In Vivo Target Gene Activation via CRISPR/Cas9-Mediated Trans-epigenetic Modulation.

Current genome-editing systems generally rely on inducing DNA double-strand breaks (DSBs). This may limit their utility in clinical therapies, as unwanted mutations caused by DSBs can have deleterious effects. CRISPR/Cas9 system has recently been repurposed to enable target gene activation, allowing regulation of endogenous gene expression without creating DSBs. However, in vivo implementation of this gain-of-function system has proven difficult. Here, we report a robust system for in vivo activation of endogenous target genes through trans-epigenetic remodeling. The system relies on recruitment of Cas9 and transcriptional activation complexes to target loci by modified single guide RNAs. As proof-of-concept, we used this technology to treat mouse models of diabetes, muscular dystrophy, and acute kidney disease. Results demonstrate that CRISPR/Cas9-mediated target gene activation can be achieved in vivo, leading to measurable phenotypes and amelioration of disease symptoms. This establishes new avenues for developing targeted epigenetic therapies against human diseases. VIDEO ABSTRACT.

Interspecies Chimerism with Mammalian Pluripotent Stem Cells.

Interspecies blastocyst complementation enables organ-specific enrichment of xenogenic pluripotent stem cell (PSC) derivatives. Here, we establish a versatile blastocyst complementation platform based on CRISPR-Cas9-mediated zygote genome editing and show enrichment of rat PSC-derivatives in several tissues of gene-edited organogenesis-disabled mice. Besides gaining insights into species evolution, embryogenesis, and human disease, interspecies blastocyst complementation might allow human organ generation in animals whose organ size, anatomy, and physiology are closer to humans. To date, however, whether human PSCs (hPSCs) can contribute to chimera formation in non-rodent species remains unknown. We systematically evaluate the chimeric competency of several types of hPSCs using a more diversified clade of mammals, the ungulates. We find that naïve hPSCs robustly engraft in both pig and cattle pre-implantation blastocysts but show limited contribution to post-implantation pig embryos. Instead, an intermediate hPSC type exhibits higher degree of chimerism and is able to generate differentiated progenies in post-implantation pig embryos.

In Vivo Amelioration of Age-Associated Hallmarks by Partial Reprogramming.

Aging is the major risk factor for many human diseases. In vitro studies have demonstrated that cellular reprogramming to pluripotency reverses cellular age, but alteration of the aging process through reprogramming has not been directly demonstrated in vivo. Here, we report that partial reprogramming by short-term cyclic expression of Oct4, Sox2, Klf4, and c-Myc (OSKM) ameliorates cellular and physiological hallmarks of aging and prolongs lifespan in a mouse model of premature aging. Similarly, expression of OSKM in vivo improves recovery from metabolic disease and muscle injury in older wild-type mice. The amelioration of age-associated phenotypes by epigenetic remodeling during cellular reprogramming highlights the role of epigenetic dysregulation as a driver of mammalian aging. Establishing in vivo platforms to modulate age-associated epigenetic marks may provide further insights into the biology of aging.

The high optical brightness of the BlueWalker 3 satellite.

Large constellations of bright artificial satellites in low Earth orbit pose significant challenges to ground-based astronomy1. Current orbiting constellation satellites have brightnesses between apparent magnitudes 4 and 6, whereas in the near-infrared Ks band, they can reach magnitude 2 (ref. 2). Satellite operators, astronomers and other users of the night sky are working on brightness mitigation strategies3,4. Radio emissions induce further potential risk to ground-based radio telescopes that also need to be evaluated. Here we report the outcome of an international optical observation campaign of a prototype constellation satellite, AST SpaceMobile's BlueWalker 3. BlueWalker 3 features a 64.3 m2 phased-array antenna as well as a launch vehicle adaptor (LVA)5. The peak brightness of the satellite reached an apparent magnitude of 0.4. This made the new satellite one of the brightest objects in the night sky. Additionally, the LVA reached an apparent V-band magnitude of 5.5, four times brighter than the current International Astronomical Union recommendation of magnitude 7 (refs. 3,6); it jettisoned on 10 November 2022 (Universal Time), and its orbital ephemeris was not publicly released until 4 days later. The expected build-out of constellations with hundreds of thousands of new bright objects1 will make active satellite tracking and avoidance strategies a necessity for ground-based telescopes.

Paravalvular leak closure after TAVI with plugs: A literature review with a specific focus on arterio-arterial loop.

More than mild paravalvular leak (PVL) following transcatheter aortic valve implantation (TAVI) is associated with a twofold increase in all-cause mortality, heart failure hospitalizations, and the need for reintervention. Successfully addressing PVL in TAVIs is more challenging than in surgical valves. The arterial-arterial (A-A) rail technique emerges as a valuable strategy for post-TAVI PVL closure, enhancing success rates by enabling the effective use of lower-profile vascular plug devices. When standard approach is ineffective, generating an A-A loop for post-TAVI PVL closure is probably the most recommended strategy to ensure procedural success.

The association of hematological markers with occurrence of thrombotic and bleeding events following left atrial appendage occlusion.

Introduction Patients undergoing left atrial appendage occlusion (LAAO) are at increased risk for bleeding or thromboembolic events. Concurrently, biomarkers are of growing importance in risk stratification for atrial fibrillation patients. We aimed to evaluate the association of hematological markers and clinical characteristics with the occurrence of thromboembolic and bleeding events following LAAO. Methods Seven implanting centers retrospectively gathered data on hematological markers (i.e. platelet count (PC), mean platelet volume (MPV), and fibrinogen) prior LAAO. Prespecified thromboembolic and major bleeding outcomes were collected and the association with pre-procedural hematological markers and clinical characteristics was evaluated using Cox regression analysis. Results In total, 1315 patients were included (74±9 years, 36% female, CHA2DS2-VASc 4.3±1.5, HAS-BLED 3.3±1.1). Over a total follow-up duration of 2682 patient years, 77 thromboembolic events and 107 major bleeding events occurred after LAAO. Baseline PC was the only biomarker showing a signal for a relation to thromboembolic events (HR 1.18, 95% CI: 1.00-1.39) per 50*109 increment, p=0.056). Thrombotic event rates, including device-related thrombus, increased within higher PC quartiles. Thromboembolism was associated with age (HR 1.05, 95% CI: 1.00-1.10, per year increase) and prior thromboembolism (HR 2.08, 95% CI: 1.07-4.03), but with none of the biomarkers in multivariate analysis. No association of any of the hematological markers with major bleeding was observed. Major bleeding following LAAO was associated with prior major bleeding (HR 5.27, 95% CI: 2.71-10.22), renal disease (HR 1.93, 95% CI: 1.17-3.18) and discharge on dual antiplatelet therapy (HR 1.71, 95% CI: 1.05-2.77). Conclusion Most thrombotic events occurred in the highest PC quartile, but no association of any of the hematological markers with thromboembolism or major bleeding was observed in our analysis. In multivariate analysis, older age and prior thromboembolism were associated with thromboembolism. Prior major bleeding, renal disease and discharge on DAPT were multivariate predictors of major bleeding after LAAO.

Usefulness of a novel sizing chart for left atrial appendage occlusion with the Amplatzer Amulet.

PURPOSE: The present article aims to compare a novel sizing chart based on both maximum and minimum diameters (novel MATRIX) with the current sizing recommendation instructions for use (IFU) based on the maximum diameter. BACKGROUND: Current IFU with the Amulet device are still based on the maximum left atrial appendage (LAA) diameter, which might lead to inappropriate oversizing, especially in elliptic appendages. METHODS: This was a retrospective analysis of patients undergoing LAA occlusion in two high-volume centers. Two hundred patients were included (100 patients with baseline cardiac computed tomography angiography [CCTA] and 100 with baseline 2D and 3D-transesophageal echocardiography [TEE]). The degree of concordance between the predicted device size recommendation and the actual device selection was the primary outcome. RESULTS: The novel MATRIX showed a higher level of concordance between the predicted and implanted device size, regardless of imaging modalities. CCTA showed the strongest, and 2D-TEE the weakest concordance between the predicted and implanted device for both MATRIX and IFU charts. The percentage of patients in whom the disagreement among the predicted and implanted device represented >1 size was higher when using the IFU chart. In elliptical LAA anatomies, the differences favoring the use of MATRIX compared to the IFU in terms of predicted/implanted agreement were higher. Finally, no significant differences in clinical or imaging endpoints were observed between the two different sizing charts. CONCLUSIONS: Incorporating both the LAA maximum and minimum diameters, as opposed to just maximum diameter, appears to improve sizing accuracy. The proposed MATRIX sizing chart offered a higher level of concordance between predicted and implanted device compared to the current IFU.

In vivo genome editing via CRISPR/Cas9 mediated homology-independent targeted integration.

Targeted genome editing via engineered nucleases is an exciting area of biomedical research and holds potential for clinical applications. Despite rapid advances in the field, in vivo targeted transgene integration is still infeasible because current tools are inefficient, especially for non-dividing cells, which compose most adult tissues. This poses a barrier for uncovering fundamental biological principles and developing treatments for a broad range of genetic disorders. Based on clustered regularly interspaced short palindromic repeat/Cas9 (CRISPR/Cas9) technology, here we devise a homology-independent targeted integration (HITI) strategy, which allows for robust DNA knock-in in both dividing and non-dividing cells in vitro and, more importantly, in vivo (for example, in neurons of postnatal mammals). As a proof of concept of its therapeutic potential, we demonstrate the efficacy of HITI in improving visual function using a rat model of the retinal degeneration condition retinitis pigmentosa. The HITI method presented here establishes new avenues for basic research and targeted gene therapies.

Survival benefit of revascularization versus optimal medical therapy alone for chronic total occlusion management in patients with diabetes.

BACKGROUND: Chronic total occlusion (CTO) is common in patients with diabetes mellitus. Data on the long-term outcomes after treatment of CTOs in this high-risk population are scarce. AIM: To compare the long-term clinical outcomes of CTO revascularization either by coronary artery bypass graft (CABG) or successful percutaneous coronary intervention (PCI) versus optimal medical treatment (MT) alone in patients with diabetes. METHODS AND RESULTS: A total of 538 consecutive patients with diabetes and at least one CTO were identified from 2010 to 2014 in our center. In the present analysis, patients were stratified according to the CTO treatment strategy that was selected. MT was selected in 61% of patients whereas revascularization in the remaining 39%. Patients undergoing revascularization were younger, had higher left ventricular ejection fraction (LVEF), lower ACEF score, and more positive myocardial ischemia detection results compared to the MT group (p < .001).Patients referred for CABG had higher rates of left main disease compared to the PCI and MT groups (32% vs. 3% and 11%, respectively; p < .001). Complete revascularization was more often achieved in the CABG group, compared to the PCI group (62% vs. 32% p < .001). Multivariable analysis showed that revascularization with CABG was associated with lower rates of all-cause and cardiac mortality rates compared to MT, [hazard ratio (HR) 0.41, 95% confidence interval (CI) 0.25-0.70, p < .001 and HR 0.40, 95% CI 0.20-81, p = .011, respectively]. Successful CTO-PCI showed a trend towards benefit in all-cause mortality (HR 0.58, 95% CI 0.33-1.04, p = .06). CONCLUSION: In our registry, CTO revascularization in diabetic patients, especially with CABG, was associated with lower long-term mortality rates as compared to MT alone.

An alternative pluripotent state confers interspecies chimaeric competency.

Pluripotency, the ability to generate any cell type of the body, is an evanescent attribute of embryonic cells. Transitory pluripotent cells can be captured at different time points during embryogenesis and maintained as embryonic stem cells or epiblast stem cells in culture. Since ontogenesis is a dynamic process in both space and time, it seems counterintuitive that these two temporal states represent the full spectrum of organismal pluripotency. Here we show that by modulating culture parameters, a stem-cell type with unique spatial characteristics and distinct molecular and functional features, designated as region-selective pluripotent stem cells (rsPSCs), can be efficiently obtained from mouse embryos and primate pluripotent stem cells, including humans. The ease of culturing and editing the genome of human rsPSCs offers advantages for regenerative medicine applications. The unique ability of human rsPSCs to generate post-implantation interspecies chimaeric embryos may facilitate our understanding of early human development and evolution.

Percutaneous left atrial appendage closure, a safe alternative to anticoagulation for patients with nonvalvular atrial fibrillation and end-stage renal disease on hemodialysis: A single center experience.

The evidence about the effectiveness and safety of oral anticoagulation in patients on hemodialysis is conflicting and scarce. Percutaneous left atrial appendage occlusion (LAAO) has demonstrated to be a valid alternative therapeutic option for stroke prevention in patients with nonvalvular atrial fibrillation (NVAF). The aim of this study is to present the outcomes of percutaneous LAAO in patients with end-stage renal disease (ESRD) on hemodialysis and NVAF in our center. We conducted a retrospective review of clinical records, demographics, LAAO procedure, complications, and outcomes of patients with NVAF and ESRD on hemodialysis who underwent a percutaneous LAAO in our center between January 2017 and January 2019. In the period of the study, eight patients with ESRD on hemodialysis underwent a percutaneous LAAO in our center. The overall mean age was 67.5 years (range 56-81; SD ± 7.2). All patients had permanent NVAF. The total mean dialysis duration was 8.49 years (range 0.83-14.8; SD ± 6.2). The mean CHA2DS2-VASc and HAS-BLED scores were high (4.75 [SD ± 1.16] and 4.62 [SD ± 0.91], respectively). All patients had history of a major hemorrhagic event (BARC Score ≥3). Most patients (n = 6) showed left ventricular hypertrophy, and the average LVEF was 54% (SD ± 6.5). All devices were implanted successfully. Postprocedural antithrombotic regimen prescribed was based on antiplatelet therapy. No deaths, cardioembolic events, or major bleeding (according to the BARC scale) were reported during a mean follow-up of 14.24 months (SD ± 9.44). Percutaneous LAAO could be of particular interest in patients with NVAF and CKD in hemodialysis. Further studies will be necessary to confirm this hypothesis.

Impact of revascularization versus medical therapy alone for chronic total occlusion management in older patients.

BACKGROUND: Chronic total occlusions (CTOs) are present in more than one third of older patients with myocardial ischemia, but controversy remains about the best therapeutic approach. AIMS: To compare long-term survival after CTO revascularization (percutaneous coronary intervention [PCI] or coronary artery bypass graft [CABG]) versus medical treatment (MT) alone in patients aged 75 and older. METHODS AND RESULTS: A total of 1,252 consecutive patients with at least one CTO were identified from 2010 to 2014 in our center. Patients were stratified by age (<75 years vs. ≥75 years) in the present analysis. All-cause and cardiac mortality were assessed at a median follow-up of 3.5 years. In the older subgroup (26%), patients were more likely to be treated with MT alone (71% vs. 43% of younger patients; p < 0.001). Patients undergoing revascularization were younger and had higher left ventricular ejection fraction (LVEF) and lower age, creatinine, ejection fraction (ACEF) score (age/LVEF +1 if creatinine >2.0 mg/dL), compared to the MT group (p < 0.05). As compared to MT, revascularization predicted lower rates of cardiac mortality and all-cause mortality in older patients, both in the subgroups treated with CABG (hazard ratio [HR] 0.35, 95% confidence interval [CI] 0.17-0.71; HR 0.39, 95%CI 0.18-0.81) and PCI (HR 0.57, 95%CI 0.33-0.98; HR 0.59, 95%CI 0.28-1.2). No differences in mortality were observed according to type of revascularization procedure. CONCLUSIONS: Among patients aged at least 75 years with a CTO, revascularization (PCI or CABG) rather than MT alone may portend a better outcome in terms of all-cause and cardiac mortality.

Predictors of renal replacement therapy after heart transplantation and its impact on long-term survival.

BACKGROUND: Renal replacement therapy (RRT) after heart transplant (HT) is associated with worse prognosis. We aimed to identify predictors of RRT and the impact of this complication on long-term survival. METHODS: Cohort study of HT patients. Univariate and multivariate competing-risk regression was performed to identify independent predictors of RRT. The cumulative incidence function was plotted for RRT. The Kaplan-Meier method was used to compare long-term survival. RESULTS: We included 103 patients. At multivariate analysis, only the emergency status of HT (short-term mechanical circulatory support as a bridge to transplant), chronic kidney disease, and low oxygen delivery were independent predictors of RRT (subhazard ratio [SHR] 4.11, 95% CI 1.84-9.14; SHR 3.17, 95% CI 1.29-7.77; SHR 2.86, 95% CI 1.14-7.19, respectively). Elective HT patients that required RRT showed a significantly reduced survival comparable to patients with emergency HT and RRT (75% ± 13% vs. 67% ± 16%). The absence of RRT implied an excellent survival in patients with an emergency status of HT and elective HT (100% vs. 93% ± 4%). CONCLUSION: The emergency status of HT, chronic kidney disease, and low oxygen delivery were independent predictors of RRT. The occurrence of RRT increases the risk of death in elective HT as much as in patients with an emergency status.

Functional rehabilitation based on therapeutic exercise training in patients with postacute COVID syndrome (RECOVER).

INTRODUCTION AND OBJECTIVES: Postacute COVID syndrome (PACS) is common after acute SARS-CoV-2 infection. One of the most frequent and disabling symptoms is exercise intolerance (EI). Recent evidence suggests that EI in PACS has a peripheral (metabolic-neuromuscular) origin, suggesting that exercise training may be an effective treatment. The aim of this study was to assess the role a therapeutic physical exercise program (TPEP) in PACS with EI. METHODS: This single-center, open-label, randomized clinical trial compared an exercise training program (intervention group) with regular physical activity recommendations (control group) in patients with PACS and EI. The intervention group underwent an 8-week TPEP. The primary endpoint was improvement in functional capacity, assessed as the change in peak VO2. RESULTS: We included 50 participants with PACS (73% women, mean age 47±7.1 years). The intervention group showed a 15% improvement in peak VO2 (peak VO2 pre- and postintervention: 25.5±7.7mL/kg/min and 29.3±4.7 mL/kg/min; P <.001) and a 13.2% improvement in predicted values (92.1±14.3% and 108.4±13.4%; P <.001). No significant changes in VO2 values were observed in the control group. Unlike the control group, the intervention group also showed improvements in all secondary outcomes: quality of life scales, muscle power, maximum inspiratory power, metabolic flexibility, and body fat percentage. CONCLUSIONS: The program improved functional capacity in patients with PACS and EI.

The Impact of Geriatric Conditions in Elderly Patients with Coronary Heart Disease: A State-of-the-Art Review.

The growing geriatric population presenting with coronary artery disease poses a primary challenge for healthcare services. This is a highly heterogeneous population, often underrepresented in studies and clinical trials, with distinctive characteristics that render them particularly vulnerable to standard management/approaches. In this review, we aim to summarize the available evidence on the treatment of acute coronary syndrome in the elderly. Additionally, we contextualize frailty, comorbidity, sarcopenia, and cognitive impairment, common in these patients, within the realm of coronary artery disease, proposing strategies for each case that may assist in therapeutic approaches.

Intervention with metabolites emulating endogenous cell transitions accelerates muscle regeneration in young and aged mice.

Tissue regeneration following an injury requires dynamic cell-state transitions that allow for establishing the cell identities required for the restoration of tissue homeostasis and function. Here, we present a biochemical intervention that induces an intermediate cell state mirroring a transition identified during normal differentiation of myoblasts and other multipotent and pluripotent cells to mature cells. When applied in somatic differentiated cells, the intervention, composed of one-carbon metabolites, reduces some dedifferentiation markers without losing the lineage identity, thus inducing limited reprogramming into a more flexible cell state. Moreover, the intervention enabled accelerated repair after muscle injury in young and aged mice. Overall, our study uncovers a conserved biochemical transitional phase that enhances cellular plasticity in vivo and hints at potential and scalable biochemical interventions of use in regenerative medicine and rejuvenation interventions that may be more tractable than genetic ones.

5-Year Outcomes of Trans Carotid Transcatheter Aortic Valve Replacement.

BACKGROUND: Trans-carotid Transcatheter Aortic Valve Replacement (TC-TAVR) has emerged as an alternative TAVR access. However, scarce data on long-term outcomes following TC-TAVR exist. This study aimed to evaluate 5-year clinical outcomes post-TC-TAVR. METHODS: One hundred ten consecutive patients who underwent TC-TAVR and completed the 5-year after the procedure were included. Baseline, procedural, and follow-up data were collected prospectively in a dedicated database. The primary endpoint was the incidence of a composite of all-cause mortality, stroke, and repeat hospitalization. Echocardiography results, New York Heart Association (NYHA) class, and quality of life (QOL) assessed with the EuroQol visual analogue scale (EQ-VAS) were examined over the 5-year follow-up. RESULTS: The median age was 77 (72-82.2) years, 42.3% were women, and the median STS risk score was 5.02% (3.4-7.5%). The incidence of the composite endpoint was 54.5%. Death from any cause occurred in 45.6% of cases (11.9 per 100 patients-years), stroke in 8.2% (1.9 per 100 patient-years), disabling stroke in 2.7% (0.7 per 100 patient-years), and rehospitalization in 27.2%. The improvements in valve hemodynamics, NYHA class, and EQ-VAS following the procedure persisted at the 5-year follow-up (p<.001). Bioprosthetic valve failure occurred in 0.9%. CONCLUSIONS: About half of the moderate-to-high-risk patients undergoing TC-TAVR survived with no major cardiovascular events at 5-year follow-up. The yearly incidence of strokes was low, and early improvements in valve hemodynamics, functional status and QOL persisted at 5 years. These results suggest the long-term safety and efficacy of TC-TAVR, and would support this approach as an alternative to surgery in non-transfemoral candidates.

Inflammation as a New Therapeutic Target among Older Patients with Ischemic Heart Disease.

Cardiovascular (CV) diseases remain a global health challenge, with ischemic heart disease (IHD) being the primary cause of both morbidity and mortality. Despite optimal pharmacological therapy, older patients with IHD exhibit an increased susceptibility to recurrent ischemic events, significantly impacting their prognosis. Inflammation is intricately linked with the aging process and plays a pivotal role in the evolution of atherosclerosis. Emerging anti-inflammatory therapies have shown promise in reducing ischemic events among high-risk populations. This review aims to explore the potential of targeted anti-inflammatory interventions in improving clinical outcomes and the quality of life for older patients with IHD.