Improved integration of single-cell transcriptome data demonstrates common and unique signatures of heart failure in mice and humans.

BACKGROUND: Cardiovascular research heavily relies on mouse (Mus musculus) models to study disease mechanisms and to test novel biomarkers and medications. Yet, applying these results to patients remains a major challenge and often results in noneffective drugs. Therefore, it is an open challenge of translational science to develop models with high similarities and predictive value. This requires a comparison of disease models in mice with diseased tissue derived from humans. RESULTS: To compare the transcriptional signatures at single-cell resolution, we implemented an integration pipeline called OrthoIntegrate, which uniquely assigns orthologs and therewith merges single-cell RNA sequencing (scRNA-seq) RNA of different species. The pipeline has been designed to be as easy to use and is fully integrable in the standard Seurat workflow.We applied OrthoIntegrate on scRNA-seq from cardiac tissue of heart failure patients with reduced ejection fraction (HFrEF) and scRNA-seq from the mice after chronic infarction, which is a commonly used mouse model to mimic HFrEF. We discovered shared and distinct regulatory pathways between human HFrEF patients and the corresponding mouse model. Overall, 54% of genes were commonly regulated, including major changes in cardiomyocyte energy metabolism. However, several regulatory pathways (e.g., angiogenesis) were specifically regulated in humans. CONCLUSIONS: The demonstration of unique pathways occurring in humans indicates limitations on the comparability between mice models and human HFrEF and shows that results from the mice model should be validated carefully. OrthoIntegrate is publicly accessible (https://github.com/MarianoRuzJurado/OrthoIntegrate) and can be used to integrate other large datasets to provide a general comparison of models with patient data.

Angiotensin Receptor-Neprilysin Inhibition (Sacubitril/Valsartan) Reduces Structural Arterial Stiffness in Middle-Aged Mice.

BACKGROUND: Increasing arterial stiffness is a prominent feature of the aging cardiovascular system. Arterial stiffening leads to fundamental alterations in central hemodynamics with widespread detrimental implications for organ function resulting in significant morbidity and death, and specific therapies to address the underlying age-related structural arterial remodeling remain elusive. The present study investigates the potential of the recently clinically available dual angiotensin receptor-neprilysin inhibitor (ARNI) sacubitril/valsartan (LCZ696) to counteract age-related arterial fibrotic remodeling and stiffening in 1-year-old mice. METHODS AND RESULTS: Treatment of in 1-year-old mice with ARNI (sacubitril/valsartan), in contrast to angiotensin receptor blocker monotherapy (valsartan) and vehicle treatment (controls), significantly decreases structural aortic stiffness (as measured by in vivo pulse-wave velocity and ex vivo aortic pressure myography). This phenomenon appears, at least partly, independent of (indirect) blood pressure effects and may be related to a direct antifibrotic interference with aortic smooth muscle cell collagen production. Furthermore, we find aortic remodeling and destiffening due to ARNI treatment to be associated with improved parameters of cardiac diastolic function in aged mice. CONCLUSIONS: This study provides preclinical mechanistic evidence indicating that ARNI-based interventions may counteract age-related arterial stiffening and may therefore be further investigated as a promising strategy to improve cardiovascular outcomes in the elderly.

Correction: Hlavicka et al. Long-Term Outcomes after Aortic Valve and Root Replacement in a Very High-Risk Population. J. Cardiovasc. Dev. Dis. 2022, 9, 197.

The authors would like to make the following corrections to the published paper [...].

Right Axillary Artery Cannulation for Veno-Arterial Extracorporeal Membrane Oxygenation in Postcardiotomy Patients: A Single-Center Experience.

Background and Objectives: To analyze the patient outcome and complication rate of axillary artery cannulation for veno-arterial extracorporeal membrane oxygenation (VA-ECMO) in patients who could not be weaned from cardiopulmonary bypass after cardiothoracic surgery. Materials and Methods: We analyzed the data of 179 patients who were supported with VA-ECMO with femoral-axillary access (FA VA-ECMO) after cardiothoracic surgery between January 2014 and January 2019 in our department. Patients requiring central aortic cannulation and patients with respiratory failure requiring veno-venous ECMO were excluded. Primary outcomes were in-hospital mortality and 1-year survival rate of patients who were weaned from VA-ECMO support. Secondary outcomes were cannulation-related complications at the axillary site, VA-ECMO-related complications, and systemic complications. Results: In our cohort, 60 (33.5%) patients were female. Mean age was 67.0 ± 10.9 years. Overall, 78 (43.5%) patients were operated upon electively, 37 (20.7%) patients underwent urgent surgery, and 64 (35.8%) patients underwent emergency surgical treatment. Sixty-seven patients (37.4%) were resuscitated preoperatively. The mean duration of VA-ECMO support was 8.4 ± 5.1 days. Weaning from VA-ECMO was successful in 87 (48.6%) patients; 62 (34.6%) patients survived the hospital stay. The 1-year survival rate was 74%. Subclavian bleeding occurred in 24 (13.4%) patients, femoral bleeding in 4 (2.2%) patients, ischemia of the upper limb in 11 (6.1%) patients, intracerebral bleeding in 9 (5%) patients, and stroke in 19 (10.6%) patients. Conclusions: In patients with acute LV dysfunction after cardiothoracic surgery who cannot be weaned from cardiopulmonary bypass, right axillary artery cannulation is a safe and reliable method for VA-ECMO support with an acceptable complication rate.

Temporary Mechanical Circulatory Support in Cardiogenic Shock Patients after Cardiac Procedures: Selection Algorithm and Weaning Strategies.

Mechanical circulatory support has proven effective in managing postcardiotomy cardiogenic shock by stabilizing patients' hemodynamics and ensuring adequate organ perfusion. Among the available device modalities, the combination of extracorporeal life support and a microaxial flow pump for left ventricular unloading has emerged as a valuable tool in the surgical armamentarium. In this publication, we provide recommendations for the application and weaning of temporary mechanical circulatory support in cardiogenic shock patients, derived from a consensus among leading cardiac centers in German-speaking countries.

Single-nuclear transcriptome profiling identifies persistent fibroblast activation in hypertrophic and failing human hearts of patients with longstanding disease.

AIMS: Cardiac fibrosis drives the progression of heart failure in ischaemic and hypertrophic cardiomyopathy. Therefore, the development of specific anti-fibrotic treatment regimens to counteract cardiac fibrosis is of high clinical relevance. Hence, this study examined the presence of persistent fibroblast activation during longstanding human heart disease at a single-cell resolution to identify putative therapeutic targets to counteract pathological cardiac fibrosis in patients. METHODS AND RESULTS: We used single-nuclei RNA sequencing with human tissues from two samples of one healthy donor, and five hypertrophic and two failing hearts. Unsupervised sub-clustering of 7110 nuclei led to the identification of 7 distinct fibroblast clusters. De-convolution of cardiac fibroblast heterogeneity revealed a distinct population of human cardiac fibroblasts with a molecular signature of persistent fibroblast activation and a transcriptional switch towards a pro-fibrotic extra-cellular matrix composition in patients with established cardiac hypertrophy and heart failure. This sub-cluster was characterized by high expression of POSTN, RUNX1, CILP, and a target gene adipocyte enhancer-binding protein 1 (AEBP1) (all P < 0.001). Strikingly, elevated circulating AEBP1 blood level were also detected in a validation cohort of patients with confirmed cardiac fibrosis and hypertrophic cardiomyopathy by cardiac magnetic resonance imaging (P < 0.01). Since endogenous AEBP1 expression was increased in patients with established cardiac hypertrophy and heart failure, we assessed the functional consequence of siRNA-mediated AEBP1 silencing in human cardiac fibroblasts. Indeed, AEBP1 silencing reduced proliferation, migration, and fibroblast contractile capacity and α-SMA gene expression, which is a hallmark of fibroblast activation (all P < 0.05). Mechanistically, the anti-fibrotic effects of AEBP1 silencing were linked to transforming growth factor-beta pathway modulation. CONCLUSION: Together, this study identifies persistent fibroblast activation in patients with longstanding heart disease, which might be detected by circulating AEBP1 and therapeutically modulated by its targeted silencing in human cardiac fibroblasts.

Ablation of palladin in adult heart causes dilated cardiomyopathy associated with intercalated disc abnormalities.

Palladin (PALLD) belongs to the PALLD/myopalladin (MYPN)/myotilin family of actin-associated immunoglobulin-containing proteins in the sarcomeric Z-line. PALLD is ubiquitously expressed in several isoforms, and its longest 200 kDa isoform, predominantly expressed in striated muscle, shows high structural homology to MYPN. MYPN gene mutations are associated with human cardiomyopathies, whereas the role of PALLD in the heart has remained unknown, partly due to embryonic lethality of PALLD knockout mice. In a yeast two-hybrid screening, CARP/Ankrd1 and FHOD1 were identified as novel interaction partners of PALLD's N-terminal region. To study the role of PALLD in the heart, we generated conditional (cPKO) and inducible (cPKOi) cardiomyocyte-specific PALLD knockout mice. While cPKO mice exhibited no pathological phenotype, ablation of PALLD in adult cPKOi mice caused progressive cardiac dilation and systolic dysfunction, associated with reduced cardiomyocyte contractility, intercalated disc abnormalities, and fibrosis, demonstrating that PALLD is essential for normal cardiac function. Double cPKO and MYPN knockout (MKO) mice exhibited a similar phenotype as MKO mice, suggesting that MYPN does not compensate for the loss of PALLD in cPKO mice. Altered transcript levels of MYPN and PALLD isoforms were found in myocardial tissue from human dilated and ischemic cardiomyopathy patients, whereas their protein expression levels were unaltered.

Long-Term Outcomes after Aortic Valve and Root Replacement in a Very High-Risk Population.

Background: Aortic valve and root replacement (AVRR) is a standardised procedure to treat patients with aortic valve and root disease. In centres with a well-established aortic valve and root repair program (valve repairs and Ross operations), only patients with very complex conditions receive AVRR; this procedure uses a mechanical or biological composite valve graft (modified Bentall-de Bono procedure). The aim of the study was to evaluate the short- and long-term results after AVRR in a high-risk population with complex pathologies. Methods: Between 2005 and 2018, a total of 273 consecutive patients (mean age 64 ± 12.8 years; 23% female) received AVRR. The indication for surgery was an acute type A aortic dissection in 18%, infective endocarditis in 36% and other pathologies in 46% patients; 39% were redo procedures. The median EuroSCORE II was 11.65% (range 1.48-95.63%). Concomitant surgery was required in 157 patients (58%). Results: The follow-up extended to 5.2 years (range 0.1-15 years) and it was complete in 96% of the patients. The 30-day mortality was 17%. The overall estimated survival at 5 and 10 years was 65% ± 3% and 49% ± 4%, respectively. Univariate and multivariate logistic regression analyses revealed the following risk factors for survival: perioperative neurological dysfunction (OR 5.45), peripheral artery disease (OR 4.4) and re-exploration for bleeding (OR 3.37). Conclusions: AVRR can be performed with acceptable short- and long-term results in a sick patient population. The Bentall-De Bono procedure may be determined to be suitable for only elderly or high-risk patients. Any other patients should receive an AV repair or the Ross procedure in well-established centres.

Results from a multicentre evaluation of plug use for left ventricular assist device explantation.

OBJECTIVES: Myocardial recovery allows for left ventricular assist device (LVAD) explantations after long-term support. Several surgical approaches, including interventional decommissioning, off-pump explantation using a custom-made plug and complete LVAD removal through redo sternotomy, have been described. We present the results from an evaluation of the long-term follow-up of patients who received a titanium sintered plug after LVAD explantation. METHODS: We performed a retrospective, European, multicentre analysis of patients who received a titanium sintered plug to seal the apical fixation ring after LVAD explantation. Data were collected from a questionnaire that included demographics, procedural details and follow-up information. RESULTS: Out of 54 contacted centres in 12 countries (n = 179 patients), a total of 68 patients were successfully included in the study. The median follow-up was 34 months (interquartile range: 17-58.5 months); 57 (84%) patients had >1-year follow-up. At the time of the last follow-up, 55 (81%) patients were alive, with a Kaplan-Meier 1-year survival of 90.1% (95% confidence interval: 84.0-98.1%) and a 5-year survival of 80.0% (95% confidence interval: 68.4-92.9%). One patient (1.5%) developed a plug infection originating from an infected part of the incorporated driveline and, after complete removal, is currently in good condition. No postoperative stroke has been reported after plug implantation. CONCLUSIONS: In this European multicentre study, the use of a custom-made titanium plug to close the apical fixation ring after LVAD explantation resulted in a low incidence of plug-related complications. With the volume of patients undergoing LVAD explantations after myocardial recovery increasing, the plug has evolved as a simple alternative to more invasive device explantation procedures or decommissioning with a high risk for infection of the remaining system or stroke.

A human cell atlas of the pressure-induced hypertrophic heart.

Pathological cardiac hypertrophy is a leading cause of heart failure, but knowledge of the full repertoire of cardiac cells and their gene expression profiles in the human hypertrophic heart is missing. Here, by using large-scale single-nucleus transcriptomics, we present the transcriptional response of human cardiomyocytes to pressure overload caused by aortic valve stenosis and describe major alterations in cardiac cellular crosstalk. Hypertrophied cardiomyocytes had reduced input from endothelial cells and fibroblasts. Genes encoding Eph receptor tyrosine kinases, particularly EPHB1, were significantly downregulated in cardiomyocytes of the hypertrophied heart. Consequently, EPHB1 activation by its ligand ephrin (EFN)B2, which is mainly expressed by endothelial cells, was reduced. EFNB2 inhibited cardiomyocyte hypertrophy in vitro, while silencing its expression in endothelial cells induced hypertrophy in co-cultured cardiomyocytes. Our human cell atlas of the hypertrophied heart highlights the importance of intercellular crosstalk in disease pathogenesis and provides a valuable resource.

The potential role of plasma miR-155 and miR-206 as circulatory biomarkers in inflammatory cardiomyopathy.

AIMS: Establishing a diagnosis of inflammatory cardiomyopathy (iCMP) by non-invasive means remains challenging despite advances in cardiac magnetic resonance imaging. Previous studies suggested the involvement of microRNAs in the pathogenesis of iCMP. We examined the association of a predefined set of circulatory microRNAs with clinical characteristics of iCMP and evaluated their diagnostic performance in suspected iCMP. METHODS AND RESULTS: Eighty-nine patients with clinical suspicion of iCMP were included in the analysis. All patients underwent cardiac catheterization with left ventricular endomyocardial biopsy, echocardiography, and cardiac magnetic resonance imaging applying the Lake Louise criteria (LLC). Plasma levels of miR-21, miR-126, miR-133a, miR-146b, miR-155, and miR-206 were determined using real-time polymerase chain reaction. Based on immunohistological findings on endomyocardial biopsy, iCMP was diagnosed in 67% of study participants (n = 60). Plasma levels of miR-155 and miR-206 were significantly increased in patients with iCMP as compared with patients with dilated cardiomyopathy (P = 0.008 and P = 0.009, respectively). In receiver operating characteristic curve analysis, miR-155 and miR-206 demonstrated superior diagnostic performance for iCMP (0.68 and 0.67, respectively) compared with LLC [area under the curve (AUC) 0.60], Troponin T (AUC 0.51), and N-terminal pro-brain natriuretic peptide (AUC 0.51). While baseline miR-155 and miR-206 plasma levels were predictive for biopsy-proven iCMP (odds ratio = 2.61, 95% confidence interval = 1.28-5.31, P = 0.008 and odds ratio = 2.65, 95% confidence interval = 1.27-5.52, P = 0.009) on univariate logistic regression analysis, the presence of positive LLC, high baseline C-reactive protein, or presence of clinical symptoms and signs of viral infection failed to predict iCMP (P > 0.05, respectively). CONCLUSIONS: The present data suggest that plasma levels of miR-206 and miR-155 are potential novel biomarkers for confirming the diagnosis of iCMP.

MicroRNA miR-29b regulates diabetic aortic remodeling and stiffening.

Patients with type 2 diabetes (T2D) are threatened by excessive cardiovascular morbidity and mortality. While accelerated arterial stiffening may represent a critical mechanistic factor driving cardiovascular risk in T2D, specific therapies to contain the underlying diabetic arterial remodeling have been elusive. The present translational study investigates the role of microRNA-29b (miR-29b) as a driver and therapeutic target of diabetic aortic remodeling and stiffening. Using a murine model (db/db mice), as well as human aortic tissue samples, we find that diabetic aortic remodeling and stiffening is associated with medial fibrosis, as well as fragmentation of aortic elastic layers. miR-29b is significantly downregulated in T2D and miR-29b repression is sufficient to induce both aortic medial fibrosis and elastin breakdown through upregulation of its direct target genes COL1A1 and MMP2 thereby increasing aortic stiffness. Moreover, antioxidant treatment restores aortic miR-29b levels and counteracts diabetic aortic remodeling. Concluding, we identify miR-29b as a comprehensive-and therefore powerful-regulator of aortic remodeling and stiffening in T2D that moreover qualifies as a (redox-sensitive) target for therapeutic intervention.

The Ubiquitin Proteasome System in Ischemic and Dilated Cardiomyopathy.

Dilated (DCM) and ischemic cardiomyopathies (ICM) are associated with cardiac remodeling, where the ubiquitin-proteasome system (UPS) holds a central role. Little is known about the UPS and its alterations in patients suffering from DCM or ICM. The aim of this study is to characterize the UPS activity in human heart tissue from cardiomyopathy patients. Myocardial tissue from ICM (n = 23), DCM (n = 28), and control (n = 14) patients were used to quantify ubiquitinylated proteins, E3-ubiquitin-ligases muscle-atrophy-F-box (MAFbx)/atrogin-1, muscle-RING-finger-1 (MuRF1), and eukaryotic-translation-initiation-factor-4E (eIF4E), by Western blot. Furthermore, the proteasomal chymotrypsin-like and trypsin-like peptidase activities were determined fluorometrically. Enzyme activity of NAD(P)H oxidase was assessed as an index of reactive oxygen species production. The chymotrypsin- (p = 0.71) and caspase-like proteasomal activity (p = 0.93) was similar between the groups. Trypsin-like proteasomal activity was lower in ICM (0.78 ± 0.11 µU/mg) compared to DCM (1.06 ± 0.08 µU/mg) and control (1.00 ± 0.06 µU/mg; p = 0.06) samples. Decreased ubiquitin expression in both cardiomyopathy groups (ICM vs. control: p < 0.001; DCM vs. control: p < 0.001), as well as less ubiquitin-positive deposits in ICM-damaged tissue (ICM: 4.19% ± 0.60%, control: 6.28% ± 0.40%, p = 0.022), were detected. E3-ligase MuRF1 protein expression (p = 0.62), NADPH-oxidase activity (p = 0.63), and AIF-positive cells (p = 0.50). Statistical trends were detected for reduced MAFbx protein expression in the DCM-group (p = 0.07). Different levels of UPS components, E3 ligases, and UPS activation markers were observed in myocardial tissue from patients affected by DCM and ICM, suggesting differential involvement of the UPS in the underlying pathologies.

Anatomically specific reactive oxygen species production participates in Marfan syndrome aneurysm formation.

Marfan syndrome (MFS) is a connective tissue disorder that results in aortic root aneurysm formation. Reactive oxygen species (ROS) seem to play a role in aortic wall remodelling in MFS, although the mechanism remains unknown. MFS Fbn1C1039G/+ mouse root/ascending (AS) and descending (DES) aortic samples were examined using DHE staining, lucigenin-enhanced chemiluminescence (LGCL), Verhoeff's elastin-Van Gieson staining (elastin breakdown) and in situ zymography for protease activity. Fbn1C1039G/+ AS- or DES-derived smooth muscle cells (SMC) were treated with anti-TGF-ß antibody, angiotensin II (AngII), anti-TGF-ß antibody + AngII, or isotype control. ROS were detected during early aneurysm formation in the Fbn1C1039G/+ AS aorta, but absent in normal-sized DES aorta. Fbn1C1039G/+ mice treated with the unspecific NADPH oxidase inhibitor, apocynin reduced AS aneurysm formation, with attenuated elastin fragmentation. In situ zymography revealed apocynin treatment decreased protease activity. In vitro SMC studies showed Fbn1C1039G/+ -derived AS SMC had increased NADPH activity compared to DES-derived SMC. AS SMC NADPH activity increased with AngII treatment and appeared TGF-ß dependent. In conclusion, ROS play a role in MFS aneurysm development and correspond anatomically with aneurysmal aortic segments. ROS inhibition via apocynin treatment attenuates MFS aneurysm progression. AngII enhances ROS production in MFS AS SMCs and is likely TGF-ß dependent.

Ischemic Cardiomyopathy Affects the Thioredoxin System in the Human Myocardium.

BACKGROUND: Oxidative stress due to reactive oxygen species (ROS) production is a key factor in the development of heart failure (HF). This study investigated the thioredoxin (Trx) system, which plays a major role in antioxidant defense, in patients suffering from ischemic (ICM) or dilated (DCM) cardiomyopathy. METHODS AND RESULTS: Myocardial tissue from ICM (n = 13) and DCM (n = 13) patients, as well as septal tissue of patients with aortic stenosis but without diagnosed hypertrophic cardiomyopathy or subaortic stenosis (control; n = 12), was analyzed for Trx1, Trx-interacting protein (TXNIP) and E3 ligase ITCH (E3 ubiquitin-protein ligase Itchy homolog) expression. Trx-reductase 1 (TXNRD1) amount and activity, cytosolic cytochrome C content, and apoptosis markers were quantified by means of enzyme-linked immunosorbent assay and multiplexing. Compared with control samples, ITCH and Trx1 expression, TXNRD1 amount and activity were reduced and TXNIP expression was increased in ICM (ITCH: P = .013; Trx1: P = .028; TXNRD1 amount: P = .035; TXNRD1 activity: P = .005; TXNIP: P = .014) but not in DCM samples. A higher level of the downstream apoptosis marker caspase-9 (ICM: 582 ± 262 MFI [P = .995]; DCM: 1251 ± 548 MFI [P = .002], control: 561 ± 214 MFI) was detected in DCM tissue. A higher expression of Bcl-2 was found in DCM (P = .011). CONCLUSION: The Trx system was impaired in ICM but not in DCM. ITCH appeared to be responsible for the down-regulation of the Trx system. ROS-induced mitochondrial instability appeared to play a role in DCM.

Acute Effect of Mitral Valve Repair on Mitral Valve Geometry.

BACKGROUND: The aim of this study was to quantify acute mitral valve (MV) geometry dynamic changes throughout the cardiac cycle using three-dimensional transesophageal echocardiography (3D TEE) in patients undergoing surgical MV repair (MVR) with ring annuloplasty and optional neochord implantation. METHODS: Twenty-nine patients (63 ± 10 years) with severe primary mitral regurgitation underwent surgical MVR using ring annuloplasty with or without neochord implantation. We recorded 3D TEE data throughout the cardiac cycle before and after MVR. Dynamic changes (4D) in the MV annulus geometry and anatomical MV orifice area (AMVOA) were measured using a novel semiautomated software (Auto Valve, Siemens Healthcare). RESULTS: MVR significantly reduces the anteroposterior diameter by up to 38% at end-systole (36.8-22.7 mm; p < 0.001) and the lateromedial diameter by up to 31% (42.7-30.3 mm; p < 0.001). Moreover, the annular circumference was reduced by up to 31% at end-systole (129.6-87.6 mm, p < 0.001), and the annular area was significantly decreased by up to 52% (12.8-5.7 cm2; p < 0.001). Finally, the AMVOA experienced the largest change, decreasing from 1.1 to 0.2 cm2 during systole (at midsystole; p < 0.001) and from 4.1 to 3.2 cm2 (p < 0.001) during diastole. CONCLUSIONS: MVR reduces the annular dimension and the AMVOA, contributing to mitral competency, but the use of annuloplasty rings reduces annular contractility after the procedure. Surgeons can use 4D imaging technology to assess MV function dynamically, detecting the acute morphological changes of the mitral annulus and leaflets before and after the procedure.

Changes in dynamic mitral valve geometry during percutaneous edge-edge mitral valve repair with the MitraClip system.

BACKGROUND: The aim of this study was to quantify the acute dynamic changes of mitral valve (MV) geometry throughout the cardiac cycle-during percutaneous MV repair with the MitraClip system by 3-dimensional transesophageal echocardiography (3D TEE). METHODS: The MV was imaged throughout the cardiac cycle (CC) before and after the MitraClip procedure using 3D TEE in 28 patients (mean age, 77 ± 8 years) with functional mitral regurgitation (FMR). Dynamic changes in the MV annulus geometry and anatomical MV orifice area (AMVOA) were quantified using a novel semi-automated software. RESULTS: Percutaneous MV repair decreased anterior-posterior diameter by up to 9% (at 50% of CC; from 34.5 to 31.9 mm; p < 0.001) throughout the CC and increased the diastolic lateral-medial diameter by up to 7% (at 60% of the CC; from 39.7 to 42.3 mm; p < 0.001), whereas the annular circumference and area were not significantly affected. Annulus sphericity index was reduced up to 13% (at 50% of the CC; from 0.89 to 0.78, p < 0.001). The AMVOA also decreased during systole, the maximum decrease being from 0.6 to 0.2 mm2 (at 0% of CC; p = 0.007), and during diastole the maximum decrease being from 4.6 to 1.6 cm2 (at 50% of CC; p < 0.001). CONCLUSIONS: Percutaneous MV repair reduces the MR by an improved coaptation of MV leaflets joint with a simultaneous indirect reduction of anterior-posterior diameter. Further, the MitraClip procedure leads to a reduction of AMVOA of more than 60% during diastole.

Gene expression profiling of acute type A aortic dissection combined with in vitro assessment.

OBJECTIVES: The mechanisms underlying aortic dissection remain to be fully elucidated. We aimed to identify key molecules driving dissection through gene expression profiling achieved by microarray analysis and subsequent in vitro experiments using human aortic endothelial cells (HAECs) and aortic vascular smooth muscle cells (AoSMCs). METHODS: Total RNA, including microRNA (miRNA), was isolated from the intima-media layer of dissected ascending aorta obtained intraoperatively from acute type A aortic dissection (ATAAD) patients without familial thoracic aortic disease (n = 8) and that of non-dissected ascending aorta obtained from transplant donors (n = 9). Gene expression profiling was performed with mRNA and miRNA microarrays, and results were confirmed by quantitative polymerase chain reaction (qPCR). Target genes and miRNA were identified by gene ontology analysis and a literature search. To reproduce the in silico results, HAECs and AoSMCs were stimulated in vitro by upstream cytokines, and expression of target genes was assessed by qPCR. RESULTS: Microarray analysis revealed 1536 genes (3.6%, 1536/42 545 probes) and 41 miRNAs (3.0%, 41/1368 probes) that were differentially expressed in the ATAAD group (versus donor group). The top 15 related pathways included regulation of inflammatory response, growth factor activity and extracellular matrix. Gene ontology analysis identified JAK2 (regulation of inflammatory response), PDGFA, TGFB1, VEGFA (growth factor activity) and TIMP3, TIMP4, SERPINE1 (extracellular matrix) as the target genes and miR-21-5p, a TIMP3 repressor, as target miRNA that interacts with the target genes. Validation qPCR confirmed the altered expression of all 7 target genes and miR-21-5p in dissected aorta specimens (all genes, P < 0.05). Ingenuity pathway analysis showed TNF-α and TGF-ß to be upstream cytokines for the target genes. In vitro experiments showed these cytokines inhibit TIMP3 expression (P < 0.05) and enhance VEGFA expression (P < 0.01) in AoSMCs but not HAECs. miR-21-5p expression increases in AoSMCs under TNF-α and TGF-ß stimulation (fold change: 1.36; P = 0.011). CONCLUSIONS: Results of our novel approach, integrating in vitro assessment into gene expression profiling, implicated chronic inflammation characterized by MMP-TIMP dysregulation, increased VEGFA expression, and TGF-ß signalling in the development of dissection. Further investigation may reveal novel diagnostic biomarkers and uncover the mechanism(s) underlying ATAAD.