Abstract The evolutionary conserved NEAT1-MALAT1 gene cluster generates large noncoding transcripts remaining nuclear, while tRNA-like transcripts (mascRNA, menRNA) enzymatically generated from these precursors translocate to the cytosol. NEAT1-/- and MALAT1-/- mice display massive atherosclerosis and vascular inflammation. Here, we identify the tRNA-like molecules as critical components of innate immunity. They appear as prototypes of a new class of noncoding RNAs distinct from others (miRNAs, siRNAs) by biosynthetic pathway and intracellular kinetics. CRISPR-generated human ΔmascRNA and ΔmenRNA monocytes/macrophages display defective innate immune sensing, loss of cytokine control, imbalance of growth/angiogenic factor expression impacting upon angiogenesis, and altered cell-cell interaction systems. Antiviral response, foam cell formation/oxLDL uptake, and M1/M2 polarization are defective in ΔmascRNA/ΔmenRNA macrophages, defining the tRNA-like molecules’ first described biological functions. menRNA and mascRNA represent novel components of innate immunity arising from the noncoding genome. Their NEAT1-MALAT1 region of origin appears as archetype of a functionally highly integrated RNA processing system.
Abstract Introduction Risk stratification based on Euroscore II (ESII) is currently used by clinicians to assist their decision to perform transcatheter aortic valve implant (TAVI) procedures. ESII is a generic, non-TAVI-specific metric. We investigated if a TAVI-specific predictive model could achieve improved predictive pre-interventional accuracy of 1-year mortality compared to ESII. Patients & Methods: In this prospective pilot study, 284 participants with severe aortic valve stenosis who underwent TAVI were enrolled. Standard clinicals metrics (ASA, NYHA, ESII) and patient-reported outcome measures (EuroQol-5D-VAS, Kansas City Cardiomyopathy Questionnaire, Clinical Frailty Scale-CFS) were assessed one day before TAVI. Using these data, we tested predictive models (logistic regression, decision tree algorithm- DTA) with 1-year mortality as the dependent variable. Results Logistic regression yielded the best prediction, with ASA and CFS as the strongest predictors. Our logistic regression model score showed significantly better prediction accuracy than ESII (AUC = 0.659 vs. 0.800; p = 0.002). By translating our results to a DTA, cut-off score values regarding 1-year mortality risk emerged: < 0.02 points for low, 0.02 – 0.15 for intermediate, > 0.15 for high risk. Treatment costs and length of stay significantly increased in high-risk patients. Conclusions & significance A novel TAVI-specific model predicts 1-year mortality, LoS and costs after TAVI using simple, established, transparent and inexpensive metrics before implantation. Based on this preliminary evidence, TAVI team members and patients can make informed decisions based on a few key metrics. Validation of this score in larger patient cohorts is needed. Comparison of ROC curves
Abstract The evolutionary conserved NEAT1-MALAT1 gene cluster generates large noncoding transcripts remaining nuclear, while tRNA-like transcripts (mascRNA, menRNA) enzymatically generated from these precursors translocate to the cytosol. NEAT1-/- and MALAT1-/- mice display massive atherosclerosis and vascular inflammation. Here, we identify the tRNA-like molecules as critical components of innate immunity. They appear as prototypes of a new class of noncoding RNAs distinct from others (miRNAs, siRNAs) by biosynthetic pathway and intracellular kinetics. CRISPR-generated human ΔmascRNA and ΔmenRNA monocytes/macrophages display defective innate immune sensing, loss of cytokine control, imbalance of growth/angiogenic factor expression impacting upon angiogenesis, and altered cell-cell interaction systems. Antiviral response, foam cell formation/oxLDL uptake, and M1/M2 polarization are defective in ΔmascRNA/ΔmenRNA macrophages, defining the tRNA-like molecules’ first described biological functions. menRNA and mascRNA represent novel components of innate immunity arising from the noncoding genome. Their NEAT1-MALAT1 region of origin appears as archetype of a functionally highly integrated RNA processing system.
Abstract Conduction disturbances after transcatheter aortic valve replacement (TAVR) remain one of the most frequent complications. The aim of this study was to analyze ECG changes after TAVR using contemporary valves and to detect risk factors for the need of further clinical evaluation according to new ESC pacing guidelines to evaluate pacemaker implantation. In this retrospective analysis we included 850 patients (mean age 80±9 years, 51% female), who underwent TAVR in our institution from January 2019 until December 2020. A mean follow-up of 8.9±8.4 months and 217 (25.5%) patients was performed. 55% of the implanted valves were self-expandable, 45% balloon-expandable. After TAVR, 77 (9.1%) patients developed new LBBB and QRS >150ms, 26 (3.1%) new PR-time >240ms. Prolongation of PR-time and prolongation of QRS duration >20ms were seen in 20 (2.4%) and 90 (10.6%) patients with preexisting conduction disturbances. 152 (17.9%) patients needed pacemaker implantation post TAVR. Developing a PR-prolongation of >20ms was associated with calcification of the annulus (OR 1.2 CI 95% 1.004–1.4; p=0.04). New LBBB (OR 0.45; CI 95% 0.25–0.79; p=0.006) and pacemaker implantation (OR 0.4; CI 95% 0.2–0.8; p=0.009) were correlated with the implantation of a self-expandable valve. Coronary heart disease (OR 3, CI 95% 1.07–8.2; p=0.04) and peripheral arterial disease (OR 2.6 CI 95% 1.18–5.6; p=0.02) were associated with prolongation of QRS >20ms. New LBBB with QRS >150ms was seen more often after post-dilatation (OR 1.03, CI 95% 1.01–1.05; p=0.05). Pre-existing AV block I° (OR 2.8, CI 95% 1.4–5.6; p<0.001), pre-existing RBBB (OR 20.5, CI 95% 7.5–56; p<0.001), nicotine abuse (OR 2, CI 95% 1.05–3.8; p=0.04), prosthesis oversizing (OR 1.06, CI 95% 1.006–1.11; p=0.03) and implantation depth (OR 1.13, CI 95% 1.006–1.26; p=0.04) were independent risk factors for pacemaker implantation. During the follow-up 161 patients (18.9%) were hospitalized in 270 inpatient stays [cardiac decompensation (n=36, 13%), pacemaker implantation (n=9, 3.3%), acute coronary syndrome (n=12, 4.4%)]. 8 patients (80%) received a pacemaker implantation because of AV Block III° and 1 (10%) patient because of sick-sinus-syndrome (SSS). Analyzing the post TAVR ECG 5 (50%) had a new LBBB (3 (30%) with QRS >150ms) and 4 (40%) patients showed LBBB together with AV Block I°. According to new guidelines 213 (25.1%) patients would have needed further clinical evaluation (EP study or ECG monitoring) after TAVR. AV-conduction abnormalities were associated with annulus calcification. Self-expandable valves were associated with new LBBB and pacemaker implantation. There seems to be a correlation between arteriosclerotic diseases and QRS width post TAVR. Pre-existing RBBB, AV block I°, implantation depth and prosthesis oversizing are important risk factors for pacemaker implantation post TAVR. New LBBB after TAVR is associated with a higher risk for pacemaker implantation in the long-term analysis. Funding Acknowledgement Type of funding sources: None.
Abstract Background Cerebral embolization in patients after Transcatheter Aortic Valve Replacement (TAVR) represents a serious complication, that was related to impaired bioprosthetic leaflet motion and new-onset atrial fibrillation (AFib). Purpose Hereafter we present the first randomized study comparing the effect of an anticoagulation plus antiplatelet with a dual antiplatelet antithrombotic treatment in patients after TAVR on cerebral embolizations as assessed by serial cerebral magnetic resonance imaging (MRI). Methods The Evaluation of Cerebral Thrombembolism After TAVR (EARTH - TAVR) study was conducted as an investigator initiated substudy of the multicenter, randomized, GALILEO study. After successful TAVR, patients without indication for chronic anticoagulation were randomly assigned to rivaroxaban 10mg plus acetylsalicylic acid 75–100mg once-daily or clopidogrel 75mg plus acetylsalicylic acid 75–100mg once-daily. Cerebral MRI scans were performed pre-TAVR as a baseline, post-TAVR (within 24–48 hours after TAVR) and 90 days after TAVR. The MRI protocol included diffusion-weighted (DWI) and fluid-attenuated inversion recovery (FLAIR) imaging. Cerebral embolic lesions were evaluated by an independent cerebral MRI core lab. The primary outcome measure of this study was the occurrence and extent of cerebral embolizations as measured by total volume of new ischaemic cerebral lesions. Results 36 patients were enrolled in the EARTH and the GALILEO study. The DWI MRI scans revealed an increase of cerebral lesions and volume post-TAVR by a median of 4.75 (95% NBCI 2.1–8.9) and 0.26cm3 (95% NBCI 0.11–0.59). On FLAIR imaging, lesion number and volume increased by a median of 3 (95% NBCI 1.5–6) and 0.1 cm3 (95% NBCI 0.04–0.31). At the 90 days MRI scan, there was no statistically significant change in cerebral lesions, if compared to the post-TAVR scan, irrespective of the treatment arm. Conclusion Thromboembolic events occur largely in the periinterventional phase post TAVR. Thereafter, the risk for additional cerebral embolization is low. An additional rivaroxaban therapy beyond antiplatelet inhibition did not impact on cerebral thromboembolism. Funding Acknowledgement Type of funding source: Private company. Main funding source(s): Bayer Pharmaceuticals
Abstract Background Neutrophil granulocytes are key players of the innate immunity, participating in the initiation and progression of atherosclerosis. However, the exact mechanisms of neutrophil activation after acute coronary syndrome (ACS) are poorly understood, especially in the context of the two predominant ACS-causing pathophysiologies - ACS with intact fibrous cap (IFC-ACS) and ACS with ruptured fibrous cap (RFC-ACS). Therefore, the current study focuses on immunophenotyping and ex-vivo functional characterization of neutrophils with regard to the molecular differences between IFC-ACS and RFC-ACS. Methods Using high-resolution optical coherence tomography (OCT) of the ACS-causing culprit lesion and re-evaluation by a second OCT-core lab, thirty-two IFC-ACS-patients were matched to thirty-two RFC-ACS-patients by gender, age and diabetes. Local and systemic blood samples were obtained from the site of the ACS-causing culprit lesion (LOC) and from the arterial sheath (SYS), respectively. Neutrophil abundance and surface marker expression were quantified by flow cytometry. Fresh neutrophils were isolated for functional analysis and ex-vivo assessment of cell-toxicity in a co-culture with human aortic endothelial cells (HAECs). Neutrophil secretion of active MMP9 was evaluated by fluorescence-based zymography in supernatants of isolated neutrophils and in patients' plasma samples. Results Neutrophils of patients with IFC-ACS show significantly higher expression of the toll-like receptor 2 (TLR2) in comparison to RFC-ACS-derived neutrophils (LOC: 1991±492.8 vs. 1615±440.2; p=0.01; SYS: 2062±464.4 vs. 1670±525.1; p=0.0056). Ex-vivo TLR2-stimulation of local neutrophils in patients with IFC-ACS led to increased toxicity of their secretome and aggravated endothelial cell death in co-culture, as compared to neutrophils from RFC-ACS patients (+59% dead HAECs, IFC-LOC vs. RFC-LOC; p=0.0078). Furthermore, TLR2-stimulation using Pam3CSK4 triggered higher activity rates of MMP9 exclusively in local neutrophils of IFC-ACS-patients (+38.9%±6.1% in IFC-LOC vs. RFC-LOC; p=0.0154). This effect was reversed in IFC-ACS-derived neutrophils being pre-treated with an anti-TLR2 neutralizing antibody (−58.4% ±5.2%, IFC-LOC-anti-TLR2 vs. IFC-LOC-vehicle; p=0.0069). Additionally, MMP9 activity was higher in plasma obtained from the culprit site of IFC-ACS patients (74.1 U/ml ±4.1 vs. 70.0 U/ml ±5.1, IFC-LOC vs. RFC-LOC; p=0.01). Conclusion The current study demonstrates novel TLR2-dependant neutrophil activation patterns at the coronary culprit lesion of IFC-ACS, leading to higher endothelial cell toxicity and MMP9 activity. Further studies need to assess whether a temporary blockade of TLR2 activation could be a possible therapeutic target in the era of personalized medicine. Funding Acknowledgement Type of funding source: Public Institution(s). Main funding source(s): Berlin Institute of Health (BIH), German Center for Cardiovascular Research
The evolutionary conserved NEAT1-MALAT1 gene cluster generates large noncoding transcripts remaining nuclear, while tRNA-like transcripts (mascRNA, menRNA) enzymatically generated from these precursors translocate to the cytosol. Whereas functions have been assigned to the nuclear transcripts, data on biological functions of the small cytosolic transcripts are sparse. We previously found NEAT1−/− and MALAT1−/− mice to display massive atherosclerosis and vascular inflammation. Here, employing selective targeted disruption of menRNA or mascRNA, we investigate the tRNA-like molecules as critical components of innate immunity. CRISPR-generated human ΔmascRNA and ΔmenRNA monocytes/macrophages display defective innate immune sensing, loss of cytokine control, imbalance of growth/angiogenic factor expression impacting upon angiogenesis, and altered cell–cell interaction systems. Antiviral response, foam cell formation/oxLDL uptake, and M1/M2 polarization are defective in ΔmascRNA/ΔmenRNA macrophages, defining first biological functions of menRNA and describing new functions of mascRNA. menRNA and mascRNA represent novel components of innate immunity arising from the noncoding genome. They appear as prototypes of a new class of noncoding RNAs distinct from others (miRNAs, siRNAs) by biosynthetic pathway and intracellular kinetics. Their NEAT1-MALAT1 region of origin appears as archetype of a functionally highly integrated RNA processing system.
Abstract Background Acute coronary syndromes (ACS) remain the most devastating clinical manifestation of cardiovascular disease, the most common cause of global mortality. ACS frequently arises from rupture of the coronary fibrous cap, in which a highly thrombogenic necrotic core is exposed to circulating blood, thereby triggering thrombus formation and impairing myocardial perfusion. While inflammation has been implicated as a key mechanism contributing to atherosclerotic plaque vulnerability and rupture, the underlying mechanisms leading to coronary plaque erosion, another form of ACS, are not well understood. In recent findings, flow cytometric (FACS) analysis of blood from the site of patients with culprit lesion plaque erosion showed significant enrichment of both CD4+ and CD8+ T-lymphocytes (+8.1% and +11.2%, respectively, both P<0.05) as well as effector molecules such as granzyme A (+22.4%), perforin (+58.8%), and granulysin (+75.4%) as compared with patients with plaque rupture culprit lesion. The proximity of eroded lesions to coronary bifurcations seen by optical coherence tomography provides further elucidations linking shear stress alterations to the occurrence of erosional ACS. Purpose We aim to further explore the underlying immune cell mechanisms of coronary plaque erosion via a novel single cell multi-omic approach, never before established from samples aspirated from the arterial sheath. Methods Patients presenting with Myocardial Infarction underwent emergent coronary angiography & percutaneous coronary intervention. Infarction was characterized as either coronary plaque rupture or erosion by optical coherence tomography, in which then blood samples were obtained by being aspirated directly along the arterial sheath. Peripheral blood mononuclear cells were isolated via density gradient within four hours of sample aspiration and cryopreserved. Samples from 24 patients were promptly thawed, washed and incubated with 137 oligonucleotide barcoded antibodies to detect surface protein expression as well as a cellular hashtag antibody. Bioinformatic integration of single cell epitope and transcriptome information was performed derived from custom pipelines. Results Our analysis found 32 unique leukocyte clusters that allowed us to rigorously compare each patients sequenced single cell data -totaling approximately 140,000 singe cells. Additionally, the discovery of T cell oligoclonality amongst certain disease patients alludes to antigen-specific activation. We also observed increased CCL4 expression in CD8+ T-lymphocytes within erosional ACS. Conclusion Our high-parametric immune cell analysis shows distinct differences in the adaptive immune system, particularly CD8+ T-lymphocytes and their effector molecules, in the pathogenesis of erosional versus ruptured ACS. Furthermore, our observations found on the oligoclonal expansion of T cell clones aids us to further elucidate underlying mechanisms of culprit lesion formation in ACS. Funding Acknowledgement Type of funding sources: Foundation. Main funding source(s): DGK - German Cardiac SocietyDGF - TRR 259 Aortic Disease German Research Foundation
