For the past 40 years, the methodology for stress echocardiography (SE) has remained basically unchanged. It is based on two-dimensional, black and white imaging, and is used to detect regional wall motion abnormalities (RWMA) in patients with known or suspected coronary artery disease (CAD). In the last five years much has changed and RWMA is not enough on its own to stratify patient risk and dictate therapy. Patients arriving at SE labs often have comorbidities and are undergoing full anti-ischemic therapy. The SE positivity rate based on RWMA fell from 70% in the eighties to 10% in the last decade. The understanding of CAD pathophysiology has shifted from a regional hydraulic disease to a systemic biologic disease. The conventional view of CAD encouraged the use of coronary anatomic imaging for diagnosis and the oculo-stenotic reflex for the deployment of therapy. This has led to a clinical oversimplification that ignores the lessons of pathophysiology and epidemiology, and in fact, CAD is not synonymous with ischemic heart disease. Patients with CAD may also have other vulnerabilities such as coronary plaque (step A of ABCDE-SE), alveolar-capillary membrane and pulmonary congestion (step B), preload and contractile reserve (step C), coronary microcirculation (step D) and cardiac autonomic balance (step E). The SE methodology based on two-dimensional echocardiography is now integrated with lung ultrasound (step B for B-lines), volumetric echocardiography (step C), color- and pulsed-wave Doppler (step D) and non-imaging electrocardiogram-based heart rate assessment (step E). In addition, qualitative assessment based on the naked eye has now become more quantitative, has been improved by contrast and based on cardiac strain and artificial intelligence. ABCDE-SE is now ready for large scale multicenter testing in the SE2030 study. Stress echocardiography (SE) is based on the assessment of inducible regional wall motion abnormalities (RWMA) for the functional evaluation of patients with known or suspected coronary artery disease (CAD). It has had a well-established clinical role for several decades  and is nSome aspects of the ABCDE protocol have already been discussed in previous review papers . HoweverIn the novel state-of-the-art ABCDE SE protocol, RWMA remains the first and main step A, which is corroborated by quantitative deformation imaging of global and regional strain. B-lines are assessed in step B, the left ventricular contractile and preload reserve in step C, Doppler-based coronary flow velocity reserve (CFVR) in the left anterior descending coronary artery or contrast myocardial perfusion imaging is assessed in step D, and ECG-based assessment of heart rate reserve in non-imaging takes place in step E. The five parameters converge conceptually, logistically and methodologically in the ABCDE protocol, which can be applied to all stresses and all patients and it allows a comprehensive color-coded risk stratification of the vulnerable patient beyond coronary artery stenosis  in the mid-distal left anterior descending coronary artery . CFVR isStep E is based on ECG, which evaluates the heart rate reserve as the peak/rest value of the heart rate derived from a 12-lead ECG or simply from the ECG lead that is automatically read in the echo monitor. It is an index of cardiac sympathetic reserve, is obtainable in all patients with 100% success rate and reproducibility. It is an index of cardiac autonomic function, and in particular, of cardiac sympathetic reserve, which can be impaired by several conditions other that CAD such as heart failure or hypertrophic cardiomyopathy . It is aMuch has changed as a result of this methodological and conceptual remodeling. The pathophysiological model has shifted from stenosis vulnerability to patient vulnerability. The prediction potential has shifted from the recurrence of angina to all events including cardiac death, non-cardiac death, arrhythmic events and progression to heart failure. Risk stratification has become multi-dimensional and color-coded , and thuNot all the problems of cardiology are solved with the ABCDE protocol. Yet, it is a simple way to capture a more comprehensive view than was possible in previous decades of the clinical use of SE. Its application is more suitable for risk stratification and functional characterization than for primary diagnosis of CAD. In general, the presence of multiple positive biomarkers points to more extensive anatomic CAD in patients with chronic coronary artery disease, but in principle, each of the four new markers can be present due to a variety of causes that are independent of the presence and severity of underlying CAD.Not all questions can be answered with the use of this protocol. However, the improvement in clinically relevant information in the last five years did not come from a technological upgrade or industrial innovation, but rather from a more integrated pathophysiological approach to the complexity of the cardiac patient. Strain is useful, contrast helps, and three-dimensional imaging is the unavoidable future, however, the conceptual and methodological framework of ABCDE can help to exploit these technological innovations in a more clinically-oriented way.More data and information are required for clinical use. In particular, we need to know whether each letter or level of the protocol predicts specific, individual endpoints, for instance, acute decompensated heart failure with step B or cardiac arrhythmias and sudden death with step E. To achieve this critical mass of data, large scale effectiveness studies are needed and are currently under way with the SE 2020, and the SE 2030 study which is about to start.Last but not least, the application of the new ABCDE protocol should always start from a clinical question or suspicion. The pathophysiological processes that underlie the protocol are always at work, but not all of them are always relevant for the treatment or interpretation of symptoms in the individual patient. Whether the steps in the protocol are essential or not may well depend on the different type of patient and the scope of testing that is required, including the primary diagnosis of CAD, identification of functional mechanisms of disease and symptoms, risk stratification, guide to therapy, or objective assessment of therapy efficacy.Apart from the limitations of the protocol as a whole, there are specific pitfalls in each step that should be considered, and which can possibly be solved by technological and cultural advances in the near future.Step A. SE has several obvious disadvantages. The three main disadvantages are patient-related , operator-related (depends on the reader’s expertise), and physiology-related . The inability to establish a good acoustic window precludes testing in a limited proportion of patients, but with the use of contrast and last-generation technology this percentage shrinks to <1%, even with the current morbid obesity pandemic. The subjective reading of RWMA by the naked eye requires training and does not allow improvisation. The training requirement for professionals in this field has been established as at least 100 studies per year with an expert supervisor to gain certification, and 100 studies per year to maintain accreditation.Step B. Lung rockets reflect the presence of an interstitial syndrome, which can be due to water, inflammation or fibrosis. B-lines must not be mixed up with Z-lines, which are frequently observed as bundle-shaped reflections arising from the pleural line, but unlike true B-lines, they do not erase A-lines, are ill-defined, less echogenic than the pleural line, are short and do not move in synchrony with respiration. The training requirement for professionals are easier than for RWMA and have been established as <50 studies per year with an expert supervisor to gain certification.Step C. The image quality required for consistent quantitative measurements of LV volumes is higher than that needed for RWMA. The delineation of LV volumes can be time-consuming on unenhanced images, and calculations are faster and more repeatable with contrast-enhancement or artificial intelligence-based software, which is now on board commercially available machines. The professional training requirements are the same as for RWMA, but the measurements of LV volumes may lack repeatability when images are of sub-optimal quality.Step D. The measurement of CFVR in LAD as a proxy for global coronary flow reserve is attractive for its simplicity but it has conceptual and practical limitations. The variations in flow velocity are proportional to the total blood flow if the diameter of the vessel lumen is kept constant. In reality, the diameter of epicardial coronary arteries increases by an average of 30% in healthy subjects following adenosine infusion Therefore, failure to take into account epicardial coronary artery vasodilation during hyperemia may cause a nonsystematic underestimation of coronary flow reserve, which can be more accurately calculated by the velocity-time integral and the cross-sectional area. CFVR assessment in the right coronary artery and left circumflex is also feasible but imaging time is too long and the technical difficulties are greater, and therefore, the clinical assessment of CFVR is restricted to one single coronary district even though CAD is a regional and multi-district disease. The feasibility of LAD flow imaging is extremely high, although 5% of patients remain unreadable even after contrast enhancement. The professional training requirements are the same as for RWMA, but the interpretation is much more quantitative and easier. Coronary flow is more difficult to obtain but easier to interpret than RWMA.Step E. If atropine is added to dobutamine or dipyridamole, the cutoff values may be higher and are not validated. During pharmacological testing, the value of heart rate reserve is not affected by physical conditioning and deconditioning, which affects the value of chronotropic incompetence during exercise. The peak heart rate cannot be recorded at a fixed time corresponding to the end of adenosine or dipyridamole infusion, since the greatest change in heart rate (measured each minute) occurs in the time interval from 0 to 5 min after drug infusion. It is important to record the peak value since there is minimal interpatient variability, whereas the peak value always corresponds with peak stress during exercise. Beta-blockers reduce the absolute rest and peak values of heart rate, but the relative rest-stress changes mirrored in heart rate reserve are essentially the same. No specific training is required because the heart rate is given in real time and automatically by any machine.As with any new technique, there is a lack of direct comparative data with other imaging techniques in different clinical CAD scenarios. If the ABCDE replaces the conventional SE protocol as the new standard, this gap could be filled quickly. In line with this principle, some of the ABCDE steps can be incorporated in other non-ultrasound cardiac functional stress testing techniques. Step E might include perfusion data from radionuclide myocardial perfusion imaging, which can also estimate LV volumes in step C, although with less temporal and spatial resolution. Cardiac magnetic resonance is ideally suited to obtain information for step C with LV volumetric imaging and for step D with myocardial perfusion imaging. Cardiac functional testing is an alternative approach to anatomic imaging with noninvasive cardiac computed tomography angiography, that is recommended in current guidelines for patients with low pre-test probability of disease. However, this approach may change when sustainability considerations are incorporated into decision-making, as a more comprehensive, less stenosis-centered approach gains popularity in the field of cardiac imaging and in line with the pathophysiological lessons learned from COURAGE, ORBITA and ISCHEMIA trials.The growth of SE in recent years is a result of the dramatically changed cultural milieu surrounding health care delivery. The concept of sustainability has emerged as a key feature in shaping the all health care practices including cardiac imaging . SustainThe second component is the direct cost. SE is considerably less expensive than other alternative imaging techniques, and economically, a test with similar accuracy and higher cost would not be considered a viable option, although the situation is different in health care in that those who buy usually do not pay. With the changing economic climate, there is less societal and political tolerance for the delivery of high cost and low value health care.The third component is safety. SE has an excellent safety profile since stress can be interrupted as soon as RWMA appear during real-time monitoring, contrast has excellent tolerability and non-ionizing radiation is used. Prioritizing safety is also important to minimize the legal concerns connected with iodine contrast nephrotoxicity. Zero radiation is essential to mitigate the cancer burden due to cumulative radiation exposure, and in Europe the law reinforces the use of radiation-free options when available .The fourth component is the environmental footprint due to carbon dioxide emissions. Echocardiography produces 2 or 3 kg of carbon dioxide emissions per examination, which is 100-fold lower than magnetic resonance cardiac imaging and is by far the lowest of all the alternative techniques . The carThe concept of sustainability is making its way into health care, and cardiac imaging is progressively affected by sustainability issues. Small individual costs, risks, and ecological footprints multiplied by billions of examinations worldwide become a significant societal cost, population risk and environmental burden. There are four stakeholders in the imaging field: the patient, the physician, the payer and the planet . SE is gThe life of SE began 40 years ago and now life begins again at forty. We need effectiveness studies that are based on real patients, real instruments and real doctors to obtain true and realistic results on the performance of new tests or technologies ,77,78,79ABCDE-SE was the result of the SE2020 study  and is tThe new core ABCDE protocol  will be


Cardiovascular magnetic resonance imaging (CMR) have described localised non-ischemic late gadolinium enhancement (LGE) lesions of prognostic importance in various non-ischemic cardiomyopathies. Ischemic LGE lesions are prevalent in diabetes (DM), but non-ischemic LGE lesions have not previously been described or systematically studied in DM.1-mapping and LGE.296 patients with type 2 DM (T2DM) and 25 sex-matched control subjects underwent echocardiography and CMR including adenosine-stress perfusion, T264 patients and all control subjects completed the CMR protocol. 78.4% of patients with T2DM had no LGE lesions; 11.0% had ischemic LGE lesions only; 9.5% had non-ischemic LGE lesions only; and 1.1% had both one ischemic and one non-ischemic lesion. The non-ischemic LGE lesions were situated mid-myocardial in the basal lateral or the basal inferolateral part of the left ventricle and the affected segments showed normal to high wall thickness and normal contraction. Patients with non-ischemic LGE lesions in comparison with patients without LGE lesions had increased myocardial mass , average E/e’, left atrial maximal volume , NT-proBNP  and high-sensitive troponin  and a higher prevalence of retinopathy  and autonomic neuropathy .A specific LGE pattern with lesions in the basal lateral or the basal inferolateral part of the left ventricle was found in patients with type 2 diabetes.Trial registrationhttps://www.clinicaltrials.gov. Unique identifier: NCT02684331. For decades it has been known that angiopathy, as seen in patients with diabetes (DM) differs from what is seen in other patients , and theIn DM, even without significant coronary atheromatosis, altered cellular metabolism and hyperglycaemia result in oxidative stress, microvascular angiopathy, and diffuse interstitial myocardial fibrosis . With ecIt is important to determine if and to what degree non-ischemic LGE lesions exist in patients with DM and to understand their relations to not only risk factors but also variables of known importance for cardiac function and prognosis. In a larger cross-sectional CMR study of patients with type 2 DM (T2DM), we therefore systematically characterized all LGE lesions and their associations with myocardial perfusion, myocardial structure, signs of diastolic dysfunction, and association to common complications of DM.2, claustrophobia or other contraindications to CMR. Sex-matched normal subjects served as a control group, all without T2DM (HbA1c < 40 mmol/mol). Antihypertensive and statin therapy were allowed in the control group.T2DM patients were recruited from the Department of Endocrinology at Næstved, Ringsted, and Slagelse Hospitals in Denmark. After written informed consent, patients were included between January 2016 and August 2019. The study complies with the Declaration of Helsinki and was approved by The Zealand Ethics Committee (SJ-490). T2DM patients aged 18–80 years and diagnosed with T2DM for at least three months prior to inclusion were eligible for inclusion. Exclusion criteria were persistent or permanent atrial fibrillation at the time of echo or CMR, eGFR < 30 mL/min/1.73 mPatients underwent full medical history, assessment of blood pressure, electrocardiogram (ECG), and echocardiography. Blood and urinary samples were collected prior to the CMR scan, which in general was performed within 14 days of other assessments. History of retinopathy (latest fundoscopy), peripheral neuropathy (latest chiropodist report), ischemic heart disease, hypertension BP > 140/90 mmHg), and medication were obtained from the patient journal. An ECG obtained during five cycles of in- and expiration was performed to allow for beat-to-beat variation being analysed. Orthostatic blood pressure measurements were obtained ½, 1½, 3, 5, and 7 min after acutely standing up 0 mmHg, a. AlbuminStandard 2D echocardiography was performed using a Vivid S5 probe on a GE healthcare Vivid E9 cardiovascular ultrasound system. The GE EchoPAC workstation was used for analysis. Peak early mitral inflow (E) was measured in the 4-chamber view using pulsed-wave Doppler with the sample volume placed between the tips of the mitral valve leaflets. Also, in the 4-chamber view, the early diastolic myocardial velocity (e′) was measured using pulsed-wave tissue velocity Doppler with the sample volume placed in the septal and lateral mitral annulus.1-mapping were performed as previously described [CMR studies were performed on a 1.5 T scanner  with patients in a supine position on the back using a surface- and spine cardiac coil with ECG gating. Cardiac short-axis steady-state free precession cine sequences were acquired during end-expiratory breath-holds in 25 phases  and in 2-, 3- and 4-chamber views. Sequence settings were TE 1.16–1.25 ms, TR 46.24–49.98 ms, Matrix 210 × 208, FoV 258 × 320–485 × 481. Myocardial perfusion images were obtained at the basal, mid-ventricular and apical cardiac short-axis level during rest and pharmacological stress (adenosine 140 µg/kg/min) using an iv. dose of 0.075 mmol/kg of gadobutrol  for each sequence as previously described . LGE imaescribed  . LV volumes and mass were determined by semi-automatic tracing of the endo- and epicardial contours in end-diastolic and end-systolic phases. Papillary muscles were excluded. LA maximum volume was measured by manual tracing in the LV end-systolic frame. For all CMR parameters we chose to report both the absolute values and indexed to body surface area because the majority of our patients were severe obese, which could bias the indexed results. A LGE lesion was considered to be present if seen in at least two perpendicular slices. Blinded to each other, two observers analysed all LGE images (AB and MS). Blinded to clinical data, the analyses were reviewed and finalized by a third observer with 10 years of CMR experience (PLvM). A LGE lesion was classified as ischemic if subendocardial or transmural and respecting the perfusion territory of a coronary artery [−1. On native and post-contrast T1-maps endocardial and epicardial borders were traced semi-automatically, and the average ECV was calculated from areas outside LGE lesions [CMR images were analysed using cvi42y artery , otherwi lesions . For det lesions .v. 7.15 .For association analysis, patients with both ischemic and non-ischemic LGE lesions were included in the group with ischemic lesions. Quantitative variables were analysed for normal distribution and are presented as mean and standard deviation (SD) or median and interquartile range (IQR) as appropriate. Categorical variables are presented as absolute values and percentages. A one-way analysis of variance (ANOVA) was applied for the comparison of continuous variables with a normal distribution. If an unequal variance was seen either visually or by a significant Levene’s test, Welch’s ANOVA was applied. Non-normal distributed continuous variables were compared using a Kruskal–Wallis test. Categorical variables were compared using a Chi-squared test. If an ANOVA was significant additional analyses were performed with the comparison of DM2 patients without LGE lesions to patients with non-ischemic LGE lesions using either the Student’s T-test, the Mann–Whitney U test, Fisher’s exact test or a Chi-squared test as appropriate. For all tests, a two-tailed P-value < 0.05 was considered statistically significant. All calculations were made in SAS Enterprise Guide 296 T2DM patients and 25 sex-matched controls were included. Twenty-five T2DM patients did not complete the CMR scan, and in 6 patients the scan was performed without gadolinium contrast. In one patient, the LGE images could not be analysed because of poor image quality leaving 264 T2DM patients for final analysis. None of the 25 control subjects had any LGE lesions. In patients with T2DM, 207 (78.4%) did not have LGE lesions; 29 (11.0%) had ischemic LGE lesions only; 25 (9.5%) had non-ischemic LGE lesions only, and 3 (1.1%) patients had both an ischemic and a non-ischemic lesion. Six T2DM patients (2.3%) with ischemic LGE lesions had no prior history of ischemic heart disease. None of the patients without LGE lesions or with non-ischemic lesions had any hypoperfused areas on the adenosine stress perfusion scan indicative of significant coronary artery disease.1 mapping slice passed through the LGE lesion in 11 patients with non-ischemic and in 18 patients with ischemic LGE lesions: Without difference between non-affected remote myocardium (31.4 (IQR29.0–33.9) % vs. 33.0 (IQR31.5–35.8) %, P = 0.11), the median ECV value for a non-ischemic lesion was lower than what was seen in an ischemic lesion .In all 28 patients with non-ischemic LGE lesions, these were located mid-myocardial in the basal lateral or the basal inferolateral part of the LV myocardium Figs. , 2. Two Patient demographics including clinical characteristics and laboratory values are presented in Table The cardiac structural and functional characteristics of the four groups are presented in Table Association of DM complications between the three groups of T2DM patients is shown in Table In a cross-sectional magnetic resonance imaging study on diabetic heart disease, 10.6%  of a 264Previous LGE CMR studies in T2DM populations have focused on ischemic heart disease with ischemic LGE lesions from myocardial infarctions –16. Of iPatients with non-ischemic LGE lesions had increased ECV, which has previously been shown to correlate well with the levels of diffuse interstitial myocardial fibrosis . Thus, oNon-ischemic LGE lesions are well-known in patients with other non-ischemic cardiomyopathies such as hypertrophic cardiomyopathy, idiopathic dilated cardiomyopathy, myocarditis, Chagas disease, sarcoidosis, Anderson-Fabry or cardiac amyloidosis . In all In other cardiomyopathies, the finding of non-ischemic LGE lesions is associated with prognosis, and if such is the case in T2DM this now has to be established. Non-ischemic LGE lesions have in other patient groups been correlated to heart failure, arrhythmias, and sudden cardiac death , 29, 30.2) since we used gadolinium contrast for the CMR scans. An overlap exists between diabetic nephropathy and albuminuria and having excluded this patient group we may have missed an association between non-ischemic LGE lesions and albuminuria. Further, we have not performed biopsies, and thus we can only speculate on the aetiology of the lesions based on statistical associations. In this study we did not perform high resolution LGE or feature tracking, both newer and emerging techniques that could possibly have helped phenotyping our cohort even more [Our work have some limitations, first the study was cross-sectional and we can only infer causality and speculate on the causes of and consequences of having non-ischemic LGE lesion from the result on other patient groups. Also we excluded patients with nephropathy (eGFR < 30 mL/min/1.73mven more , 34. HowA localised non-ischemic LGE lesion pattern situated mid-myocardial, basal lateral or basal inferolateral is associated with a severe variant of the diabetic cardiac phenotype with increased myocardial mass, increased myocardial extracellular volume, impaired diastolic parameters, increased biomarkers and prevalent complications of diabetes. Further studies correlating magnetic resonance imaging findings with prognosis and pathology are now needed to explain these lesions and their importance for prognosis.


Mesenchymal stem cells (MSCs) from several tissue sources have been investigated in clinical trials for multiple disorders, and mixed results from these trials have been documented , indicat2 strain) heart failure model to study cardiac repair mechanisms mediated by MSCs [2 hamster heart is known to exhibit an early aging phenotype due to progressive loss of cardiomyocytes and functional decline, it is important to determine whether the aging heart of older TO2 hamster may be able to achieve functional improvement in response to MSC therapy. Echocardiography performed 1 month after MSC injection shows that both the saline (HBSS) - and MSC-treated old TO2 hamsters exhibited a similar decline in function as indicated by indistinguishable fractional shortening (FS) and left ventricular end-diastolic dimension (LVDd) between the two groups (This host tissue deficit remains a major challenge in regenerative medicine because the aging population usually require the therapy. We previously used a hamster (TO by MSCs . However2 hamsters, it is ineffective in treating the older cardiomyopathic hamsters. Advanced age typically exhibits more prevalent adverse events in humans. Although the human heart has been known to harbor a significant number of resident stem cells possessing limited regeneration capacity, age-related ECM remodeling and stem cell senescence can lead to declining cardiomyocyte populations and myocardial dysfunction [2 heart may profoundly interfere with the growth factor signaling cascade mediated by the administered MSCs. The finding highlights the progressive nature of the fibrogenic process in the cardiomyopathic heart, which can interfere with the regenerative therapy. Thus, host tissue aging/fibrosis represents a major consideration in the design of MSC therapy.Thus, although MSC therapy is effective in treating younger TOfunction ,8. Similfunction ,10. Sinc


Aim: Left ventricular non-compaction (LVNC) is perceived as a rare high-risk cardiomyopathy characterized by excess left ventricular (LV) trabeculation. However, there is increasing evidence contesting the clinical significance of LV hyper-trabeculation and the existence of LVNC as a distinct cardiomyopathy. The aim of this study is to assess the association of LV trabeculation extent with cardiovascular morbidity and all-cause mortality in patients undergoing clinical cardiac magnetic resonance (CMR) scans across 57 European centers from the EuroCMR registry.Methods and Results: We studied 822 randomly selected cases from the EuroCMR registry. Image acquisition was according to international guidelines. We manually segmented images for LV chamber quantification and measurement of LV trabeculation (as per Petersen criteria). We report the association between LV trabeculation extent and important cardiovascular morbidities  and all-cause mortality prospectively recorded over 404 ± 82 days of follow-up. Maximal non-compaction to compaction ratio (NC/C) was mean (standard deviation) 1.81 ± 0.67, from these, 17% were above the threshold for hyper-trabeculation (NC/C > 2.3). LV trabeculation extent was not associated with increased risk of the defined outcomes  in the whole cohort, or in sub-analyses of individuals without ischaemic heart disease, or those with NC/C > 2.3.Conclusion: Among 882 patients undergoing clinical CMR, excess LV trabeculation was not associated with a range of important cardiovascular morbidities or all-cause mortality over ~12 months of prospective follow-up. These findings suggest that LV hyper-trabeculation alone is not an indicator for worse cardiovascular prognosis. Tertiarmpaction –5. LVNC mpaction , 7. Idenmpaction .Increased awareness of LVNC and improved imaging techniques have led to a surge in its diagnosis. However, studies of healthy cohorts have identified fulfillment of the LVNC imaging criteria in a high proportion of individuals with no clear association to adverse outcomes –11. SimiThere is increasing uncertainty regarding the clinical significance of LV hyper-trabeculation and some have questioned the status of LVNC as a distinct cardiomyopathy . HoweverThere remains some uncertainty regarding generalisability of findings from these low-risk asymptomatic cohorts to patients seen in clinical practice. It is important to ascertain the significance of LV trabeculation in a real-life population with clinical indication for CMR. We present the first prospective multicenter multinational study of the relationship of the extent of LV trabeculation with cardiovascular morbidity and all-cause mortality in real-life patients undergoing clinical CMR in 57 European center.The EuroCMR registry was set up to assess the clinical utility and prognostic impact of CMR in real-life clinical scenarios. Over 37,000 consecutive patients undergoing routine clinical CMR were recruited from 57 center in 15 European countries . Scans wIn order to ensure quality control within the EuroCMR registry, there was requirement for each center to submit a selection of random cases for quality assessment. This created a bank of 980 randomly selected scans collated from all participating center. The current study is based on analysis of this sample. All scans were anonymised, and the demographic and clinical data were not available at the time of image analysis. After quality control checks, 158 scans were excluded due to sub-optimal image quality, the remaining 822 scans are included in this analysis .CMR studies were analyzed using cvi42 software . Left ventricular end diastolic volume (LVEDV), end systolic volume (LVESV), and end-diastolic mass (LVM) were obtained from cine short-axis images covering the LV from base to apex. End-systole and end-diastole were defined by the cardiac phases with the largest and smallest blood pool area at the mid-ventricular level. The operator defined the slice range (base to apex). Endocardial and epicardial contours were manually drawn at end-diastole and end-systole for each slice according to expert consensus recommendations . Papillap < 0.0001 for all parameters). The interobserver variability in the same 100 studies of LV measurements in our group showed very good to excellent reproducibility; ICC values: LVEDVi 0.97, LVESVi 0.88, LVEF 0.71, LVMi 0.92; p < 0.0001 for all measurements.Three long-axis  cine images were used for measuring the thickness of compacted myocardium and trabeculations at the centre of 16 segments of the AHA model . CompactWe considered the following outcomes: all-cause mortality, stroke, atrial fibrillation, severe heart failure (defined as New York Heart Association class 3 and 4). Outcomes were obtained from standardized EuroCMR interviews with follow up duration of 404 ± 82 days. Additionally, we investigated the association of demographic and baseline characteristics on LV trabeculation extent.http://www.R-project.org/]. Continuous variables are summarized with mean  or median . Categorical variables are presented as frequencies and percentages. Differences between LV trabeculation quintiles were evaluated by the analysis of variance (ANOVA) with a post-hoc Tukey test for continuous variables and chi-squared test for categorical variables. Univariate linear regression models were used to assess the relationship between demographic factors, clinical data and LV structural parameters as exposure variables and maximal NC/C as the outcome variable. Cox proportional hazards regression models were used to estimate hazard ratios (HR) and the associated 95% confidence interval (CI) for mortality. Univariable logistic-regression models were used to estimate the odds ratio (OR) and the associated 95% CI for the endpoints. LV trabeculation may be more relevant to individuals with suspected non-ischaemic cardiomyopathies. Therefore, subgroup analysis was performed on a subgroup of 569 subjects without ischaemic heart disease (IHD), defined as having no history of myocardial infarction, coronary revascularisation, or evidence of IHD on CMR.Statistical analysis was performed using R version 3.6.1 (2019-07-05) [R Core Team (2013). R: A language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria. URL: n = 516) of participants were male. Mean age was 59 ± 14 years (range: 16–90). There was substantial burden of cardiovascular risk factors: hypertension 60%, diabetes 14%, dyslipidaemia 40%, smoking history 29%. The most common indications for CMR were evaluation of coronary artery disease (81.9%), cardiomyopathy (15.7%), and myocarditis (8.5%), there was non-exclusivity of indications.Demographic data are presented in n = 59), dilated cardiomyopathy (n = 33), arrhythmogenic right ventricular cardiomyopathy (n = 1), unclassified cardiomyopathy (n = 18). Myocarditis was diagnosed in 29 patients.No abnormality was detected in 49% of the CMR scans. IHD was diagnosed in 25% of patients. Amongst these, myocardial ischaemia was detected in 147 cases, syndrome X in 64, and cardiac thrombus in one patient. Non-ischaemic cardiomyopathies were diagnosed in 111 patients: HCM (n = 142), and in at least two segments for 5.1% (n = 42). The maximal NC/C had a log-normal distribution. The mean maximal NC/C for each participant's analyzed segments was 1.81 ± 0.67. Greater magnitude of trabeculation was detected in lateral segments, and increased from base to apex. LV trabeculation was not seen in the basal infero-septal segment of any participant.NC/C was calculated in 10,456 (79.5%) segments. The remaining 2,696 segments either had insufficient contrast between the blood pool and the myocardium or were off-axis impeding confident measurement of NC/C. NC/C >2.3 was measured in at least one segment for 17.2% of participants (≤35%) was detected in 5.6% (n = 46). There were no differences between the quintiles of maximal NC/C in the frequency of severe LV systolic dysfunction (p = 0.39).LV volumes and functional parameters are presented in p < 0.05) and decreased with age . Maximal NC/C was larger by 0.5 for every 100 ml/m2 increase in LVEDVi (p < 0.0001), and by 0.4 for every 100 ml/m2 increase in LVESVi (p < 0.0001). Maximal NC/C was also larger by 0.06 units for every 10% decrease in LV ejection fraction (p < 0.01).NC/C was higher in women than in men .Mortality data was available for 530 participants with mean follow-up of 404 ± 82 days. During the follow up period, 10 deaths were recorded (death rate of 1.9%). The extent of maximal NC/C was not associated with increased mortality. There was no difference in survival between quintiles of maximal NC/C . Similarn = 333) were in NYHA class I, 11.7% (n = 60) in NYHA class II, 3.7% (n = 19) in NYHA class III, and <1% (n = 5) in NYHA class IV. Increasing quintiles of the maximal NC/C were not associated with greater odds of severe heart failure defined as NYHA class III and IV (Data on heart failure status assessed by the New York Heart Association (NYHA) classification was available for 513 participants. Of these, 65% (I and IV . Sensitip = 0.16) (Twenty-eight (3.4%) participants had prior history of stroke. There were no differences in the prevalence of stroke between the quintiles of maximal NC/C ( = 0.16) . One par = 0.16) .p = 0.16) (Twenty (2.4%) participants had a history of atrial fibrillation. There were no differences in the frequency of atrial fibrillation among quintiles of maximal NC/C ( = 0.16) .n = 10), severe heart failure (n = 24), and stroke . Some participants had more than more of these outcomes, overall, 57 (12.8%) individuals had at least one MACE. The extent of maximal NC/C expressed as a continuous variable and in quintiles was not associated with frequency of MACE  and more likely to be women (p < 0.05). They had lower prevalence of diabetes (p < 0.0001) and hypertension (p < 0.05) . There was no association between maximal NC/C and mortality  or MACE . The number of individual outcomes  was too small in this subgroup for sufficiently powered statistical analysis.The 569 individuals without IHD were younger ( < 0.05) . A greatIn this multicenter multinational study of real-life patients with clinical indication for CMR, we identified no association between the extent of LV trabeculation and increased risk of all-cause mortality, severe heart failure, stroke, atrial fibrillation, or MACE composed of all these parameters. The same result was observed with subgroup analysis of individuals without IHD and with stratified analysis using the threshold of NC/C >2.3. Women and younger individuals had greater magnitudes of LV trabeculation, as did those with larger cavity volumes and lower LVEF.n = 292), we limited our analysis to individuals with an explicitly documented mortality outcome . Of course, there is still potential for bias, e.g., relating to excess deaths in the missing cohort, however, we believe our approach is the method with least potential for bias within the limitations of the information available. Our observations reflect short-term risk with an average follow-up period of just over 12 months; we cannot exclude longer term prognostic significance of LV trabeculation.Whilst the inclusion of consecutive real-life clinical patients is a strength of this study, analysis of individuals with relevant symptoms in a more selective manner may provide more meaningful context for interpretation of clinical significance of LV trabeculation. However, this approach would introduce ascertainment bias, which has hampered previous studies with highly selective cohorts. We only used one method to quantify trabeculation extent. However, whilst other methods exist , method n = 142), however this did not translate to an exaggerated rate of adverse events. Similar prevalence of LV hyper-trabeculation has been reported in multiple studies of healthy cohorts with no association to poor outcomes (de-novo appearance and spontaneous disappearance of LV trabeculae in pregnancy and post-partum as had been suggested (The proportion of individuals in our study with NC/C >2.3 in at least one segment was high (17%, outcomes , 11. Notoutcomes . Amzulesoutcomes . High raoutcomes . Studiesoutcomes . These cuggested .Ivanov et al. present the only other study, aside from ours, assessing the significance of LV trabeculation extent in a real-life clinical setting . Consistin-utero “myocardial compaction” phase (in-utero rather than a distinct genetic cardiomyopathy.Thus, there is accumulating evidence from multiple clinical studies, with a variety of study designs, in different populations disputing the clinical significance of LV hyper-trabeculation. Furthermore, the genetic and embryologic origins of LVNC have been questioned. Although several genetic links to LVNC have been established, there is significant overlap with other cardiomyopathies and no single gene has been reliably and specifically linked to LVNC . Addition” phase –26. The n” phase . It is cWhilst initially presented as a rare genetic cardiomyopathy with poor outcomes, growing evidence from a number of sources in different populations shows high prevalence of LV hyper-trabeculation by imaging criteria with no evidence of associated prognostic significance. Furthermore, the genetic and embryologic evidence for the etiology of LVNC are equivocal, with the absence of a clear gene-phenotype link and lack of evidence for an embryologic myocardial compaction phase. It seems increasingly likely that the initial reported poor outcomes relating to LVNC were the biased results of highly selective tertiary centre cohorts. It is clear from the available evidence, that the current imaging criteria for quantification of LV trabeculation are too broad, capturing high proportions of individuals with no underlying cardiac disease and no increased risk of adverse events.This study adds to the growing evidence that LV hyper-trabeculation in isolation is not a predictor of risk and does not mandate further investigation or follow-up. In cases, where there is uncertainty regarding the existence of LVNC as a distinct cardiomyopathy, physicians should take a holistic approach and exert caution in making this diagnosis.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.All participating center had approval from local institutional ethics review boards, and all patients provided written informed consent in accordance to the Declaration of Helsinki, under standards of the EuroCMR registry. The patients/participants provided their written informed consent to participate in this study.FZ, SEP, and SM conceived the study. FZ collated, analyzed the CMR images, and led and performed the statistical analysis. ZR-E, FZ, and SEP wrote the manuscript. ZR-E contributed to cross-checking of the analysis. MK contributed to image analysis. All co-authors contributed to critical revision of the manuscript and approved the final version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Patients with left ventricular assist devices (LVAD) require specific anesthetic and hemodynamic considerations. We report the specific anesthetic preparation and management in this scenario.We present the case of a 66-year-old male with a HeartMate II LVAD undergoing robotic prostatectomy for prostate cancer in the steep Trendelenburg position. We employed central venous and radial arterial access, LVAD pump parameters, near-infrared sensor of cerebral oximetry, and transesophageal echocardiography for monitoring. Hemodynamics were managed with nicardipine, dobutamine, epinephrine, and phenylephrine during abdominal insufflation, operative positioning, and desufflation. The patient had a successful procedure, was discharged on postoperative day 2, and achieved surgical cure of his prostate cancer.By presenting the first detailed account of anesthetic management in this scenario, we provide a clinical vignette for use by the clinical anesthesiologist in his or her preparation prior to caring for this type of patient. The use of left ventricular assist devices (LVADs) for patients with heart failure has become widespread. Although cardiac transplantation is considered definitive therapy, shortage of donor hearts has led to the increasing use of LVADs as a bridge to transplant or final therapy devices , 2. PatiA 66-year-old male patient with a HeartMate II model LVAD  4 months postoperative from implant was planned for robotic prostatectomy for prostate cancer after multidisciplinary oncologic case discussion, with a goal for a surgical cure in order to obtain candidacy for heart transplantation. His past medical history included viral cardiomyopathy with severe left ventricular dysfunction (EF 20%), ventricular tachycardia status post-implanted cardiac defibrillator (ICD), stage 2 chronic kidney disease, and hypertension.Preoperative evaluation with a ramp transthoracic echocardiogram (TTE) demonstrated an optimal pump speed of 5300 revolutions per minute (RPM). This type of TTE is a protocolized assessment of LVAD operation during which continuous evaluation of the heart is used to determine an optimal RPM at which the patient’s ventricular function, aortic opening, and cardiac output are optimal . The TTEWarfarin was held the night prior to the surgery. On the morning of prostatectomy, prothrombin international normalized ratio (INR) resulted as 3.2. He was given prothrombin complex concentrate . PT-INR was again drawn upon arrival in the operating room and resulted at 1.4.The intraoperative team included the urologic surgeon, general anesthesiologist, pharmacist, and a cardiac perfusionist. After placement of ASA standard monitors, a 20-gage right radial arterial line was placed under ultrasound guidance. Central venous access was obtained via the right internal jugular vein. Additional variables monitored included near-infrared cerebral oximetry , bispectral index monitor , and LVAD power index and rotation speed communicated to the anesthesia team by the cardiac perfusionist.Anesthesia was induced using fentanyl 1.5 mcg/kg, propofol 1 mg/kg, and rocuronium 1.2 mg/kg, which resulted in relative hypotension that responded to a bolus of 250 ml of 5% albumin. Rapid sequence induction was chosen to rapidly secure the airway and reach a stable plane of anesthesia using a volatile anesthetic (sevoflurane). Albumin was chosen as it is our institutional colloid of choice. Full details of the anesthetic record are provided in Fig. Surgical insufflation of the abdomen resulted in an acute rise in blood pressure, which was associated with the decrease in LVAD flow rate and increased power requirement presumed due to high afterload. This was treated with 0.75 mg/kg bolus of propofol and initiation of dobutamine and nicardipine infusions with good effect. The patient was placed in a Trendelenburg position in a stepwise manner while monitoring hemodynamics and LVAD indices. Trendelenburg positioning was associated with a rise in the central venous pressure (CVP), though this was tolerated. The LVAD showed the following parameters: pulsatility index 3.9, flow 4.5 L/min, rpm 5300, and power 3.8 W.Approximately midway through the procedure, the pulsatility index began to decrease and central venous pressure (CVP) increased by 50%. This generated concern for worsening right ventricular function creating decreased left ventricular and LVAD preload. Cardiac anesthesia was called for intraoperative transesophageal echocardiogram (TEE). TEE demonstrated mild bowing of the intraventricular septum into the right ventricle and moderate right ventricular dysfunction see Fig. . InhaledRobotic prostatectomy was completed in the standard fashion. Ten minutes prior to abdominal desufflation, nicardipine infusion was paused and dobutamine infusion rate was reduced. At the time of desufflation, there was an acute drop in arterial blood pressure which required initiation of phenylephrine and epinephrine infusions as well as a crystalloid bolus of 250 ml. Neuromuscular blockade was reversed using sugammadex , and the patient was extubated. All vasoactive medications were able to be discontinued at this time, and the patient was transported to the cardiac surgery intermediate care unit in stable condition.Postoperatively, he did not require vasoactive infusions. PT-INR was drawn postoperatively at 5 h after administration of prothrombin concentrate and resulted at 1.5. His preoperative dose of warfarin was resumed on the evening of postoperative day 0, and INR on postoperative day 1 was 2.0. The remainder of his postoperative laboratory studies was stable from preoperative values. He was discharged home on postoperative day 2. Subsequent evaluation by urologic surgery has determined that the procedure was curative. He is currently undergoing evaluation for cardiac transplant.In this case report, we share our experience with the anesthetic management of a patient with an LVAD undergoing laparoscopic abdominal surgery in steep Trendelenburg positioning.The ramp TTE is perhaps the most important preoperative test in this situation and was initially reported as a means to optimize LVAD pump speed and to diagnose device thrombosis . The tesThe hemodynamics of patients with LVAD devices have unique considerations with respect to cardiac output . Blood fThe literature does not provide specific recommendations for perioperative management of anticoagulation in LVAD patients. The majority of literature focuses on institutional experience and case reports, though it appears that the majority of patients with LVAD presenting for non-cardiac surgery receive some type of pharmacologic reversal .Our case supports previous findings that patients with LVAD devices may safely tolerate procedures that require abdominal insufflation as well as potentially problematic positioning such as steep Trendelenburg. This was demonstrated by Khemees et al., although that report and others for laparoscopic nephrectomy  and sleeAs the number of patients with ventricular assist devices presenting for non-cardiac procedures grows, the economics and workforce supply considerations of who manages these patients may favor generalist responsibility with specialist consultation as needed. In our case, preoperative evaluation and optimization facilitated this strategy. If a combination of system-based approaches, local expertise, and interdisciplinary collaboration can be established, the value proposition of managing these patients using a generalist model may create substantial savings for healthcare systems that care for these patients.In this report, we present a case of moderately complex non-cardiac surgery delivered by one potential configuration of the care team with a specific focus on anesthetic management and considerations. General anesthesiologists with adequate motivation, proper preparation, and collaborative systems may be able to provide safe anesthetic care and positive outcomes for LVAD patients undergoing non-cardiac surgery.


The molecular mechanism for worsening left ventricular (LV) function after mitral valve (MV) repair for chronic mitral regurgitation remains unknown. We wished to assess the LV transcriptome and identify determinants associated with worsening LV function post-MV repair. A total of 13 patients who underwent MV repair for chronic primary mitral regurgitation were divided into two groups, preserved LV function (N = 8) and worsening LV function (N = 5), for the study. Specimens of LV from the patients taken during surgery were used for the gene microarray study. Cardiomyocyte cell line HL-1 cells were transfected with gene-containing plasmids and further evaluated for mRNA and protein expression, apoptosis, and contractile protein degradation. Of 67,258 expressed sequence tags, microarrays identified 718 genes to be differentially expressed between preserved-LVF and worsening-LVF, including genes related to the protein ubiquitination pathway, bone morphogenetic protein (BMP) receptors, and regulation of eIF4 and p70S6K signaling. In addition, worsening-LVF was associated with altered expressions of genes pathologically relevant to heart failure, such asdownregulated apelin receptors and upregulated peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PPARGC1A). HL-1 cardiomyocyte cells transfected with ubiquitination-related genes demonstrated activation of the protein ubiquitination pathwaywith an increase in the ubiquitin activating enzyme E1 (UAE-E1). It also led to increased apoptosis, downregulated and ubiquitinated X-linked inhibitor of apoptosis protein (XIAP), and reduced cell viability. Overexpression of ubiquitination-related genes also resulted in degradation and increased ubiquitination of α-smooth muscle actin (SMA). In conclusion, worsening-LVF presented differential gene expression profiles from preserved-LVF after MV repair. Upregulation of protein ubiquitination-related genes associated with worsening-LVF after MV repair may exert adverse effects on LV through increased apoptosis and contractile protein degradation. Severe, chronic, isolated mitral regurgitation (MR) is characterized by progressive dilation of the left ventricle (LV) due to significant volume overload.Severe MR also leads to a considerate remodeling of cardiac pathology including chamber dilation, cardiac fibrosis, and cardiomyocyte hypertrophy and apoptosis. The LV ejection fraction (EF) can be falsely normal due to the favorable condition for LV shortening dynamics when the LV myocardium has been irreversibly injured by severe MR. Surgery is the only promising therapy for severe MR and, as per the guidelines, an early surgery in symptomatic or asymptomatic patients is recommended before LV contractile function begins to deteriorate .However, the timing of surgical correction of MR remains debatable ,3. TherePatients with normal LV EF before surgery may undergo LV remodeling and dysfunction after MV repair. The literature available describing LVremodelingafter repair of chronic MR is scarce and the mechanism is largely unclear . It remaTherefore, in this study, we explored the molecular mechanism of progressive LV remodeling despite mitral valve repair using a genome-wide approach. We revealed differential gene expression profiles in LV from patients with severe MR, with and without progressive LV remodeling after MV repair.The clinical characteristics of patients are listed in p < 0.05), among which 138 genes were upregulated and 580 genes were downregulated  were measured by microarray and 718 differentially expressed genes were identified (>25% fold-change) between worsening-LVF and preserved-LVF patients  to be upregulated in worsening-LVF compared to preserved-LVF, we further evaluated the effects of these genes in cardiomyocytes. Plasmids containing cDNAs of the genes that were upregulated in worsening-LVF, namely USP15, PSMD7, UBE2D1, DNAJC15, DNAJC8, and pcDNA3.1, as their control were individually transfected into HL-1 cardiomyocytes. The efficacy of transfection was confirmed by Western blot .Ubiquitin requires activation by UAE-E1 before ligation to substrate proteins, which are recognized and degraded by proteasomes. Therefore, we measured the expression level of UAE-E1 in the HL-1 cells using a Ubiquitin Human ELISA Kit 24-h after plasmid transfection. The expression of UAE-E1 was significantly upregulated in HL-1 cardiomyocytes transfected with plasmids containing USP15, PSMD7, UBE2D1, DNAJC15, and DNAJC8 compared with pcDNA3.1, suggesting that transfection with plasmid containing ubiquitination-related genes would significantly activate protein ubiquitination activity A.Since worsening myocardial function may be attributed to loss of cardiomyocytes as well as to loss of intracellular contractile proteins, we further evaluated the effect of these ubiquitination-related genes on cardiomyocyte apoptosis and intracellular contractile protein degradation. The results obtained showed that both the apoptotic activity B and myoCaspase-3 and caspase-9 are proteases well known for their roles in carrying out apoptosis. X-linked inhibitor of apoptosis protein (XIAP) is one of the key proteins exerting anti-apoptotic activity by inhibiting caspase-3 and caspase-9. XIAP is known to be ubiquitinated and degraded by the UPS . In HL-1The expression of active/cleaved caspase-9, relative to pro-caspase-9-, was significantly increased, and the expression of XIAP was significantly downregulated in cardiomyocytes transfected with USP15, UBE2D1, DNAJC15, and DNAJC8 compared with pcDNA3.1 control A. These To evaluate if XIAP and the contractile protein, α-smooth muscle actin (SMA), were labeled by ubiquitin, we performed co-immunoprecipitation to study the binding of XIAP and SMA with ubiquitin. The cell lysates were immunoprecipitated with anti-ubiquitin antibody and the expression of XIAP/SMA was evaluated by Western blot using an anti-XIAP and anti-SMA antibody. We performed Western blot for SMA and XIAP on the same membrane. The levels of total ubiquitinated proteins in HL-1 cardiomyocytes transfected with different plasmids were presented in In the present study, we identified differentially expressed genes in LV after MV repair in patients with worsening LV function as compared to patients with worsening LV function (worsening-LVF) with normal pre-operative LV function. The results of the microarray showed the expression of NPPB, which codes for brain natriuretic peptide (BNP), a well-known protein associated to decompensated heart failure, was around 15-fold higher in worsening-LVF compared with preserved-LVF . It implChronic, severe MR causes LV volume overload, resulting in LV remodeling and leading to heart failure. Several signaling pathways and phenotypes have been reported to be altered in humans with chronic MR and animal models of volume overload, including activated β-adrenergic signaling, calcium-handling proteins , and dowThe UPS is a major intracellular system responsible for the degradation of proteins, and plays a major role in regulating many cellular processes ,14. An iIt has been shown that the UPS regulates cardiac apoptosis . PiacentUbiquitin-specific protease 15 (USP15), a member of cysteine protease deubiquitinases. It has been shown USP15 would remove ubiquitin from pro-caspase 3 , which mTaken together, our findings suggest both UPS and XIAP have the potential to be novel therapeutic targets in preventing the progression of MR-related heart failure.We showed that the expression of apelin receptor (APLNR), a G-protein coupled receptor widely expressed throughout the heart, was downregulated in worsening-LVF. Targeting APLNR helps prevent heart failure resulting from pressure overload via suppression of angiotensin-converting enzyme expression and pathogenic angiotensin II signaling . In humaThe sample size of the study is small, and may not reflect the whole profile of genetic changes and the mechanism for worsening LV function after surgical mitral valve repair. The duration of follow-up echocardiography after MV surgery in the study is not long, which may not reflect the long-term alterations in the patients. Considering the heterogeneity of patients, future studies with bigger sample sizes will advance the findings. We did not perform quantitative protein analysis due to the unavailability of human specimens. The analysis of mRNA transcripts may not exactly reflect the expression of proteins and their function, as we showed, and altered UPS may enhance protein degradation.We explored the different molecular signaling associated with worsening LV function in patients with previously preserved LV function after MV repair for the first time, whichmay help reveal novel potential therapeutic targets for heart failure.The study protocol was approved by the institutional review board of ChangGung Memorial Hospital. Patients with severe, chronic, isolated MR and LV EF >55% who underwent MV repair were included in the study. Between December 2013 and June 2015, 18 participants were recruited and LV specimens were obtained after patient consent. These patients were operated on by the same surgeon with a technical unit to avoid any deviation in the procedure. All 18 patients were followed up and the LV EF and diameter were evaluated by echocardiography up to six to nine months after surgical mitral repair. The patients following the criteria were recruited into two groups; group A included patients with improved or unchanged post-operative LV EF without increase in LV end-diastolic diameter (LVEDD) and group B included patients with reduced post-operative LV EF >10% with or without increase in LVEDD. The LV specimens of patients from both groups were collected for a microarray study. Patients who showed improved or unchanged LV EF but increased LVEDD were not included in the study. Therefore, of the 18 patients, eight and five patients were included in group A  and group B  for furtherstudy, respectively.The isolation of tissue RNA and its quality and quantity analysis were carried out as described previously . Gene exGenes of interest were analyzed using ingenuity pathway analysis . Fisher’s exact test was applied to examine the likelihood that the association between genes of interest and related pathways was not random. Functions that were predicted to be influenced by the differentially expressed genes were ranked according to their order of significance and were further analyzed by relevance to cardiovascular diseases or second messenger and intracellular signaling.TRIzol reagent was used for RNA extraction from the LV specimens, and q-PCR was performed as previously described . GlyceraHL-1 cardiomyocytes were cultured in the Claycomb medium and subjected to field stimulation as described previously .A set of plasmids containing cDNAs of USP15, PSMD7, UBE2D1, DNAJC15, and DNAJC8P were purchased from OriGene . The vectors were transfected into HL-1 cardiomyocytes using Lipofectamine 2000  and used for experiments 24 h after transfection.HL-1 cells were grown to confluence in 60 mm dishes. The concentration of UAE-E1 in the HL-1 cells was determined by a mouse E1/ubiquitin-activating enzyme ELISA kit  after plasmid transfection, as per the manufacturer’s instructions.Apoptosis was detected by TUNEL assay. After plasmid transfection, the HL-1 cells were grown in 24-well culture dishes for 24 h. The cells were then fixed on slides and TUNEL reaction mixture  was added to the sections according to the manufacturer’s instructions, followed by incubation at 37 °C for 60 min. After removal of the TUNEL reagent, the slides were rinsed with PBS, and TUNEL-positive cells were evaluated using a confocal microscope  with excitation at 488 nm with an argon laser. Emission was recorded using a Longpass> 550 nm filter set to acquire two-dimensional images (512 × 512 pixel).The cell viability was measured by cell counting kit-8  according to the manufacturer’s protocol. Briefly, HL-1 cardiomyocytes were seeded into a 96-well plate at 37 °C with 5% CO2. Then, cells were transfected with different plasmids for 24 h. Afterwards, 10 μL of CCK-8 reagent was added to each well and incubated for 2 h at 37 °C. Finally, the absorbance at 450 nm was determined using a microplate reader .Western blot was performed following the protocol as described previously . Briefly2, 0.2 mmol/L EDTA, 0.5% Nonidet P-40, and 0.5 mmol/L dithiothreitol) and immunoprecipitated with anti-ubiquitinylated protein antibodies  for 2 h at 4°C. Following incubation with 50 μL of Protein A-Sepharose for 1 h at 4°C, beads were collected, washed three times, and resuspended in SDS sample buffer. The immunocomplexes were resolved by SDS-PAGE and analyzed by Western blot with anti-XIAP and anti-SMA .The lysates from HL-1 cells were harvested with lysis buffer  primary antibody followed by Cy3 -conjugated secondary antibody. Nuclei were counterstained using DAPI. Myosin degradation was quantified as the cytoplasmic myosin (MHC) area divided by the nuclear area. The relative expression levels of MHC were normalized to the control level. For each analysis, at least five fields were selected at random to observe >30 myocytes.p value of <0.05 and relative fold-change >25%. The data in the figures are presented with box-and-whisker plots. Unpaired Student’s t test and one-way analysis of variance (ANOVA) with post hoc Tukey tests were applied for the two groups and multiple comparisons, respectively. A p value of <0.05 was considered statistically significant. The original data files of the microarray have been deposited in Gene Expression Omnibus (http://www.ncbi.nlm.gov/geo) with the access number GSE150661.For patient characteristics, mean ± standard deviation for continuous variables, and count and percent for categorical variables are presented, respectively. Fisher’s exact test was used for category comparisons between groups. For microarray analysis, the data were normalized to the expression of housekeeping genes like GAPDH for each set and the difference between the two groups was considered significant for a


The role of neutrophil extracellular traps (NETs) in acute heart failure is unknown. We recently showed that interleukin 8, a putative NETs stimulator, was associated with myocardial recovery in acute heart failure complicating ST-elevation myocardial infarction (STEMI). In this exploratory post-hoc study, we aimed to investigate the role of NETs components in relation to myocardial function and interleukin 8 in STEMI patients with symptomatic acute heart failure.In 61 STEMI patients developing acute heart failure within 48 hours of successful revascularization, wall motion score index (WMSI), global longitudinal strain (GLS) and left ventricular ejection fraction (LVEF) were assessed by echocardiography at baseline and on day 5. Blood drawn at baseline and days 1, 2 and 5 was used to quantify double-stranded DNA (dsDNA), myeloperoxidase-DNA complexes (MPO-DNA) and citrullinated histone 3 (CitH3). The area under the curve (AUC) of each NETs marker and interleukin 8 was approximated for the first 5 days.AUC and MPO-DNAAUC correlated significantly with change in WMSI from baseline to day 5 , whereas NETs AUCs did not correlate with changes in GLS and LVEF. dsDNAAUC was significantly correlated with interleukin 8AUC . However, mixed model regression could not identify a significant effect of the NETs components on myocardial function parameters.dsDNAIn this cohort with acute heart failure complicating STEMI, NETs components were partly correlated with myocardial function and interleukin 8 levels, yet no causal relationship between NETs components and myocardial recovery could be established.NCT00324766.ClinicalTrials.gov, identifier:  Although patient outcomes after ST-elevation myocardial infarction (STEMI) have improved considerably over the last decades, the incidence of heart failure during hospitalization for the index myocardial infarction (MI) is still estimated at nearly 30% , necessiAmong the inflammatory mediators that are elevated post-MI, interleukin 8 (IL-8) attracts neutrophils that infiltrate the myocardium within hours of infarction and contribute to myocardial injury through recruitment of other inflammatory mediators, degranulation and production of reactive oxygen species . We receWe hypothesized that increased levels of circulating NETs components might reflect an unfavourable immune response as part of sterile inflammation post-MI, potentially affecting myocardial recovery after ischaemia and reperfusion (IR) injury. The present exploratory study therefore aimed to investigate whether NETs component levels were associated with echocardiographic indices of myocardial function in patients with AHF complicating STEMI. Secondarily, we explored whether there was any association between NETs components and circulating IL-8 levels as a putative stimulator of NETosis.n = 61) were enrolled in the LEvosimendan in Acute heart Failure (LEAF) trial at the Oslo University Hospital Ullevål from 2006–2010. Results and details of the study design have previously been published [Patients included in this post hoc observational sub-study  despite volume therapy. A subgroup of patients in cardiogenic shock were stratified by block randomization, and the inclusion criteria were systolic blood pressure <90 mmHg after one hour of adequate volume therapy, or systolic blood pressure 90–100 mmHg with ongoing inotropic treatment, in addition to clinical signs of hypoperfusion including oliguria, cold and clammy extremities, or reduced consciousness.NCT00324766.The study conforms with the principles outlined in the Declaration of Helsinki, and was approved by the South East Regional Ethics Committee, Norway. All participants gave written informed consent. The LEAF trial is registered with ClinicalTrials.gov, identifier: Echocardiography was performed by two experienced echocardiographers at baseline, on days 1 and 5, and at the 6-week follow-up. A single observer performed all echocardiographic analyses, scoring 16 predefined segments of the left ventricle (LV) as either normal or hyperkinetic (1 point), hypokinetic (2 points), akinetic (3 points), or dyskinetic (4 points) using a digital ultrasonic device . WMSI is calculated as the sum of scores divided by number of assessed segments, yielding a score = 1 for a normally functioning LV, and >1 in the case of LV dysfunction. Upon study inclusion, patients were required to have decreased LV wall motion in at least 3 of 16 segments. Left ventricular ejection fraction (LVEF) was assessed using Simpson’s biplane method or by visual approximation. Global longitudinal strain (GLS) measurements were derived from speckle tracking echocardiography images of the myocardium in three apical views, with automatic calculations by the aforementioned software. Analysis required optimal tracking in at least 14 segments and absence of AF.Infarct size was quantified using 99 m-tetrofosmin ECG-gated myocardial single photon emission computed tomography (SPECT), expressing the perfusion defect as percentage of LV mass.® tubes, and then kept on ice until centrifugation within 30 min at 3000g and 4°C for 20 min. Serum was centrifuged at 2500g for 10 min within an hour of sampling. Samples were ultimately stored at -80°C pending analysis.Venepuncture was performed at baseline on study inclusion (median 22 hours after PCI), and subsequently on days 1, 2 5, and 42 (6-week follow-up). Plasma was collected using EDTA and citrated Vacutainer®  and a fluorimeter  with an inter-assay coefficient of variation (CV) of 8.2%. Serum levels of myeloperoxidase-DNA (MPO-DNA) complexes were assessed using an enzyme-linked immunosorbent assay (ELISA) technique previously described by Kessenbrock et al. [Plasma (EDTA) levels of double-stranded DNA (dsDNA) were quantified using a fluorescent nucleic acid stain, Quant-iT PicoGreenk et al. . BrieflyCirculating levels of IL-8 were quantified in serum using a commercial ELISA  with an inter-assay CV of 7.6%. Peak cardiac troponin T (TnT) was measured in serum using a commercial immunoassay .th, 75th percentile), or proportions (%) as appropriate. The distribution of data was judged based on skewness, kurtosis and Q-Q plots (rs) was used to perform correlation analyses, and 95% confidence intervals (CI) for rho were calculated using Fisher’s Z transformation. Continuous data were compared by use of the unpaired two-sample Student t-test or Mann-Whitney U test depending on data distribution. The Friedman test was performed to assess differences in levels of the NETs components across repeated samples, whereas Wilcoxon signed-rank tests were used to assess differences in the levels of the markers pairwise compared to baseline levels. Area under the curve (AUC) from baseline to day 5 was approximated for each NETs marker and IL-8 using the trapezoidal rule (n = 53 due to missing samples day 2). Longitudinal mixed model regression analysis was performed to assess the effects of NETs levels on myocardial function, testing for potential confounders  , median (25-Q plots . As the eak TnT) . The levon 4.0.2 .The patients had a mean age of 65 years and approximately 30% were women . Only onp ≤ 0.017 for all) (p < 0.001). No significant changes were observed in CitH3 levels during the first five days. When comparing levels of the NETs components at baseline through day 5 to the levels at six weeks, both dsDNA and MPO-DNA were significantly higher at all previous time points . CitH3 was only significantly higher at baseline, day 2 and day 5 (p < 0.04) and just missed significance on day 1 (p = 0.06).There was a significant difference in the levels of all three NETs components across the five sampling time points from baseline to six weeks (day 42), assessed using Friedman tests  . dsDNA lAUC, MPO-DNAAUC and CitH3AUC, were 2000 ng/ml∙days , 1.29 OD∙days , and 26.4 ng/ml∙days , respectively.The AUCs for the three NETs components, expressing total “burden” over the first five days, hereafter referred to as dsDNAn = 9) or absence (n = 52) of cardiogenic shock at baseline, there were no significant between-group differences in dsDNA levels at baseline or at 6-week follow-up, but dsDNA was significantly higher in the group with shock as compared to patients without shock on day 1, 2, and 5 (n = 6), as compared to those without, had significantly greater dsDNAAUC  vs 1982 , p = 0.02) and CitH3AUC  vs. 24.3 , p = 0.02), but no significant difference was observed between the two groups for MPO-DNAAUC (p = 0.10).Stratifying patients according to the presence  (AUC. There were no significant correlations between NETs AUCs and change in GLS or LVEF during the first five days (dsDNActively) . No signive days . In ordeTo investigate potential effects of NETs components on changes in myocardial function as illustrated in AUC was significantly correlated with baseline IL-8 . However when investigating potential mediating effects of the NETs components on IL-8 effects in the aforementioned mixed model regression analysis, no significant effects were observed.Correlations between NETs levels and IL-8 at corresponding time points are shown in AUC correlated significantly with peak TnT levels , whereas MPO-DNAAUC and CitH3AUC did not. Neither peak TnT nor SPECT-determined infarct size differed significantly between cardiogenic shock groups.Myocardial infarct size, as determined by SPECT after 6 weeks, did not correlate significantly with levels of the NETs components at any time point, nor with NETs AUCs . dsDNAAUIn this study of 61 patients with AHF complicating a PCI-treated STEMI, we could not confirm that NETs components play a role in myocardial recovery after STEMI. The association with IL-8, although weak, may support IL-8 to be a potential NETs inducer in this setting. Although no assertions can be made with respect to causality, to the best of our knowledge, this is the first report on the temporal profile of circulating NETs components in AHF complicating STEMI.Although the total burden of dsDNA and MPO-DNA were associated with change in WMSI from baseline to day 5, longitudinal mixed model analysis showed neutral effects with respect to NETs and myocardial recovery. Although not specifically examining AHF patients, NETs burden has previously been linked to infarct size and left ventricular function ,15. NeutDespite some significant correlations between WMSI and NETs burden, the findings for GLS and LVEF were inconsistent. The observations with respect to changes in LVEF may be obscured by the compensation of non-infarcted myocardium, supported by evidence from one cohort with myocardial infarction in which most had LVEF within the normal range, whilst 70% had impaired GLS . DespiteNETs burden as quantified by dsDNA and CitH3 was significantly higher amongst patients with cardiogenic shock, but with no association to final infarct size, supporting the observation that NETs burden may be partly related to heart failure with increas0065d filling pressure and low cardiac output, and not only myocardial necrosis. Increased NETosis in cardiogenic shock may suggest that NETs components are released as a result of systemic hypoperfusion and organ hypoxia, with NETosis potentially being triggered on a broader scale. In line with this, dsDNA, MPO-DNA and histones have all been reported to be elevated in patients with septic shock ,23, leadin vitro data showing that IL-8 stimulates NETs release [An association between IL-8 and NETosis was only corroborated by the correlation between dsDNA and IL-8 at baseline. In this cohort, we have previously shown that IL-8 levels were significantly higher at baseline than at all later points , seeming release , though limit the inflammatory response. For instance, NETs could perhaps restrict injury to a smaller area, trigger compensation of the remote non-infarcted myocardium, or initiate repair, however such causal inferences cannot be drawn from the present study. The crucial role of neutrophils in the conversion of macrophages to a reparative phenotype, as well as the timely development of a fibrotic scar, is evident from studies in mice showing that neutrophil depletion leads to poorer myocardial function and the development of AHF [With respect to the neutral effects of NETs components on myocardial recovery, it might be discussed whether some of the neutrophil components also act to t of AHF . Recent t of AHF  and yet t of AHF . Other bt of AHF , we coult of AHF .Limitations of this study include sample size, potential impact of outliers, and the number of analyses performed, although the study is exploratory and observational in nature, precluding conclusions with respect to causality. Whether dsDNA is a nonspecific marker of cellular injury or a marker of NETosis cannot be determined from our data, nor can we dissect whether our observations are related primarily to MI or AHF. Moreover, we can only speculate as to possible reasons for the inconsistent observations between the different NETs components, potentially due to differential release of the individual components, differing half-life in the circulation, or methodological issues. The primary endpoint (change in WMSI from baseline to day 5) was in part chosen based on the pharmacokinetic properties of levosimendan, thus limiting the study hypothesis to neutrophil effects in the acute and subacute phase. Finally, concurrent conditions such as infections may have affected neutrophil activity and general inflammation.Our results provide novel knowledge of how levels of circulating NETs components evolve in patients with large myocardial infarctions and symptomatic heart failure. Despite some correlations between NETs components and myocardial recovery after STEMI, and the observation that IL-8 might be involved in enhancing NETs formation in AHF, no significant cause-effect relationships could be shown. The mechanistic links and potential clinical utility of NETs components as biomarkers in AHF require further investigation.S1 Fig(DOCX)Click here for additional data file.S1 Table(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file.S3 Table(DOCX)Click here for additional data file.S4 Table(DOCX)Click here for additional data file.S1 File(PDF)Click here for additional data file.S2 File(XLSX)Click here for additional data file.


Myocardial rupture after thrombolysis is an often fatal consequence. For patients with myocardial infarction and ischemic stroke within a short timeframe, a catheter‐based therapy to retrieve emboli poses a valid therapeutic option in case of a treatment target in CT‐Angiography. Myocardial rupture after thrombolysis is an often fatal consequence. For patients with myocardial infarction and ischemic stroke within a short timeframe, a catheter‐based therapy to retrieve emboli poses a valid therapeutic option in case of a treatment target in CT‐Angiography. Intravenous fibrinolytic therapy given late in the setting of myocardial infarction is associated with an increased risk of hemorrhagic necrosis and myocardial rupture. Due to the development of primary percutaneous coronary interventions for myocardial infarction instead of intravenous fibrinolysis, a dramatic decrease in regard to mortality has been registered, mainly due to less myocardial re‐infarction. Patients with a recent myocardial infarction and subsequent stroke present a dilemma to the treating physicians due to the increased potential of intramyocardial hemorrhage and myocardial perforation after iv thrombolysis for ischemic stroke.In this article, we present the case of a 79‐year‐old patient with myocardial rupture and following fatal outcome after iv thrombolysis applied for a stroke in the setting of emergent PCI for an unstable proximal RCA lesion.Perforation of the myocardium after iv thrombolysis presented a common challenge during the era of fibrinolytic therapies applied for myocardial infarction. Current guidelines regarding early causative therapy for ischemic stroke do not view a recent myocardial infarction as a contraindication for iv thrombolysis. The concomitant occurrence of myocardial infarction and ischemic stroke poses a challenge to the treating Heart and Brain team. Due to the high mortality of myocardial rupture when iv lysis is given later than 12‐24 hours after an acute MI, the interdisciplinary discussion should consider mechanical thrombectomy instead of lysis when feasible.2A 79‐year‐old male patient with no prior cardiac diagnosis presented to a local hospital on the 14th January due to acute onset of angina pectoris after physical exertion , and known localized cancer of the prostate (in remission) for which the patient had been treated with radiotherapy. Regarding cardiac risk factors, the patient presented with a known arterial hypertension and hypercholesterinemia. At time of admission, the patient was receiving statin treatment (simvastatin 40 mg/day), antihypertensive therapy  and was also taking a platelet aggregation inhibitor (aspirin 100 mg/day).2, thus a highly significant stenosis. Due to LM involvement, the patient was transferred to our tertiary center on the 14th January for further evaluation and possible subacute coronary artery bypass graft (CABG). The use of cardiopulmonary bypass machine was deemed possible despite the previously described, albeit distant intracerebral hemorrhage, however, with increased risk of bleeding complications.Acute coronary angiography was carried out via a radial approach, in which a three vessel coronary artery disease (CAD) was documented due to stenoses of the Left main (LM), proximal Left anterior descending (LAD), and mid‐Right coronary artery (RCA). An intravascular ultrasound (IVUS) study of the LM showed a minimum lumen area (MLA) of 3.1 mmDuring continuous monitoring, a nonsustained ventricular tachycardia for 30 seconds was registered. Blood gas analysis showed a transient elevation of lactate. The next morning, the hsTnT rose further from 201 at admission to 423 pg/mL and elevated creatine kinase (CK) was detected (276 U/L) with a relevant (18%) proportion of CK‐MB. Due to hemodynamic destabilization with progressive hypotension, immediately an ad hoc heart team reversed the strategy with the recommendation to intervene with PCI targeting the unstable RCA lesion. In addition to the existing aspirin therapy, loading with a P2Y12 inhibitor occurred with 600 mg of clopidogrel. On the 15th, the RCA as culprit lesion  was carried out; initial echocardiography showed no pericardial effusion. No return of spontaneous circulation (ROSC) could be achieved. After 30 minutes with no ROSC, CPR was terminated.After transfer to the monitoring ward one hour after the PCI, the patient developed aphasia without any motor deficit. Initial neurological assessment showed a score of 7 on the National Institutes of Health Stroke Scale (NIHSS) and a score of 0 on the modified Rankin Scale (mRS). Further imaging studies were initiated immediately due to the working diagnosis of a peri‐interventional stroke. In the cranial computed tomography (cCT) and angiography (CT‐A), no acute intracerebral hemorrhage, ischemic tissue demarcation, or occluded cerebral artery was registered. Because of the persisting symptoms, a magnetic resonance imaging of the neurocranium (cMRI) with diffusion‐weighted imaging (DWI) was carried out. Due to the demarcation of the left precentral gyrus in the DWI analysis Figure , persistIn the autopsy, the pathologists were able to detect a rupture of the free wall of the right ventricle with acute myocardial hemorrhage and pericardial effusion with 300 mL of serosanginous fluid. The implanted stents showed no obstruction. Studies of the cerebral tissue were able to document no intracerebral bleeding.3The incidence of myocardial rupture with subsequent cardiac tamponade and fatal outcome after intravenous fibrinolytic therapy for MI has been described previously. Before the advent of coronary interventions, iv thrombolysis was the only feasible option to treat patients with an acute MI. Patel et al describe an incidence of 0.85% myocardial perforations in a cohort of more than 100 000 patients treated with iv thrombolysis after ST‐segment elevation myocardial infarction (STEMI).The concomitant appearance of MI and stroke remains a dilemma facing the interdisciplinary Heart and Brain team of cardiologists and neurologists. In 2.3% of all patients with an MI, an ischemic stroke can be documented in the first month after myocardial infarction, which in comparison to a control cohort is a 44‐fold increased risk for stroke.In the updated version of the American Heart Association/American Stroke Association (AHA/ASA) stroke guidelines, in patients with recent MI an iv lysis is considered reasonable.Since the incidence of catheter‐associated strokes is rising due to the number cardiac catheterizations being carried out, this treatment dilemma will become a more frequent challenge.All considered, myocardial rupture after iv thrombolysis is a rare, but often fatal consequence. For patients with MI and ischemic stroke within a short timeframe, no concrete recommendations can be given. A catheter‐based therapy to retrieve emboli that seem to be causative for stroke after coronary angiography poses a valid treatment option and requires further investigation.None declared.BB and PC: involved in conception, literature research, and writing of the case report. CW, DLR, and SB: involved in research and critical revision of the manuscript.Written and informed consent was obtained from the patients next of kin in regard to publication of this article.