Phosphomimetic S3D cofilin binds but only weakly severs actin filaments.

Many biological processes, including cell division, growth, and motility, rely on rapid remodeling of the actin cytoskeleton and on actin filament severing by the regulatory protein cofilin. Phosphorylation of vertebrate cofilin at Ser-3 regulates both actin binding and severing. Substitution of serine with aspartate at position 3 (S3D) is widely used to mimic cofilin phosphorylation in cells and in vitro The S3D substitution weakens cofilin binding to filaments, and it is presumed that subsequent reduction in cofilin occupancy inhibits filament severing, but this hypothesis has remained untested. Here, using time-resolved phosphorescence anisotropy, electron cryomicroscopy, and all-atom molecular dynamics simulations, we show that S3D cofilin indeed binds filaments with lower affinity, but also with a higher cooperativity than wild-type cofilin, and severs actin weakly across a broad range of occupancies. We found that three factors contribute to the severing deficiency of S3D cofilin. First, the high cooperativity of S3D cofilin generates fewer boundaries between bare and decorated actin segments where severing occurs preferentially. Second, S3D cofilin only weakly alters filament bending and twisting dynamics and therefore does not introduce the mechanical discontinuities required for efficient filament severing at boundaries. Third, Ser-3 modification (i.e. substitution with Asp or phosphorylation) "undocks" and repositions the cofilin N terminus away from the filament axis, which compromises S3D cofilin's ability to weaken longitudinal filament subunit interactions. Collectively, our results demonstrate that, in addition to inhibiting actin binding, Ser-3 modification favors formation of a cofilin-binding mode that is unable to sufficiently alter filament mechanical properties and promote severing.

Cardiac amyloidosis and left atrial appendage closure. The CAMYLAAC study.

INTRODUCTION AND OBJECTIVES: Transthyretin cardiac amyloidosis (ATTR-CA) patients often have atrial fibrillation and increased bleeding/thrombogenic risks. We aimed to evaluate outcomes of left atrial appendage closure (LAAC) compared with patients without a known diagnosis of CA. METHODS: Comparison at long-term of patients diagnosed with ATTR-CA who underwent LAAC between 2009 and 2020 and those without a known diagnosis of CA. RESULTS: We studied a total of 1159 patients. Forty patients (3.5%) were diagnosed with ATTR-CA; these patients were older and had more comorbidities, higher HAS-BLED and CHA2DS2-VASc scores, and lower left ventricular function. Successful LAAC was achieved in 1137 patients (98.1%) with no differences between groups. Regarding in-hospital and follow-up complications, there were no differences between the groups in ischemic stroke (5% vs 2.5% in those without a known diagnosis of CA; P=.283), hemorrhagic stroke (2.5% and 0.8% in the control group; P=.284), major or minor bleeding. At the 2-year follow-up, there were no significant differences in mortality (ATTR-CA: 20% vs those without known CA: 13.6%, 0.248); however, the at 5-year follow-up, ATTR-CA patients had higher mortality (40% vs 19.2%; P <.001) but this difference was unrelated to hemorrhagic complications or ischemic stroke. CONCLUSIONS: LAAC could reduce the risk of bleeding complications and ischemic cerebrovascular events without increasing the rate of early or mid-term complications. Although long-term survival was impaired in ATTR-CA patients, it was comparable to that of patients without a known diagnosis of CA at the 2-year follow-up, suggesting that LAAC for patients with ATTR-CA might not be futile.

Effectiveness and Safety of an Outpatient Program for Percutaneous Left Atrial Appendage Occlusion.

BACKGROUND: Left atrial appendage occlusion (LAAO) is a safe and effective alternative to oral anticoagulation for thromboprophylaxis in patients with nonvalvular atrial fibrillation. Technological development in devices and imaging techniques, as well as accumulated experience, have increased procedural success rates and decreased complications. Same-day discharge protocols have been proposed in the field of structural heart disease, but this approach has not been studied in detail for the LAAO procedure. AIM: The aim of this study is to assess the safety and efficacy of an outpatient program for LAAO when compared to the conventional treatment approach. METHODS: We present a retrospective, non-randomized single-center study of 262 consecutive patients undergoing LAAO. Patients were divided into two groups, the first (n = 131) followed a conventional protocol (CP), and the second (n = 131) an outpatient protocol (OP). The primary composite endpoint comprised MACCE (death, stroke, and bleeding), cardiac tamponade, vascular complication, or attendance in the emergency department after hospital discharge at 30 days. RESULTS: The overall success rate was 99.6%, with a periprocedural complication rate of 2.29%. With regards to the CP versus OP group, there were no differences between incidences of the primary composite endpoint (6.1% PC vs. 3.0% PA, p = 0.24), or after an analysis, with propensity score matching. No differences were observed in the individual endpoints. There was a decrease in hospital length of stay in the same-day discharge group (p < 0.01). CONCLUSIONS: A same-day discharge LAAO program is safe, effective, and feasible when compared to the conventional strategy. Moreover, it reduces hospital length of stay, which might have clinical and economic benefits.

Left Atrial Appendage Occlusion in Hereditary Haemorrhagic Telangiectasia Patients (Rendu Osler Syndrome) With Non-Valvular Atrial Fibrillation: Prevention of Cardioembolic Events While Avoiding the Long-Term Risks of Oral Anticoagulation.

BACKGROUND/PURPOSE: Hereditary haemorrhagic telangiectasia (HHT) (Osler-Weber-Rendu Syndrome) is an inherited vascular disorder, associated with arteriovenous malformations and bleeding events. Despite potential benefits, data regarding the use of left atrial appendage occlusion (LAAO) in HHT patients with non-valvular atrial fibrillation (NVAF) is scarce. The aim of the present paper was to assess the outcomes of HHT patients undergoing LAAO. METHODS/MATERIALS: This multicentre observational study included all consecutive patients with NVAF and a previous diagnosis of HHT who underwent LAAO between 2015 and December 2020. The follow-up protocol included a clinical control at 3 months after hospital discharge, and then yearly. RESULTS: Overall, 822 subjects undergoing LAAO were initially screened for the study. Among them, 7 patients had previous diagnosis of HHT. Device implant success was achieved in all patients and non-procedural adverse event were observed. At the maximum follow-up (median follow up = 470.5 days), no ischemic strokes or device related thrombosis (DRT) were reported. Absence of antithrombotic treatment after LAAO was not associated with an increased risk of stroke or DRT. CONCLUSIONS: LAAO might represent a promising non-pharmacological alternative to prevent thromboembolic events in patients with HHT and NVAF.

Outcome Analysis in Elective Electrical Cardioversion of Atrial Fibrillation Patients: Development and Validation of a Machine Learning Prognostic Model.

Background: The integrated approach to electrical cardioversion (EC) in atrial fibrillation (AF) is complex; candidates can resolve spontaneously while waiting for EC, and post-cardioversion recurrence is high. Thus, it is especially interesting to avoid the programming of EC in patients who would restore sinus rhythm (SR) spontaneously or present early recurrence. We have analyzed the whole elective EC of the AF process using machine-learning (ML) in order to enable a more realistic and detailed simulation of the patient flow for decision making purposes. Methods: The dataset consisted of electronic health records (EHRs) from 429 consecutive AF patients referred for EC. For analysis of the patient outcome, we considered five pathways according to restoring and maintaining SR: (i) spontaneous SR restoration, (ii) pharmacologic-cardioversion, (iii) direct-current cardioversion, (iv) 6-month AF recurrence, and (v) 6-month rhythm control. We applied ML classifiers for predicting outcomes at each pathway and compared them with the CHA2DS2-VASc and HATCH scores. Results: With the exception of pathway (iii), all ML models achieved improvements in comparison with CHA2DS2-VASc or HATCH scores (p < 0.01). Compared to the most competitive score, the area under the ROC curve (AUC-ROC) was: 0.80 vs. 0.66 for predicting (i); 0.71 vs. 0.55 for (ii); 0.64 vs. 0.52 for (iv); and 0.66 vs. 0.51 for (v). For a threshold considered optimal, the empirical net reclassification index was: +7.8%, +47.2%, +28.2%, and +34.3% in favor of our ML models for predicting outcomes for pathways (i), (ii), (iv), and (v), respectively. As an example tool of generalizability of ML models, we deployed our algorithms in an open-source calculator, where the model would personalize predictions. Conclusions: An ML model improves the accuracy of restoring and maintaining SR predictions over current discriminators. The proposed approach enables a detailed simulation of the patient flow through personalized predictions.

Plaque modification in calcified chronic total occlusions: the PLACCTON study.

INTRODUCTION AND OBJECTIVES: Severe calcification is present in> 50% of coronary chronic total occlusions (CTOs) undergoing percutaneous intervention. We aimed to describe the contemporary use and outcomes of plaque modification devices (PMDs) in this context. METHODS: Patients were included in the prospective, consecutive Iberian CTO registry (32 centers in Spain and Portugal), from 2015 to 2020. Comparison was performed according to the use of PMDs. RESULTS: Among 2235 patients, wire crossing was achieved in 1900 patients and PMDs were used in 134 patients (7%), requiring more than 1 PMD in 24 patients (1%). The selected PMDs were rotational atherectomy (35.1%), lithotripsy (5.2%), laser (11.2%), cutting/scoring balloons (27.6%), OPN balloons (2.9%), or a combination of PMDs (18%). PMDs were used in older patients, with greater cardiovascular burden, and higher Syntax and J-CTO scores. This greater complexity was associated with longer procedural time but similar total stent length (52 vs 57mm; P=.105). If the wire crossed, the procedural success rate was 87.2% but increased to 96.3% when PMDs were used (P=.001). Conversely, PMDs were not associated with a higher rate of procedural complications (3.7 vs 3.2%; P=.615). Despite the worse baseline profile, at 2 years of follow-up there were no differences in the survival rate (PMDs: 94.3% vs no-PMDs: 94.3%, respectively; P=.967). CONCLUSIONS: Following successful wire crossing in CTOs, PMDs were used in 7% of the lesions with an increased success rate. Mid-term outcomes were comparable despite their worse baseline profile, suggesting that broader use of PMDs in this setting might have potential technical and prognostic benefits.

Percutaneous Closure of Mitral Paravalvular Leak: Long-Term Results in a Single-Center Experience.

BACKGROUND: Paravalvular leak occurs in 5-17% of patients following surgical valve replacement, more often in mitral position. The prognosis without treatment is poor. Percutaneous device closure represents an alternative to repeat surgery. The objective of this work is to evaluate the medium and long-term results in the percutaneous closure of PVL in mitral prosthesis. METHODS: This observational study is based on a retrospective registry including consecutive mitral PVL cases undergoing percutaneous closure at a single tertiary-care center from April 2010 to December 2020. The safety and efficacy results of the procedure, at 90 days and in the long term, were analyzed. Also, predictors of procedure failure and long-term events were identified. RESULTS: A total of 128 consecutive mitral paravalvular leak closure procedures were included. Technical success was achieved in 115 (89.8%) procedures. The presence of multiple PVLs was the sole factor that independently predicted procedural failure. Median follow-up of our sample was 41.8 months (mean 47.7 ± 35.7 months). Underlying hemolytic anemia as the indication for PVL closure, a recent admission for decompensated HF, and lack of improvement in functional class emerged as consistent predictors of MACE and death during long-term follow-up, while lack of procedural success during the first PVL procedure and chronic kidney disease were also associated with MACE during follow-up. CONCLUSIONS: Percutaneous mitral PVL closure displayed high technical and procedural success rates, with an acceptable safety profile, in a high-risk population. Percutaneous mitral PVL closure achieved an improvement in short- and long-term functional class and a reduction of hemolysis in the vast majority of patients. In addition, long-term survival in our study was good, in particular for patients undergoing successful PVL closure procedures.

Formation of virus-like particles from human cell lines exclusively expressing influenza neuraminidase.

The minimal virus requirements for the generation of influenza virus-like particle (VLP) assembly and budding were reassessed. Using neuraminidase (NA) from the H5N1 and H1N1 subtypes, it was found that the expression of NA alone was sufficient to generate and release VLPs. Biochemical and functional characterization of the NA-containing VLPs demonstrated that they were morphologically similar to influenza virions. The NA oligomerization was comparable to that of the live virus, and the enzymic activity, whilst not required for the release of NA-VLPs, was preserved. Together, these findings indicate that NA plays a key role in virus budding and morphogenesis, and demonstrate that NA-VLPs represent a useful tool in influenza research.

[Penetrating foreign body in the left ventricle with undetected chronic evolution]. / Cuerpo extraño penetrante en el ventrículo izquierdo con evolución crónica inadvertida.

Penetrating cardiac trauma constitutes a serious threat for life. Less than the third part of all patients who suffer it arrives at the hospital alive, and half of them die. Most of the penetrating foreign bodies in the heart are metallic, and are frequently caused by firearms and rarely by self-injury. The accidental penetration is uncommon but inadvertent penetration is extremely rare. We present the case of a patient who suffered a closed-chest trauma and did not notice the penetration of a foreign body (copper wire fragment) in the heart. It remained lodged in the left ventricle for more than 3 months. This it is the only case reported in the literature where a strange body: a) crosses the free wall of the right ventricle, the right ventricular cavity, interventricular septum, and the mitral valve apparatus; b) occupies almost all the anteroposterior diameter of the heart, and c) did not produce acute or chronic bleeding.

Short-term effects of a 3-week interval training program on heart rate variability in chronic heart failure. A randomised controlled trial.

BACKGROUND: Exaggerated sympathetic nervous system activity associated with low heart rate variability (HRV) is considered to trigger cardiac arrhythmias and sudden death. Regular exercise training is efficient to improve autonomic balance. OBJECTIVE: We aimed to verify the superiority of high-intensity interval training (HIIT) to enhance HRV, cardiorespiratory fitness and cardiac function as compared with moderate intensity continuous training (MICT) in a short, intense cardiac rehabilitation program. METHODS: This was a prospective, monocentric, evaluator-blinded, randomised (1:1) study with a parallel two-group design. Overall, 31 individuals with voluntary chronic heart failure (CHF) (left ventricular ejection fraction [LVEF]<45%) were allocated to MICT (n=15) or HIIT (n=16) for a short rehabilitation program (mean [SD] 27 [4] days). Participants underwent 24-hr electrocardiography, echocardiography and a cardiopulmonary exercise test at entry and at the end of the study. RESULTS: High-frequency power in normalized units (HFnu%) measured as HRV increased with HIIT (from 21.2% to 26.4%, P<0.001) but remained unchanged with MICT (from 23.1% to 21.9%, P=0.444, with a significant intergroup difference, P=0.003). Resting heart rate (24-hr Holter electrocardiography) decreased significantly for both groups (from 68.2 to 64.6 bpm and 66.0 to 63.5 bpm for MICT and HIIT, respectively, with no intergroup difference, P=0.578). The 2 groups did not differ in premature ventricular contractions. Improvement in peak oxygen uptake was greater with HIIT than MICT (+21% vs. +5%, P=0.009). LVEF improved with only HIIT (from 36.2% to 39.5%, P=0.034). CONCLUSIONS: In this short rehabilitation program, HIIT was significantly superior to the classical MICT program for enhancing parasympathetic tone and peak oxygen uptake. CLINICALTRIALS. GOV IDENTIFIER: NCT03603743.

Automated and simplified identification of normal and abnormal plasma cells in Multiple Myeloma by flow cytometry.

BACKGROUND: Multiple myeloma (MM) is an incurable disease characterized by clonal plasma cell (PC) proliferation within the bone marrow (BM). Next-generation flow cytometry has become the reference tool to follow minimal residual disease (MRD). We developed a new simpler and cheaper flow cytometry method to analyze bone marrow samples in patients with MM. METHODS: To identify and characterize abnormal PCs, we designed a simple panel composed of anti-CD38, antikappa, and antilambda light chain antibodies, combined with two antibody pools with the same fluorophore (anti-CD19 and anti-CD27 for the negative pool and anti-CD56, anti-CD117, and anti-CD200 antibodies for the positive pool). We also developed dedicated software for the automated identification of malignant PCs and MRD assessment. We then compared PC identification with our simple antibody panel and with the larger antibody panel routinely used at Montpellier University Hospital Center in 52 patients with MM (M-CHU cohort). RESULTS: Results for total PC detection (r2  = 0.9965; P < 0.001; n = 52) and malignant PC detection (r2  = 0.9486; P < 0.001; n = 38) obtained with the two panels were significantly correlated. Moreover, comparison of the results obtained by automated detection with our software and by manual gating analysis in 80 BM samples (38 from the M-CHU cohort and 42 patients from another MM cohort) showed strong correlation for both total and malignant PC selection (respectively, r2  = 0.936; P < 0.001 and r2  = 0.9505; P < 0.001). CONCLUSIONS: Our simple and automated strategy for MRD assessment in MM could help increasing reproducibility and productivity without compromising sensitivity and specificity, while decreasing the test cost. © 2017 International Clinical Cytometry Society.

Hemagglutinin pseudotyped lentiviral particles: characterization of a new method for avian H5N1 influenza sero-diagnosis.

BACKGROUND: Highly pathogenic avian influenza (HPAI) H5N1 has spread globally in birds and infected over 270 humans with an apparently high mortality rate. Serologic studies to determine the extent of asymptomatic H5N1 infection in humans and other mammals and to investigate the immunogenicity of current H5N1 vaccine candidates have been hampered by the biosafety requirements needed for H5N1 micro-neutralization tests. OBJECTIVE: Development of a serodiagnostic tool for highly pathogenic influenza that reproduces H5N1 biology but can be used with less biohazard. STUDY DESIGN: We have generated and evaluated H5 hemagglutinin pseudotyped lentiviral particles encoding the luciferase reporter (H5pp). RESULTS: H5pp entry into target cells depends on alpha2-3 cell surface sialic acids and requires low pH for membrane fusion. H5pp infectivity is specifically neutralized by sera from patients and animals infected with H5N1 and correlates well with conventional microneutralization test. CONCLUSIONS: H5pp reproduce H5N1 influenza virus entry into target cells and potentially provides a high-throughput and safe method for sero-epidemiology.

Chemical messages from an ancient buried bottle: metabolomics for wine archeochemistry.

Restoration works in the old Clunisian Saint-Vivant monastery in Burgundy revealed an unidentified wine bottle (SV1) dating between 1772 and 1860. Chemical evidence for SV1 origin and nature are presented here using non-targeted Fourier Transform Ion Cyclotron Resonance Mass Spectrometry and Nuclear Magnetic Resonance analyses. The SV1 chemical diversity was compared to red wines (Pinot Noir) from the Romanée Saint Vivant appellation and from six different vintages spanning from 1915 to 2009. The close metabolomic signature between SV1 and Romanée Saint Vivant wines spoke in favor of a filiation between these wines, in particular considering the Pinot noir grape variety. A further statistical comparison with up to 77 Pinot noir wines from Burgundy and vintages from nearly all the 20th century, confirmed that SV1 must have been made more than one hundred years ago. The increasing number of detected high masses and of nitrogen containing compounds with the ageing of the wine was in accordance with known ageing mechanisms. Besides, resveratrol was shown here to be preserved for more than one hundred years in wine. For the first time, the age of an old unknown wine along with its grape variety have been assessed through non-targeted metabolomic analyses.

Investigation of the binding and cleavage characteristics of N1 neuraminidases from avian, seasonal, and pandemic influenza viruses using saturation transfer difference nuclear magnetic resonance.

OBJECTIVES: The main function of influenza neuraminidase (NA) involves enzymatic cleavage of sialic acid from the surface of host cells resulting in the release of the newly produced virions from infected cells, as well as aiding the movement of virions through sialylated mucus present in the respiratory tract. However, there has previously been little information on the binding affinity of different forms of sialylated glycan with NA. Our objectives were then to investigate both sialic acid binding and cleavage of neuraminidase at an atomic resolution level. DESIGN: Nuclear magnetic resonance (NMR) spectroscopy was used to investigate pH and temperature effects on binding and cleavage as well as to interrogate the selectivity of human-like or avian-like receptors for influenza neuraminidase N1 derived from a range of different influenza virus strains including human seasonal H1N1, H1N1pdm09 and avian H5N1. RESULTS: We demonstrated that an acidic pH and physiological temperature are required for efficient NA enzymatic activity; however a change in the pH had a minimum effect on the NA-sialic acid binding affinity. Our data comparing α-2,3- and α-2,6-sialyllactose indicated that the variation in neuraminidase activity on different ligands correlated with a change in binding affinity. Epitope mapping of the sialylglycans interacting with NAs from different viral origin showed different binding profiles suggesting that different binding conformations were adopted. CONCLUSIONS: The data presented in this study demonstrated that physicochemical conditions (pH in particular) could affect the NA enzymatic activity with minor effect on ligand binding. NA cleavage specificity seemed to be associated with a difference in binding affinity to different ligands, suggesting a relationship between the two events. These findings have implications regarding the replication cycle of influenza infection in the host where different sialidase activities would influence penetration through the respiratory mucin barrier and the release of the newly generated virus from the infected cells.