Vision and Change Through the Genome Consortium for Active Teaching using Next-Generation Sequencing (GCAT-SEEK)

Development of the Genome Consortium on Active Teaching using Next Generation Sequencing (GCAT-SEEK) is described. Workshops, educational modules, assessment resources, data analysis software and computer hardware available for faculty are described

Evaluation of an open-source pipeline to create patient-specific left atrial models: A reproducibility study

This work presents an open-source software pipeline to create patient-specific left atrial models with fibre orientations and a fibrDEFAULTosis map, suitable for electrophysiology simulations, and quantifies the intra and inter observer reproducibility of the model creation. The semi-automatic pipeline takes as input a contrast enhanced magnetic resonance angiogram, and a late gadolinium enhanced (LGE) contrast magnetic resonance (CMR). Five operators were allocated 20 cases each from a set of 50 CMR datasets to create a total of 100 models to evaluate inter and intra-operator variability. Each output model consisted of: (1) a labelled surface mesh open at the pulmonary veins and mitral valve, (2) fibre orientations mapped from a diffusion tensor MRI (DTMRI) human atlas, (3) fibrosis map extracted from the LGE-CMR scan, and (4) simulation of local activation time (LAT) and phase singularity (PS) mapping. Reproducibility in our pipeline was evaluated by comparing agreement in shape of the output meshes, fibrosis distribution in the left atrial body, and fibre orientations. Reproducibility in simulations outputs was evaluated in the LAT maps by comparing the total activation times, and the mean conduction velocity (CV). PS maps were compared with the structural similarity index measure (SSIM). The users processed in total 60 cases for inter and 40 cases for intra-operator variability. Our workflow allows a single model to be created in 16.72 ± 12.25 min. Similarity was measured with shape, percentage of fibres oriented in the same direction, and intra-class correlation coefficient (ICC) for the fibrosis calculation. Shape differed noticeably only with users' selection of the mitral valve and the length of the pulmonary veins from the ostia to the distal end; fibrosis agreement was high, with ICC of 0.909 (inter) and 0.999 (intra); fibre orientation agreement was high with 60.63% (inter) and 71.77% (intra). The LAT showed good agreement, where the median ± IQR of the absolute difference of the total activation times was 2.02 ± 2.45 ms for inter, and 1.37 ± 2.45 ms for intra. Also, the average ± sd of the mean CV difference was -0.00404 ± 0.0155 m/s for inter, and 0.0021 ± 0.0115 m/s for intra. Finally, the PS maps showed a moderately good agreement in SSIM for inter and intra, where the mean ± sd SSIM for inter and intra were 0.648 ± 0.21 and 0.608 ± 0.15, respectively. Although we found notable differences in the models, as a consequence of user input, our tests show that the uncertainty caused by both inter and intra-operator variability is comparable with uncertainty due to estimated fibres, and image resolution accuracy of segmentation tools

Blood outgrowth endothelial cell migration and trapping in vivo: a window into gene therapy

Human blood outgrowth endothelial cells (hBOEC) may be useful delivery-cells for gene therapy. hBOEC have high expansion capacity and stable phenotype. If incorporated into blood vessels, hBOEC could release therapeutic agents directly into the blood stream. However, little is known about lodging and homing of hBOEC in vivo. We examined the homing patterns of hBOEC in mice, and explored extending cell-based FVIII gene therapy from mice to larger animals. hBOEC were injected into NOD/SCID mice to determine where they localize, how localization changes over time and if there were toxic effects on host organs. The presence of hBOEC in mouse organs was determined by qPCR and immunofluorescence microscopy. hBOEC lodged most notably in mouse lungs at 3 h, but by 24 h there were no differences between 9 organs. hBOEC longevity was assessed up to 7 months in vivo. hBOEC expanded well and then plateaued in vivo. hBOEC from older cultures expanded equally well in vivo as younger. hBOEC caused no noticeable organ toxicity up to 3 days post-injection. When mice were pretreated with antibodies to E-selectin, P-selectin or anti-α4 integrin prior to hBOEC injection, the number of hBOEC in lungs at 3h was inhibited. Preliminary studies infusing hemophilic dogs with autologous canine BOEC over-expressing FVIII (B-domain deleted) showed improvement in whole blood clotting times (WBCT). In conclusion, the survivability, expandability and lack of toxicity of BOEC in vivo indicate that they may be valuable host cells for gene therapy

Predicting atrial fibrillation recurrence by combining population data and virtual cohorts of patient-specific left atrial models

Background: Current ablation therapy for atrial fibrillation is suboptimal, and long-term response is challenging to predict. Clinical trials identify bedside properties that provide only modest prediction of long-term response in populations, while patient-specific models in small cohorts primarily explain acute response to ablation. We aimed to predict long-term atrial fibrillation recurrence after ablation in large cohorts, by using machine learning to complement biophysical simulations by encoding more interindividual variability. Methods: Patient-specific models were constructed for 100 atrial fibrillation patients (43 paroxysmal, 41 persistent, and 16 long-standing persistent), undergoing first ablation. Patients were followed for 1 year using ambulatory ECG monitoring. Each patient-specific biophysical model combined differing fibrosis patterns, fiber orientation maps, electrical properties, and ablation patterns to capture uncertainty in atrial properties and to test the ability of the tissue to sustain fibrillation. These simulation stress tests of different model variants were postprocessed to calculate atrial fibrillation simulation metrics. Machine learning classifiers were trained to predict atrial fibrillation recurrence using features from the patient history, imaging, and atrial fibrillation simulation metrics. Results: We performed 1100 atrial fibrillation ablation simulations across 100 patient-specific models. Models based on simulation stress tests alone showed a maximum accuracy of 0.63 for predicting long-term fibrillation recurrence. Classifiers trained to history, imaging, and simulation stress tests (average 10-fold cross-validation area under the curve, 0.85±0.09; recall, 0.80±0.13; precision, 0.74±0.13) outperformed those trained to history and imaging (area under the curve, 0.66±0.17) or history alone (area under the curve, 0.61±0.14). Conclusion: A novel computational pipeline accurately predicted long-term atrial fibrillation recurrence in individual patients by combining outcome data with patient-specific acute simulation response. This technique could help to personalize selection for atrial fibrillation ablation

Atrial Fibrillation and In-Hospital Mortality in Covid-19 patients

BACKGROUND: There are conflicting data on whether new-onset atrial fibrillation (AF) is independently associated with poor outcomes in COVID-19 patients. This study represents the largest dataset curated by manual chart review comparing clinical outcomes between patients with sinus rhythm, pre-existing AF, and new-onset AF. OBJECTIVE: The primary aim of this study was to assess patient outcomes in COVID-19 patients with sinus rhythm, pre-existing AF, and new-onset AF. The secondary aim was to evaluate predictors of new-onset AF in patients with COVID-19 infection. METHODS: This was a single-center retrospective study of patients with a confirmed diagnosis of COVID-19 admitted between March and September 2020. Patient demographic data, medical history, and clinical outcome data were manually collected. Adjusted comparisons were performed following propensity score matching between those with pre-existing or new-onset AF and those without AF. RESULTS: The study population comprised of 1241 patients. A total of 94 (7.6%) patients had pre-existing AF and 42 (3.4%) patients developed new-onset AF. New-onset AF was associated with increased in-hospital mortality before (odds ratio [OR] 3.58, 95% confidence interval [CI] 1.78-7.06, P < .005) and after (OR 2.80, 95% CI 1.01-7.77, P < .005) propensity score matching compared with the no-AF group. However, pre-existing AF was not independently associated with in-hospital mortality compared with patients with no AF (postmatching OR: 1.13, 95% CI 0.57-2.21, P = .732). CONCLUSION: New-onset AF, but not pre-existing AF, was independently associated with elevated mortality in patients hospitalised with COVID-19. This observation highlights the need for careful monitoring of COVID-19 patients with new-onset AF. Further research is needed to explain the mechanistic relationship between new-onset AF and clinical outcomes in COVID-19 patients

Atrial CARdiac Magnetic resonance imaging in patients with embolic stroke of unknown source without documented Atrial Fibrillation (CARM-AF): Study design and clinical protocol

Background: Initiation of anticoagulation therapy in ischemic stroke patients is contingent on a clinical diagnosis of atrial fibrillation (AF). Results from previous studies suggest thromboembolic risk may predate clinical manifestations of AF. Early identification of this cohort of patients may allow early initiation of anticoagulation and reduce the risk of secondary stroke. Objective: This study aims to produce a substrate-based predictive model using cardiac magnetic resonance imaging (CMR) and baseline noninvasive electrocardiographic investigations to improve the identification of patients at risk of future thromboembolism. Methods: CARM-AF is a prospective, multicenter, observational cohort study. Ninety-two patients will be recruited following an embolic stroke of unknown source (ESUS) and undergo atrial CMR followed by insertion of an implantable loop recorder (ILR) as per routine clinical care within 3 months of index stroke. Remote ILR follow-up will be used to allocate patients to a study or control group determined by the presence or absence of AF as defined by ILR monitoring. Results: Baseline data collection, noninvasive electrocardiographic data analysis, and imaging postprocessing will be performed at the time of enrollment. Primary analysis will be performed following 12 months of continuous ILR monitoring, with interim and delayed analyses performed at 6 months and 2 and 3 years, respectively. Conclusion: The CARM-AF Study will use atrial structural and electrocardiographic metrics to identify patients with AF, or at high risk of developing AF, who may benefit from early initiation of anticoagulation

High prevalence of new clinically significant findings in patients with embolic stroke of unknown source evaluated by cardiac magnetic resonance imaging

Background: Embolic stroke of unknown source (ESUS) accounts for one in six ischaemic strokes. Current guidelines do not recommend routine cardiac magnetic resonance (CMR) imaging in ESUS and, beyond the identification of cardio-embolic sources, there are no data assessing new clinical findings from CMR in ESUS. This study aimed to assess the prevalence of new cardiac and non-cardiac findings and to determine their impact on clinical care in patients with ESUS.Methods and Results: In this prospective, multicentre, observational study, CMR was performed within 3-months of ESUS. All scans were reported according to standard clinical practice. A new clinical finding was defined as one not previously identified through prior clinical evaluation. A clinically significant finding was defined as one resulting in further investigation, follow-up or treatment. A change in patient care was defined as initiation of medical, interventional, surgical or palliative care. From 102 patients recruited, 96 underwent CMR. One or more new clinical findings were observed in 59 patients (61%). New findings were clinically significant in 48 (81%) of these patients. Of 40 patients with a new clinically significant cardiac finding, 21 (53%) experienced a change in care (medical therapy, n=15; interventional/surgical procedure, n=6). In 12 patients with a new clinically significant extra-cardiac finding, 6 (50%) experienced a change in care (medical therapy, n=4; palliative care, n=2). Conclusions: CMR imaging identifies new clinically significant cardiac and non-cardiac findings in half of patients with recent ESUS. Advanced cardiovascular screening should be considered in patients with ESUS.<br/

Erythroid-Specific Expression of β-globin from Sleeping Beauty-Transduced Human Hematopoietic Progenitor Cells

Gene therapy for sickle cell disease will require efficient delivery of a tightly regulated and stably expressed gene product to provide an effective therapy. In this study we utilized the non-viral Sleeping Beauty (SB) transposon system using the SB100X hyperactive transposase to transduce human cord blood CD34+ cells with DsRed and a hybrid IHK–β-globin transgene. IHK transduced cells were successfully differentiated into multiple lineages which all showed transgene integration. The mature erythroid cells had an increased β-globin to γ-globin ratio from 0.66±0.08 to 1.05±0.12 (p = 0.05), indicating expression of β-globin from the integrated SB transgene. IHK–β-globin mRNA was found in non-erythroid cell types, similar to native β-globin mRNA that was also expressed at low levels. Additional studies in the hematopoietic K562 cell line confirmed the ability of cHS4 insulator elements to protect DsRed and IHK–β-globin transgenes from silencing in long-term culture studies. Insulated transgenes had statistically significant improvement in the maintenance of long term expression, while preserving transgene regulation. These results support the use of Sleeping Beauty vectors in carrying an insulated IHK–β-globin transgene for gene therapy of sickle cell disease