rAAV9 airway delivery results in effective knockdown of mutant alpha 1-antitrypsin in the liver while upregulating wildtype alpha 1-antitrypsin in the lung

Alpha 1-Antitrypsin (AAT) deficiency is a human genetic disease resulting in the production of mutant AAT, a hepatocyte produced serine protease inhibitor that functions to prevent alveolar epithelial damage by inhibiting neutrophil elastase. Patients with AAT deficiency have increased lung disease, due to decreased proteolytic protection, as well as sporadic severe liver disease secondary to accumulation of mutant AAT, especially a common mutant form termed PiZ, within hepatocytes. We previously showed, in a PiZ mutant mouse model, simultaneous knock-down of mutant PiZ-AAT and augmentation of wild-type AAT production through intravenous delivery of a recombinant adeno-associated viral (rAAV) vector encoding both a miRNA targeting PiZ-AAT and a miRNA-resistant wild-type AAT gene. In this study we tested the hypothesis that rAAV2/9 vector administered intra-nasally or intra-tracheally can deliver a gene of interest to both the airways and liver. Initially C57Bl/6 mice were administered intra-nasally 1011 genome copies (GC) of rAAV2/9 vector expressing a firefly luciferase, which resulted in increased luminescence in the nasal passages, liver, and lung 21 days post delivery. Next, 1012 GC of rAAV2/9 vector expressing GFP and miRNAs targeting PiZ-AAT were delivered via oro-tracheal intubation to PiZ mice. This resulted in decreased serum AAT levels in the PiZ mice and GFP expression in both the liver and lungs. Finally, 1012 GC of rAAV2/9 vector encoding miRNA resistant wild-type AAT and miRNAs targeting PiZ-AAT were delivered via oro-tracheal intubation. This resulted in both systemic and local (liver and lung) elevations in wild-type AAT as well as decreased PiZ-AAT levels. In conclusion, tracheal delivery of rAAV2/9 resulted in expression of AAT in the liver and lung of treated animals, with sufficient targeting of the liver to mediate knock-down of mutant AAT to a similar degree as intravenous delivery, representing a potential non-invasive delivery route for gene therapy in AAT deficient patients

Women do have an improved long-term outcome after non–ST-elevation acute coronary syndromes treated very early and predominantly with percutaneous coronary intervention A prospective study in 1,450 consecutive patients

AbstractObjectivesThis study sought to assess gender-based differences in long-term outcome after very early aggressive revascularization for non–ST-elevation acute coronary syndromes (NSTACS).BackgroundThe Fragmin and fast Revascularization during InStability in Coronary artery disease (FRISC) II study suggested that women have less to gain from an early invasive strategy.MethodsWe conducted a prospective cohort study in 1,450 consecutive patients with NSTACS undergoing coronary angiography and subsequent coronary stenting of the culprit lesion as the primary revascularization strategy within 24 h of admission. The combined primary end point was defined as death or nonfatal myocardial infarction (MI) and recorded for a mean of 20 months.ResultsPercutaneous coronary intervention was performed in more than 50% of patients in women and men and accompanied with stenting in 80%. The percutaneous coronary intervention:coronary artery bypass grafting ratio was 4:1 in men and 5:1 in women. The primary end point occurred in 29 (7.0%) women as compared with 108 (10.5%) men (hazard ratio for women, 0.65; 95% confidence interval [CI] 0.42 to 0.99; p = 0.045). Backward-stepwise multivariate Cox regression analysis identified female gender as an independent predictor of death or MI (hazard ratio for female gender, 0.51; 95% CI, 0.28 to 0.92; p = 0.024). Kaplan-Meier analysis showed that women had consistently lower event rates during the entire follow-up period (p = 0.037 by log-rank for death or MI).ConclusionsWomen treated with very early aggressive revascularization with coronary stenting of the culprit lesion as the primary revascularization strategy have a better long-term outcome as compared with men

Favourable neurological outcome following paediatric out-of-hospital cardiac arrest: a retrospective observational study

BACKGROUND Out-of-hospital cardiac arrest (OHCA) in children is rare and can potentially result in severe neurological impairment. Our study aimed to identify characteristics of and factors associated with favourable neurological outcome following the resuscitation of children by the Swiss helicopter emergency medical service. MATERIALS AND METHODS This retrospective observational study screened the Swiss Air-Ambulance electronic database from 01-01-2011 to 31-12-2021. We included all primary missions for patients ≤ 16 years with OHCA. The primary outcome was favourable neurological outcome after 30 days (cerebral performance categories (CPC) 1 and 2). Multivariable linear regression identified potential factors associated with favourable outcome (odd ratio - OR). RESULTS Having screened 110,331 missions, we identified 296 children with OHCA, which we included in the analysis. Patients were 5.0 [1.0; 12.0] years old and 61.5% (n = 182) male. More than two-thirds had a non-traumatic OHCA (67.2%, n = 199), while 32.8% (n = 97) had a traumatic OHCA. Thirty days after the event, 24.0% (n = 71) of patients were alive, 18.9% (n = 56) with a favourable neurological outcome (CPC 1 n = 46, CPC 2 n = 10). Bystander cardiopulmonary resuscitation (OR 10.34; 95%CI 2.29-51.42; p = 0.002) and non-traumatic aetiology (OR 11.07 2.38-51.42; p = 0.002) were the factors most strongly associated with favourable outcome. Factors associated with an unfavourable neurological outcome were initial asystole (OR 0.12; 95%CI 0.04-0.39; p < 0.001), administration of adrenaline (OR 0.14; 95%CI 0.05-0.39; p < 0.001) and ongoing chest compression at HEMS arrival (OR 0.17; 95%CI 0.04-0.65; p = 0.010). CONCLUSION In this study, 18.9% of paediatric OHCA patients survived with a favourable neurologic outcome 30 days after treatment by the Swiss helicopter emergency medical service. Immediate bystander cardiopulmonary resuscitation and non-traumatic OHCA aetiology were the factors most strongly associated with a favourable neurological outcome. These results underline the importance of effective bystander and first-responder rescue as the foundation for subsequent professional treatment of children in cardiac arrest

STED-FLCS:An Advanced Tool to Reveal Spatiotemporal Heterogeneity of Molecular Membrane Dynamics

Heterogeneous diffusion dynamics of molecules play an important role in many cellular signaling events, such as of lipids in plasma membrane bioactivity. However, these dynamics can often only be visualized by single-molecule and super-resolution optical microscopy techniques. Using fluorescence lifetime correlation spectroscopy (FLCS, an extension of fluorescence correlation spectroscopy, FCS) on a super-resolution stimulated emission depletion (STED) microscope, we here extend previous observations of nanoscale lipid dynamics in the plasma membrane of living mammalian cells. STED-FLCS allows an improved determination of spatiotemporal heterogeneity in molecular diffusion and interaction dynamics via a novel gated detection scheme, as demonstrated by a comparison between STED-FLCS and previous conventional STED-FCS recordings on fluorescent phosphoglycerolipid and sphingolipid analogues in the plasma membrane of live mammalian cells. The STED-FLCS data indicate that biophysical and biochemical parameters such as the affinity for molecular complexes strongly change over space and time within a few seconds. Drug treatment for cholesterol depletion or actin cytoskeleton depolymerization not only results in the already previously observed decreased affinity for molecular interactions but also in a slight reduction of the spatiotemporal heterogeneity. STED-FLCS specifically demonstrates a significant improvement over previous gated STED-FCS experiments and with its improved spatial and temporal resolution is a novel tool for investigating how heterogeneities of the cellular plasma membrane may regulate biofunctionality

Correlation of disability with quality of life in patients with multiple sclerosis treated with natalizumab: primary results and post hoc analysis of the TYSabri ImPROvement study (PROTYS).

BACKGROUND In patients with multiple sclerosis (MS), relapses and disability progression have been associated with decreased health-related quality of life (HRQoL). METHODS PROTYS, a prospective, multicentre, single-arm, observational study in seven Swiss MS centres, evaluated correlations between change in disability status (measured through the Expanded Disability Status Scale (EDSS)) and HRQoL changes (measured through the global Multiple Sclerosis International Quality of Life (MusiQoL) index questionnaire) in 35 patients with relapsing remitting MS on natalizumab for 1 year. In addition, several other scales were also used, such as: Multiple Sclerosis Intimacy and Sexuality Questionnaire-19, EuroQoL-5 Dimension, and Fatigue Scale of Motor and Cognitive Function. A post hoc analysis further assessed the association between HRQoL changes after 1 year and the MusiQoL subscores and other patient-reported outcome (PRO) measures. RESULTS At 1 year, patients were categorised into 'EDSS improved' (6/35), 'EDSS stable' (28/35) and 'EDSS worsened' (1/35). Mean disability scores decreased for 'EDSS improved' and 'EDSS stable' but increased for 'EDSS worsened'. Mean MusiQoL index score for 'EDSS improved' increased from 61.2 at baseline to 66.3 at 1 year, while the 'EDSS stable' group increased from 67.9 to 70.8. No meaningful statistical relationship was observed between EDSS group and changes in MusiQoL score. For the post hoc analysis, patients were categorised in 'MusiQoL improved' (n=21) and 'MusiQoL worsened' (n=14) groups. MusiQoL subscores for 'symptoms,' 'psychological well-being' and 'activities of daily living', as well as scores for several related PRO measures, correlated with improvement of the MusiQoL global index. There was no correlation between the changes in MusiQoL global index and EDSS score. CONCLUSIONS Natalizumab treatment for 1 year resulted in either improved or stable EDSS status in most patients, and although no significant relationship was observed between global HRQoL change and EDSS change, several domains of HRQoL seemed to improve with natalizumab treatment. TRIAL REGISTRATION NUMBER NCT02386566

Targeted gene delivery to the enteric nervous system using AAV: a comparison across serotypes and capsid mutants

Recombinant adeno-associated virus (AAV) vectors are one of the most widely used gene transfer systems in research and clinical trials. AAV can transduce a wide range of biological tissues, however to date, there has been no investigation on targeted AAV transduction of the enteric nervous system (ENS). Here, we examined the efficiency, tropism, spread, and immunogenicity of AAV transduction in the ENS. Rats received direct injections of various AAV serotypes expressing green fluorescent protein (GFP) into the descending colon. AAV serotypes tested included; AAV 1, 2, 5, 6, 8, or 9 and the AAV2 and AAV8 capsid mutants, AAV2-Y444F, AAV2-tripleY-F, AAV2-tripleY-F+T-V, AAV8-Y733F, and AAV8-doubeY-F+T-V. Transduction, as determined by GFP-positive cells, occurred in neurons and enteric glia within the myenteric and submucosal plexuses of the ENS. AAV6 and AAV9 showed the highest levels of transduction within the ENS. Transduction efficiency scaled with titer and time, was translated to the murine ENS, and produced no vector-related immune response. A single injection of AAV into the colon covered an area of ~47 mm(2). AAV9 primarily transduced neurons, while AAV6 transduced enteric glia and neurons. This is the first report on targeted AAV transduction of neurons and glia in the ENS

Indications and practical approach to non-invasive ventilation in acute heart failure

In acute heart failure (AHF) syndromes significant respiratory failure (RF) is essentially seen in patients with acute cardiogenic pulmonary oedema (ACPE) or cardiogenic shock (CS). Non-invasive ventilation (NIV), the application of positive intrathoracic pressure through an interface, has shown to be useful in the treatment of moderate to severe RF in several scenarios. There are two main modalities of NIV: continuous positive airway pressure (CPAP) and pressure support ventilation (NIPSV) with positive end expiratory pressure. Appropriate equipment and experience is needed for NIPSV, whereas CPAP may be administered without a ventilator, not requiring special training. Both modalities have shown to be effective in ACPE, by a reduction of respiratory distress and the endotracheal intubation rate compared to conventional oxygen therapy, but the impact on mortality is less conclusive. Non-invasive ventilation is also indicated in patients with AHF associated to pulmonary disease and may be considered, after haemodynamic stabilization, in some patients with CS. There are no differences in the outcomes in the studies comparing both techniques, but CPAP is a simpler technique that may be preferred in low-equipped areas like the pre-hospital setting, while NIPSV may be preferable in patients with significant hypercapnia. The new modality 'high-flow nasal cannula' seems promising in cases of AHF with less severe RF. The correct selection of patients and interfaces, early application of the technique, the achievement of a good synchrony between patients and the ventilator avoiding excessive leakage, close monitoring, proactive management, and in some cases mild sedation, may warrant the success of the technique

Therapeutic rAAVrh10 Mediated SOD1 Silencing in Adult SOD1(G93A) Mice and Nonhuman Primates

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease; survival in ALS is typically 3-5 years. No treatment extends patient survival by more than three months. Approximately 20% of familial ALS and 1-3% of sporadic ALS patients carry a mutation in the gene encoding superoxide dismutase 1 (SOD1). In a transgenic ALS mouse model expressing the mutant SOD1(G93A) protein, silencing the SOD1 gene prolongs survival. One study reports a therapeutic effect of silencing the SOD1 gene in systemically treated adult ALS mice; this was achieved with a short hairpin RNA, a silencing molecule that has raised multiple safety concerns, and recombinant adeno-associated virus (rAAV) 9. We report here a silencing method based on an artificial microRNA termed miR-SOD1 systemically delivered using adeno-associated virus rAAVrh10, a serotype with a demonstrated safety profile in CNS clinical trials. Silencing of SOD1 in adult SOD1(G93A) transgenic mice with this construct profoundly delayed both disease onset and death in the SOD1(G93A) mice, and significantly preserved muscle strength and motor and respiratory functions. We also document that intrathecal delivery of the same rAAVrh10-miR-SOD1 in nonhuman primates significantly and safely silences SOD1 in lower motor neurons. This study supports the view that rAAVrh10-miR-SOD1 merits further development for the treatment of SOD1-linked ALS in humans