data indicate that CNS-directed, AAV-mediated SMN augmentation is highly efficacious in addressing both neuronal and muscular pathologies in a severe mouse model of SMA.

Long-term bone marrow culture and its clinical potential in chronic wound healing.

Bone marrow-derived cells have long been regarded to play a crucial role in the homeostasis of skin. We have previously described the clinical benefit of directly applying autologous bone marrow aspirate and cultured bone marrow cells to recalcitrant chronic skin wounds. The initial response to treatment appears to be vascular in nature with the formation of new blood vessels. The difficulty in consistently growing adequate numbers of cells for delivery to patients was, however, a limiting factor. Here, in a subsequent protocol, we describe an improved bone marrow culture system yielding a reliable growth of bone marrow cells and leading to a greater clinical response. Cells expressing markers of endothelial progenitors including CD133, CD146, and particularly CD14 are enhanced in these cultures. CD14-isolated cells produced colonies in endothelial cell assays and sprouting in matrigel assays. Angiogenic cytokines, including angiogenin, epithelial neutrophil-activating protein-78, growth-regulated oncogene, growth-regulated oncogene-alpha, Interleukin-8, CXC16, and monocyte chemoattractant protein-1, were found to be elevated in these cultures. Administration of improved culture cells to patients with chronic wounds present for >1 year lead to an enhanced clinical response.