Early detection of myocardial involvement can be relevant in coronavirus disease 2019 (COVID-19) patients to timely target symptomatic treatment and decrease the occurrence of the cardiac sequelae of the infection.The aim of the present study was to assess the clinical value of cardiovascular magnetic resonance (CMR) in characterizing myocardial damage in active COVID-19 patients, through the correlation between qualitative and quantitative imaging biomarkers with clinical and laboratory evidence of myocardial injury.In this retrospective observational cohort study, we enrolled 27 patients with diagnosis of active COVID-19 and suspected cardiac involvement, referred to our institution for CMR between March 2020 and January 2021.Clinical and laboratory characteristics, including high sensitivity troponin T (hs-cTnT), and CMR imaging data were obtained. Relationships between CMR parameters, clinical and laboratory findings were explored.Comparisons were made with age-, sex- and risk factor–matched control group of 27 individuals, including healthy controls and patients without other signs or history of myocardial disease, who underwent CMR examination between January 2020 and January 2021.The median (IQR) time interval between COVID-19 diagnosis and CMR examination was 20 (13.5–31.5) days. Hs-cTnT values were collected within 24 h prior to CMR and resulted abnormally increased in 18 patients (66.6%). A total of 20 cases (74%) presented tissue signal abnormalities, including increased myocardial native T1 (n = 11), myocardial T2 (n = 14) and extracellular volume fraction (ECV) (n = 10), late gadolinium enhancement (LGE) (n = 12) or pericardial enhancement (n = 2). A CMR diagnosis of myocarditis was established in 9 (33.3%), pericarditis in 2 (7.4%) and myocardial infarction with non-obstructive coronary arteries in 3 (11.11%) patients. T2 mapping values showed a moderate positive linear correlation with Hs-cTnT . A high degree positive linear correlation between ECV and Hs-cTnT was also found .CMR allows in vivo recognition and characterization of myocardial damage in a cohort of selected COVID-19 individuals by means of a multiparametric scanning protocol including conventional imaging and T1–T2 mapping sequences. Abnormal T2 mapping was the most commonly abnormality observed in our cohort and positively correlated with hs-cTnT values, reflecting the predominant edematous changes characterizing the active phase of disease.The online version contains supplementary material available at 10.1186/s12968-021-00764-x. Myocardial injury is not uncommon in coronavirus disease 19 (COVID-19) and recognizes a complex multifactorial pathogenesis, including direct viral toxicity, uncontrolled immune-activation , stress cardiomyopathy, mismatch ischemia and prothrombotic activation with plaques formation and microvascular disease . By defiThis heterogeneity probably relies on the different thresholds applied in cTnT assays as well as on the clinical severity of the cases included, which has shown to be significantly related with the extent of myocardial damage. A recent metanalysis summarized that acute myocardial injury rate was 13-fold higher in intensive care unit patients as compared to those with mild forms of infection . NeverthWhilst it remains debated whether cTnT rise necessarily reflects direct cardiac infection, there is a clear independent association between myocardial injury and mortality rate. Fatal outcomes were reported in 37.5% in patients with elevated levels of cTnT and raised to nearly 70% in the presence of pre-existing CVD comorbidities  and out-Despite the evidence, the American College of Cardiology does not recommend routine testing of cTnT levels in COVID-19 patients, unless the diagnosis of acute myocardial infarction (MI) is suspected on clinical grounds .This potentially excludes from the screening most of pauci- or asymptomatic patients in which myocardial injury yields prognostic significance.In this scenario, early detection of myocardial involvement can be relevant to timely target symptomatic treatment and decrease the occurrence of the devastating cardiac sequelae of the infection.Our endpoint is to assess the clinical value of a non-invasive and highly sensitive tool like cardiovascular magnetic resonance (CMR) in characterizing myocardial damage in COVID-19 patients, through the correlation of qualitative and quantitative CMR features with clinical and laboratory evidence of myocardial injury. Our quantitative analysis relies on the use of parametric mapping, which is an innovative and reproducible method to provide unique quantitative data about T1 and T2 relaxation time changes in the myocardium.As a further advantage, CMR may by integrated by the comprehensive assessment of heart, pulmonary vessels and lung parenchyma in a one-stop-shop approach, which makes it potentially suitable to contemporary exclude thromboembolic complications and follow-up pulmonary disease progression in COVID-19 patients, using a radiation-fee imaging procedure .This was a single-center observational retrospective study on a cohort of 27 patients with a confirmed active COVID-19 by reverse transcription–polymerase chain reaction (RT-PCR) on nasopharyngeal swabs.at least one high sensitive Troponin T (hs-cTnT) measurement above 99th percentile (> 0.014 ng/ml) in absence of ST-elevation or other signs of MI;newly observed reduced (< 50%) left ventricle (LV) ejection fraction (LVEF) detected by a point of care bedside echocardiography;no obstructive coronary artery disease (CAD) at coronary angiography despite infarct-like clinical presentation.Patients were considered eligible if RT-PCR nasopharyngeal swab, collected within 72 h prior to CMR scan, confirmed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, COVID-19 related clinical signs and symptoms were persistent and at least one the following inclusion criteria was fulfilled:Exclusion criteria were general contraindications to CMR, unstable clinical conditions or inability to perform repeated breath-holds.2/FiO2) ratio.In all patients, the following routine blood tests and arterial blood gas tests were collected in the 48 h before the CMR examination: C-reactive protein (CRP), D-dimer, white blood cells (WBC) count, lymphocyte count, and arterial oxygen partial pressure/fraction of inspired oxygen  were considered abnormal. Data on respiratory rate and oxygen saturation were also reported.Disease severity score was evaluated for all individuals, using the Chinese Center of Disease Control (CDC) criteria : mild, sBased on computed tomography (CT) scoring by Pan et al. , a semi-As explained elsewhere, chest CT scans were performed on two multidetector scanners . Acquisition parameters were settled according to manufacturers’ recommendations for standard chest CT examination and images reconstruction was performed with filtered back-projection at 1.00 mm thickness, with a B20 kernel for soft tissue and a B60 kernel for pulmonary parenchyma .A group of 27 individuals, including healthy controls and patients without clinical signs or history of myocardial disease, who underwent a CMR between January 2020 and January 2021, was also enrolled. All patients were matched for age, sex and cardiovascular risk factors  and performed the same CMR protocol on the same scanner as for COVID patients.All CMR examinations were performed between March 2020 and January 2021 on a fully dedicated COVID-19 1.5 T CMR scanner  equipped with SQ-engine gradients  and a 16-channel phased-array cardiac coil. According to international recommendations  of 0.029 (0.007–0.057) ng/ml.12, all patients were classified to have a “late phase” disease (> 7 days); in addition, the median time between SARS-CoV-2 infection diagnosis and CMR was 20 (13.5–31.5) days.Depending on timing of symptoms’ onset9/l) in 7/27 (25.9%) patients.Increased D-dimer (> 500 ng/ml) was found in 21/27 (77.8%) patients with a median value (IQR) of 750 (580–1823) ng/ml and increased CRP (> 0.5 mg/dl) was present in 19/27 (70.4%) patients with a median value (IQR) of 1.4 (0.43–3.93) mg/dl. WBC count was elevated (> 11.3 × 109/l) was observed in 11/27 (40.7%) patients, decreased O2 saturation (≤ 95%) in 5/27 (18.5%), and decreased PaO2/FiO2 ratio (< 300) in 4/27 (14.8%) patients.Decreased lymphocyte count , corresponding to a mean global CT score of 9.6 ± 4.8 (range 0–15). Mean time interval between chest CT scan and CMR was 7.3 ± 3.8 days.Underlying CVD comorbidities were present in 12/27 patients (37%), including hypertension  and type II diabetes , dyslipidemia . No patients had known previous CVD or history of obstructive CAD.No procedural complications were observed in any patient or control, even though four patients interrupted the exam, renouncing the post-contrast MOLLI images acquisition.CMR features are displayed in Table LV dilation  was found in 5/27 (18.5%) patients, whereas 13/27 (48.15%) patients showed impaired LV systolic function (LVEF < 50%) and 13/27 had regional wall motion abnormalities consisting in hypo- or akinesia (the number of LV segments involved was 1–3 in four patients and 4–7 in nine patients), with a predominant involvement of mid anteroseptal and mid anterior wall segment.A total of 20/27 (74.1) patients showed myocardial tissue signal abnormalities, including at least one of the following: increased myocardial native T1 (11/27), T2 (14/27) and ECV (10/23), areas of LGE (12/27) or edema on STIR images (10/27), or pericardial enhancement (2/27).Nine patients had a non-ischemic-type LGE pattern, while 3 showed transmural LGE. Among cases with non-ischemic LGE pattern, eight patients showed a subepicardial and midwall distribution, whereas midwall LGE area was present in one patient; the LV wall regions involved were septum in 4/9 patients, anterior wall in 1/9, anterolateral wall in 3/9 patients, inferolateral wall in 6/9 and inferior wall in 4/9 patients.Isolated elevation of one myocardial parametric value was found in 7/27 patients (3/27 patients with increased native T1 and 4/27 with increased T2).Final diagnosis of myocarditis was established in 9/27 (33.3%), pericarditis in 2/27 (7.4%) and myocardial infarction with non-obstructive coronary arteries (MINOCA) in 3/27 11.1%) patients , when these variables were considered as categorical. The correlation analysis using the Pearson Correlation index showed a moderate positive linear relationship between the T2 and the hs-cTnT values .A significant dependence between T2 and the hs-cTnT values was found using the Pearson’s X2 0.59 and a r 0.77.The linear regression analysis was conducted to examine the relationship between the T2 and hs-cTnT values. In particular the best fitting model was obtained after adding 4 parameters, hs-cTnT, WBC count, lymphocytes count, CRP value, with a rNo significant correlation was found between the native T1 and the hs-cTnT, both considering as categorical or quantitative variables (p 0.78–0.13).2 test (p 0.029) and a high degree positive correlation between these two parameters, using the Pearson Correlation Index .All patients with an increased ECV had an elevated hs-cTnT; with a significant dependence between the altered ECV and hs-cTnT using the Pearson XROC analysis identified a hs-cTnT value > 0.0215 ng/ml as the best threshold for distinction between the healthy controls and patients with myocardial edema  and ECV (p 0.025). No significant differences were found in native T1 , T2 (p 0.428) and ECV (p 0.807).To the best of our knowledge, this is the first study using CMR to explore the nature of myocardial damage in a selected cohort of individuals with active phase of COVID-19 disease.Besides the definition of myocardial injury, corresponding to the rise of cTnT levels above the 99th percentile upper reference limit , our resThe heterogeneity of CMR patterns observed in our population reflects the complexity of disease pathogenesis, offering insight into the predominant mechanisms involved , with potential implications on patient management and prognosis .As expected, the most common CMR disease pattern we observed (33.3% of patients) was consistent with a diagnosis of clinically suspected acute myocarditis, characterized by the combination of myocardial edema with evidence of non-ischemic myocardial injury, as suggested by the new CMR diagnostic criteria .CMR pattern of diffuse inflammation was more commonly detected, probably reflecting the increased interstitial macrophage recruitment and, in a limited percentage of cases, the multifocal lymphocytic infiltration, reported in previous pathological studies .Coherently with the acute phase of COVID-19-related myocardial involvement characterizing our study population, the most common CMR feature observed was the increase of myocardial T2 (14/27 patients), mirroring prior studies in non-COVID-19 literature and reflecting the predominant edematous expression of the process .Our average T2 values (52.2 ms) significantly differed from an earlier publication by Esposito et al. , reportiSimilar observations were published by Puntmann et al. , who fouIsolated increase of native T1 values (3/27 patients), as a non-specific expression of interstitial fibrosis of unknown origin and potentially pre-existing to the SARS-CoV-2 infection, was considered non-necessarily attributable to COVID-19 related myocardial injury.Among our COVID-19 subjects, the main differential diagnoses of acute myocarditis was pericarditis and myocardial infarction with non-obstructive coronary arteries (MINOCA). Pericardial inflammation in COVID-19 has been described as a purely inflammatory response to the systemic insult rather than a local infectious process . AccordiPossible explanations have been attributed to the presence of high levels of angiotensin converting enzyme 2 receptors in pericytes and endothelial cells, determining severe microvascular dysfunction enhanced by the cytokines storm . MismatcA further distinguished pathological CMR pattern (observed in 4 cases) showed an isolated increase of T2 without associated CMR features of myocardial damage on LGE, native T1 and ECV. This CMR phenotype, not corresponding to a diagnosis of acute myocarditis, likely represents the imaging correlate of the diffuse edematous myocardial involvement induced by the uncontrolled cytokine release, characterizing the late-phases of the infection. Presence of myocardial interstitial macrophage infiltration without myocyte injury was reported in 86% of cases from a recent multicenter study including the autopsies of 21 consecutive COVID-19 patients . The autSimilar findings were published by Xu et al.  reportin2: 0.75; p < 0.001).Systemic Capillary Leak Syndrome has been advocated as a possible mechanism of pathogenesis in COVID-19 patients . It causCMR follow-up data are missing in literature with these regards and will certainly provide better comprehension of the underlying mechanism of injury and its transient nature with or without permanent tissue abnormalities.Interestingly, two patients showed newly diagnosed mild reduction of biventricular systolic function in absence of myocardial signal alterations both in conventional sequences (T2 STIR and LGE) and in relaxometric imaging. Interpretation of this finding remains questionable as a pre-existent ventricular impairment cannot be excluded and could not be differentiated from a chronic evolution of myocardial damage.We found a positive linear relationship between the T2 mapping and hs-cTnT values.Besides myocardial necrosis, hs-cTnT assays can be detectable in COVID-19 as the consequence of the transient ischemic or inflammatory conditions associated with the disease, including respiratory and renal failure, hypoxemia, tachyarrhythmias and thrombo-embolic disease .Regardless of the underlying pathogenetic mechanisms, troponin levels have been hypothesized to be the expression of a general hyperinflammatory status . At the Coherently, the stepwise linear regression analysis showed that the increase of each of the inflammatory biomarkers  values, was associated with higher T2; these results confirm that patients with myocardial injury showed a more pronounced inflammatory response with direct impact on T2 measurements.ROC analysis indicated that a hs-cTnT value > 0.0215 ng/ml would define the best cut-off value to differentiate normal versus increased T2 . This threshold identified 13/14 patients as having evidence of myocardial pathological involvement on T2 and may represent a potential cut-off value to direct patients to CMR examination.A further interesting finding is the non-correlation between the extent of pulmonary disease and occurrence of myocardial involvement, which further supports the theory that cardiac involvement is not affected or a complication of pulmonary pathology, even though they likely share common pathogenic mechanisms.active COVID-19.Our sample size was modest, but nonetheless represents, up to date, one of the largest cohorts of patients who have undergone CMR with Because of the complexity of the diagnostic exam, requiring repeated breath holds and an average scanning time of 50 min, we had a selection bias regarding the clinical stage of patients enrolled, almost all presenting with a mild form of disease.Myocardial injury proportionally correlated with the severity of the clinical manifestations of COVID-19 and potentially identifies patients with worse baseline clinical status. This has an obvious impact on the prevalence and extent of tissue damage observed in our study.Similarly, no early phase disease patients  were included in our cohort. This indirectly confirms that myocardial involvement begins or persists days after disease onset.Our study did not provide prognostic data, which are likely to further refine the role of CMR imaging in this complex clinical scenario and to better elucidate possible inclusion criteria for patient’s selection.None of our patients performed an endomyocardial biopsy, therefore the diagnosis of myocarditis in our patients should be considered as not definitively confirmed.Finally, we did not include asymptomatic COVID-19 individuals, in which multiparametric imaging data would allow subclinical detection of structural damage with potentially relevant implications on early diagnosis and clinical decision-making.CMR allows recognition and characterization of myocardial damage in a cohort of selected COVID-19 individuals with active disease. This consists in heterogeneous patterns of injury ranging from acute myocarditis, to MINOCA, pericarditis, and CMR evidence of isolated edematous changes. The prompt recognition of pattern disease is pivotal to drive therapy and patient’s management. Myocardial T2 appears to be the prevalent CMR imaging biomarker in active COVID-19 patients and the best predictor of myocardial injury.Additional file 1: Table S1. CMR scanning protocol parameters in detail.


Hypertension plays a dominant role in the development of left ventricular (LV) remodeling and heart failure, in addition to being the main risk factor for coronary artery disease. In this review, we focus on the focal geometric and functional tissue aspects of the LV septal base, since basal septal hypertrophy (BSH), as the early imaging biomarker of LV remodeling due to hypertensive heart disease, is detected in cross-sectional clinic studies. In addition, the validation of BSH by animal studies using third generation microimaging and relevant clinical observations are also discussed in the report. Finally, an evaluation of both human and animal quantitative imaging studies and the importance of combined cardiac imaging methods and stress-induction in the separation of adaptive and maladaptive phases of the LV remodeling are pointed out. As a result, BSH, as the early imaging biomarker and quantitative follow-up of functional analysis in hypertension, could possibly contribute to early treatment in a timely fashion in the prevention of hypertensive disease progression to heart failure. A variety of stress stimuli in etiopathogenesis and the difficulty of diagnosing pure hemodynamic overload mediated BSH lead to an absence of the certain prevalence of this particular finding in the population. Heart failure (HF) is a progressive process and associated with the left ventricular (LV) remodeling . HyperteIt is known that LV segmental remodeling could be the case in early-stage hypertension described as borderline hypertension, and the septal wall is speculated to be the first LV wall to become hypertrophic in clinical practice according to cross-sectional studies ,11,12. IOur earlier reports showed a high heart rate–pressure product and high LV outflow tract blood flow as well as basal septal hyperdynamic tissue under stress in BSH patients with hypertension ,18,19,20In the animal validation studies, we also detected a remarkable velocity increment in the early time points after transaortic construction in pathologic stress, which was consistent with the findings in cross-sectional clinic studies . HoweverBP monitorization under stress is insufficient in patients with LV septal hypertrophy during clinical practice. In fact, no attempt was made for BP recording during exercise even in the comprehensively planned CMR study in which physiologic exercise in a healthy population was speculated to be the reason for septal hypertrophy . We emphBSH plays a role as a common denominator for variety of stress stimuli beyond basic hemodynamic stress-mediated LV remodeling. Due to this complex nature of BSH, we previously described this morphological finding as the ‘stressed heart morphology’ (SHM), which was documented as an imaging figure in both acute and chronic stress-mediated heart conditions such as acute stress cardiomyopathy  and hypeIn fact, while a smooth curvature is the case in some patients , anotherSympathetic overdrive more dominantly involves the LV base compared to the midapical myocardial segments. However, despite the fact that it is a potentially possible etiologic mechanism, there has not been any chronic mental or emotional disorder documented or described except acute stress cardiomyopathy in the development of SHM. Heterogeneity involved in remodeling in humans produces a dilemma not only in a certain diagnosis of pure hemodynamic overload-mediated BSH in hypertensive patients, but in the certain determination of BSH prevalence in the population.On the other hand, years later, we have determined LV segmental remodeling in an animal model that is associated with a very regular distribution a,b underHF with preserved ejection fraction (HFPEF) is a relatively new clinical entity and the number of HFPEF patients is increasing worldwide. As the majority of the HFPEF patients have systemic hypertension, HFPEF is usually accepted as a hypertension-mediated phenomenon. The preserved global systolic function has been documented by freehand three-dimensional echocardiography at rest in HFPEFpatients . HoweverThe efficacy of preventive measures to slow the progression of hypertensive heart disease supports the concept that hypertrophy is possibly a reversible stage of LV remodeling . HoweverHypertension-mediated HF describes a mild LV remodeling mostly observed in female patients . It is eIt has been mentioned that myocardial fibrosis is associated with diastolic HF in hypertensive heart disease . The accAge-related myocardial fibrosis may adversely affect the contractile reserve, especially in elderly hypertensive individuals. Recent data have shown that contractility may become blunted by stress induction in patients older than 70 . InteresThe collected data showed that blood pressure control is an essential step in patients with hypertension-mediated consequences leading to vascular mortality ,51. The According to another comprehensive study from the Johns Hopkins Medical Institutions, HFPEF patients who are on an effective medical therapeutic regimen (68% use of an ACE inhibitor) have similar endocardial and mid-wall fractional shortening compared to that of hypertensive LVH . This prAtrial fibrillation (AF) is an important problem during follow-up in hypertensive patients. HFPEF with AF is associated with a greater mean LV mass compared to systolic HF patients with AF, which supports the special importance of long-standing hypertension as the risk factor in HFPEF patients . It may As a result, since LV remodeling leading to HF is a progressive process, an LV segmental evaluation seems important, not only in the determination of BSH as the early imaging biomarker, but in a severity assessment of LV myocardial regions in advance hypertensive cases with global LV remodeling. It is documented in the literature that precise patientmonitorization using novel cardiac imaging can successfully contribute to differentiating the stages of disease course and successful, widespread long-term medical management.


A 2-group PLS-Discriminant Analysis was performed to verify whether the two HCM groups differed also based on the metabolic fingerprint. A clear clustering was shown (ANOVA p = 0.014). The most discriminating metabolites resulted as follows: in the NOHCM Group, there were higher levels of threitol, aminomalonic acid, and sucrose, while the OHCM Group presented higher levels of amino acids, in particular those branched chains, of intermediates of glycolysis (lactate) and the Krebs cycle , of fatty acids , of ketone bodies (2-OH-butyrate). Our data point out a different systolic function related to a specific metabolic activity in the two HCM phenotypic forms, with specific metabolites associated with better contractility in OHCM.The purpose of this study was to assess whether metabolomics, associated with echocardiography, was able to highlight pathophysiological differences between obstructive (OHCM) or non-obstructive (NOHCM) hypertrophic cardiomyopathy. Thirty-one HCM patients underwent standard and advanced echocardiography; a plasma sample was collected for metabolomic analysis. Results. Patients with OHCM compared with subjects with NOHCM had higher values of 2DLVEF (66.5 ± 3.3% vs. 60.6 ± 1.8%,  Hypertrophic cardiomyopathy (HCM) is the most common of cardiovascular pathologies with the genetic transmission. It is caused by a variety of mutations, mainly concerning genes encoding cardiac sarcomeric proteins, and is characterized by rather heterogeneous clinical expression, with individual pathophysiology and natural history .2+ of myofilaments. The result is an altered energy metabolism of cardiomyocytes, with an increase in ATP consumption by the cellular contractile apparatus, but with reduced efficiency of electro-mechanical coupling [In vitro studies and animal models have shown that some sarcomeric HCM mutations are associated with subclinical impairment in contractile function and excessive sensitivity to CaMetabolomics is the “omics” science that aims to analyze the profile of small molecules in a biological organism. Reflecting the exact endpoints of biological events determined by the sum of genes, RNA, proteins, and environmental factors, metabolomics provides an overall and instantaneous functional view of an organism . TherefoIn this study we used gas chromatography-mass spectrometry (GC-MS)-based metabolomic analysis associated with standard and advanced echocardiography to a) highlight the metabolic specifications of the HCM, and b) distinguish and characterize the energetic substrate of both the obstructive (OHCM) and non-obstructive (NOHCM) phenotype.The anthropometric and clinical data of the study population are shown in We found no significant differences between the two groups regarding comorbidities or cardiovascular risk factors .The risk of sudden 5-year cardiac death calculated with the HCM-Sudden Cardiac Death (SCD) score  had a mep < 0.05). Patients with OHCM showed higher weight  and, consequently, BMI (27.9 ± 2.0 vs. 23.5 ± 3.6 kg/m2. p < 0.01) and BSA . The pharmacologic treatment did not show significant differences between the two groups.According to the literature, the HCM-SCD was higher in the OHCM group than in the NOHCM group , dilatation of the left atrium (Left Atrium Volume Indexed (LAVI): 52.8 ± 20.2 mL/m2), a normal LV Ejection Fraction (LVEF: 63.6 ± 5.2%) and reduction of the LV longitudinal function, assessed both by Speckle Tracking Echocardiography  2D: 14.0 ± 3.6%); 3D-GLS values were comparable to those measured with two-dimensional evaluation .Standard and advanced echocardiography. The whole population study showed the typical characteristics of the underlying disease, with a marked increase in the Inter-Ventricular Septum (IVS) thickness (18.7 ± 3.8 mm) and of the Left Ventricular (LV) mass measured with 3D echocardiography, both absolute (173 ± 39 g) and indexed (105.5 ± 20.2 g/mp < 0.01), S wave  and 3D GLS .Metabolomics. To trace metabolic alterations correlated to myocardial contractility, we applied a regression analysis  between the metabolomic data (X variables) of the whole population and GLS wave (Y variable), used as a landmark continuous variable of systolic function, obtaining an excellent correlation (R2 (R2x = 0.889) and Q2 (0.622); the good quality of the model was also confirmed by the permutation test and cross-validated by ANOVA (p = 0.01).The hypothesis that the 2 groups differed significantly on the basis of the phenotypic characteristics of HCM, specifically the obstructive and non-obstructive subtype, was subsequently tested; for this purpose, a 2-group PLS-Discriminant Analysis (PLS-DA) was performed, showing a clear clustering , Panel AThe results of PLS-DA have also been reported as a loading score, which allows highlighting the weight (importance) that the variables have in separating the a priori identified classes. It is interesting to observe how it is possible to identify, even visually, a more intense metabolic activity in the OHCM group compared to that of the non-obstructive form , Panel B2y = 0.947; Q2 = 0.727), rather than a worsening, as would have been expected if the clustering was due to anthropometric differences.To rule out that the observed separation was due to differences in nutritional status, (a) the existence of a gradient along the Principal Component (PC) 1 or PC2 that followed the distribution of body weight or BMI was excluded; (b) a PLS-DA was carried out on two subgroups comparable by weight and BMI, obtaining an improvement of the model  allowed us to identify the most discriminating and expressed metabolites in the two classes . In the The present study was aimed at assessing whether metabolomics, in combination with standard and advanced echocardiographic data, was able to highlight metabolic differences underlying diverse phenotypes of HCM. The results showed: a. an excellent correlation between the contractile state, documented by S wave and GLS measurement, and the metabolomic profile in the entire population; b. a better-preserved systolic function, as demonstrated by the 2D and 3D parameters, in patients with OHCM, compared to those with NOHCM; c. a clear metabolomic clustering of OHCM and NOHCM. In this context, the most discriminating and expressed metabolites were threitol, aminomalonic acid, and sucrose in the NOHCM group, and amino acids, in particular BCAA, lactate, fumarate, succinate, citrate, fatty acids, and ketone bodies, in the OHCM group.The alteration in energy metabolism in HCM was widely documented, by a lower value of the ratio between cardiac phosphocreatine (PCr) and adenosine triphosphate (ATP) , regardlIn recent years, metabolomics was successfully adopted to decipher the biomolecular substrate of this energy dysfunction. A mouse knock-in for the MYBPC3 gene HCM model was treated with perhexiline, a metabolic drug known to improve the myocardial production of energy. Six weeks of treatment resulted in a partial improvement in the anterior wall thickness and ventricular mass, accompanied by a substantial change of 272 metabolites, mostly involved in the cardiac energy metabolism .In humans, an integrated echocardiographic and metabolomic approach was used to study 34 HCM subjects carrying the MYBPC3-Q1061X mutation, 19 mutation carriers without hypertrophy, and 20 relatives without both mutation and hypertrophy. This study found that concentrations of BCAAs, triglycerides, and ether-phospholipids were higher in HCM-bearing mutation than in controls and carriers of non-hypertrophic mutations. Furthermore, these molecules correlated with both LV hypertrophy and systo-diastolic dysfunction, suggesting their potential remodeling function in HCM [The correlation between metabolome and cardiac function has been confirmed by our results. Indeed, the TDI-derived S wave and GLS, the more sensitive parameter of systolic performance, showed a strong correlation with the metabolic profile in both patient groups.On the other hand, S wave, a well-known longitudinal dysfunction parameter , even thThe evidence of a higher LV longitudinal function in OHCM patients seems to be a particularly relevant finding of this study: despite similar LV sizes and thicknesses in both HCM groups, it could suggest a different ratio between contractile and non-contractile tissue, as previously demonstrated using cardiac Magnetic Resonance Imaging (MRI) and Positron Emission Tomography (PET) .In line with this evidence, the metabolomic analysis revealed a clear difference between the two HCM subpopulations, demonstrating metabolic hyperactivity in the OHCM group, characterized by a relative hypercontractility. These findings could be explained by a relatively greater amount of metabolically active cardiomyocytes and a lesser presence of fibrotic tissue, as found by Potios et al., who identified a higher prevalence of large late gadolinium enhancement (LGE) burden on MRI and microvascular ischemia by PET in NOHCM patients . This evAnalyzing the VIP obtained through multivariate analysis, we found that the BCAAs in the OHCM group were the most represented molecules, in accordance with previous investigations on hypertrophied rat hearts  and HCM Finally, in the OHCM patients the identification of increased levels of arachidonic acid and palmitoleic acid, both associated with an increased risk of ventricular arrhythmias ,19,20, iAmong the molecules found to have increased in patients with NOHCM, the aminomalonic acid and threitol have recently been identified as biomarkers of pulmonary hypertension; these findings seem to be in line with the higher values of PAPS observed in our study in this group of patients.  For its Threitol, an end-product of the xylose metabolism, is linked to the pentose-phosphate pathway through the glucuronate cycle . Notewor2+ and disrupting the homeostasis of this ion at the sarcoplasmic level [Sucrose, which we also found increased in patients with NOHCM, is a molecule capable of reducing the sensitivity of contractile myofilaments to Caic level , with reic level .The whole data identify a different systolic function related to a specific metabolic activity in the two HCM phenotypic forms, with better contractility in OHCM associated with a more intense metabolism, and this finding seems to confirm previous evidence of a greater amount of fibrosis in NOHCM patients (lower number of active cardiomyocytes -> lower metabolic activity -> worse systolic function), thus helping to explain the differences observed in the two forms of the same pathology.Our study presents some limitations: the small sample size and the absence of a direct measurement of the fibrosis degree; however, 15 subjects per arm is dimed sufficient for a pilot study  and prevMoreover, our work does not pretend to be conclusive or exhaustive; rather, it would be a hypothesis-generating study and would stimulate further research on this topic in the view of a deeper knowledge of the correlation between pathophysiology and clinical phenotype of HCM.Thirty-one patients  affected by HCM, diagnosed according to the criteria established by the relating 2014 European guidelines , were enPatients were divided into two groups, based on the presence (OHCM) or absence (NOHCM) of left ventricular outflow tract (LVOT) obstruction. As indicated in the ESC HCM Guidelines , LVOT obOn this basis, 15 patients presented OHCM, and 16 a NOHCM phenotype . This sample size resulted adequate to assure the minimum precision requested for a pilot study based on the work of Julious SA , which eThe study was approved by the Ethics Committee of the “Azienda Ospedaliero-Universitaria di Cagliari" and was performed in accordance with the Declaration of Helsinki. Participants were informed of the purpose and methodology of the study and their written consent was obtained prior to inclusion. All patients underwent clinical evaluation, 12-lead electrocardiogram, echocardiography with the evaluation of the standard systo-diastolic parameters, and Global Longitudinal Strain (GLS) by both 2D and 3D speckle tracking echocardiography (STE). In the morning time (08–13), after a fasting night, a 4 mL heparinized venous blood sample was collected from all patients for metabolomic analysis. The risk of sudden 5-year cardiac death was calculated by the HCM-SCD score .Standard echocardiography. A complete mono- and two-dimensional and Color Doppler echocardiographic evaluation was performed. The volumes and ventricular thicknesses were measured, and the left ventricle ejection fraction (LVEF) was calculated using the Simpson’s biplane method from the apical projection of four and two chambers; a value ≤ 50% has been considered abnormal. The diastolic function was evaluated by recording the velocities with the pulsed Doppler in the four apical chambers view. Using Tissue Doppler imaging (TDI), we evaluated LV longitudinal function by measuring the velocities of the mitral valve annulus. Myocardial velocity patterns were obtained from the average values measured by placing the sample volume in the basal portion of the interventricular septum (IVS) and lateral wall. Peak systolic velocity (S wave), peak velocity in proto-diastole (E’ wave), peak velocity of atrial contraction (A’ wave), and LV isovolumetric relaxation time (IVRT) were measured [measured ,33. S waLVOT gradient was measured during a rest echocoardiography.Raw data were also acquired using the STE technique.3D echocardiography. The “full-volume” 3D data sets were acquired using a complete matrix array transducer , positioning the probe in apical view and performing the acquisition during a brief breath interruption.To ensure the inclusion of the entire LV within the pyramidal volume scanning and a valuable volume rate, the data sets were acquired using the "wide-angle" mode associated with a multi-beat triggered acquisition (3–4 beats) under electrocardiographic trace and during a 5–7 s apnea .2D STE. The 2D STE analysis was performed using software  able to perform the analysis independently of the equipment used for the acquisition, to obtain absolute results and not vendor-specific. GLS was measured by performing a manual tracking of the endocardial border in the 3 apical views.After STE analysis of the “frame-by-frame” LV endocardium during a cardiac cycle, the software provides the regional strain curves in each view, from which the average peak strain value is determined. The adequacy of the tracking was visually verified and, in case of tracing considered not optimal, a manual correction of the endocardial border was performed . If the 3D STE. 3D volumetric analysis and 3D strain measurements of the LV were performed using the 3D-STE. The “full volume” 3D data sets were analyzed using the same software as in the 2D analysis but with a specific application .The papillary muscles were included in the cavity of the left ventricle. Manual adjustments of the endocardial surface were performed when needed.The software performed 3D tracking analysis throughout the cardiac cycle and LV mass was also determined.For the 3D strain analysis, the software provided an average longitudinal strain from which the peak GLS was determined. As the software did not provide an automatic evaluation of the adequacy of the images, the accuracy of the tracking was visually assessed on the 2D images extracted from the 3D data sets. When the data was considered inadequate after manual correction the subjects were excluded from the analysis ; GLS wasMetabolomic analysis. Specimens of all study participants were centrifuged at 2000 rpm for 10 min: the supernatant was transferred in Eppendorf tubes and stored at −80 °C until analysis. Plasma samples were thawed at room temperature. 100 μL of each sample were collected to form a pooled sample to use for quality control and to form an average composition sample to analyze among the others. 400 μL of plasma were treated with 1200 μL of cold methanol in 2 mL Eppendorf tubes, vortex mixed, and centrifuged 10 min at 14,000 rpm (16.9 G × 1000). 400 μL of the upper phase were transferred in glass vials (1.5 mL) and evaporated to dryness overnight in an Eppendorf vacuum centrifuge. 50 μL of a 0.24 M (20 mg/mL) solution of methoxylamine hydrochloride in pyridine was added to each vial, samples were vortex mixed and left to react for 17 h at room temperature in the dark. Then 50 μL of MSTFA (N-Methyl-N-trimethylsilyltrifluoroacetamide) were added and left to react for 1 h at room temperature. The derivatized samples were diluted with hexane (100 μL) with tetracosane (0.01 mg/mL) as internal standard, just before GC-MS analysis.Instrumental parameters. Samples were analyzed using an Agilent 5975C interfaced to the GC 7820 (new 5977B/7890B) equipped with a DB-5ms column (J & W), injector temperature at 230 °C, detector temperature at 280 °C, helium carrier gas flow rate of 1 mL/min. The GC oven temperature program was 90 °C initial temperature with 1 min hold time and ramping at 10 °C/min to a final temperature of 270 °C with 7 min hold time. One μL of the derivatized sample was injected in split (1:4) mode. After a solvent delay of 3 min, mass spectra were acquired in full scan mode using 2.28 scans/s with a mass range of 50–700 Amu .http://chemdata.nist.gov/mass-spc/amdis, accessed on 7 September 2021) that identified each peak by comparison of the relative mass spectra and the retention times with those stored in an in-house made library comprising 255 metabolites. Other metabolites were identified using NIST08  and the Golm Metabolome Database . Through this approach, 113 compounds were accurately identified, while 28 other metabolites were tentatively assigned relying on GMD and NIST libraries. AMDIS analysis produced an Excel datasheet that was successively subjected to chemometric analysis.Mass Spectral deconvolution. Each acquired chromatogram was analyzed by means of the free software AMDIS , an unsupervised method, the Partial Least Square Analysis (PLS), a supervised regression analysis method, and the Partial Least-Square Discriminant Analysis, (PLS-DA) which uses a Y-matrix containing information about the a priori class to which the sample belongs (NOHCM or OHCM) and which is used to evaluate the statistical significance of the classification [fication .p < 0.05 was considered statistically significant. The analyses were performed using IBM SPSS v. 25.Univariate statistical analysis. Continuous variables  were compared using the two-tailed T-test for non-paired samples, while the Fisher exact test was used for the categorical ones. A corrected value of two-tailed Overall, the echocardiographic differences between NOHCM and OHCM patients seem ascribable–not only but also–to a different type of hypertrophy present in the two phenotypic forms. In turn, the metabolomic profiles, different in the 2 groups of HCM patients, a. correlate with the echocardiographic parameters of contractile function, b. could be related to a different modulation of hypertrophy/fibrosis, and c. seem to identify two different metabolic states, dependent mainly on the respective energy efficiency.We indeed acknowledge that our results will have to be strengthened and extended with the enrollment of a larger number of patients. However, we consider the identification of specific metabolic signatures in the HCM subgroups and the related pathophysiological mechanisms as hopeful anticipation. Our data could help physicians to know better the pathophysiology and the clinical characteristics of their HCM patients. Moreover, the current results, although not conclusive, seem able to generate further research on this topic with an eminently translational approach that could lead to innovative, tailored therapies based on the identification of new molecular targets , able to modify the natural history of the pathology modulating the pathways involved in its progression.


Plasma and urinary cyclic guanosine monophosphate (cGMP) excretion levels were higher  with tadalafil plus SC BNP versus tadalafil alone. There was no improvement in renal response as measured by GFR, renal plasma flow, sodium excretion, and urine flow with tadalafil plus SC BNP compared to tadalafil alone. In subjects with PDD, tadalafil alone resulted in no improvement in cardiac adaptation, while tadalafil and SC BNP resulted in enhanced cardiac adaptation to VL.Preclinical diastolic dysfunction (PDD) results in impaired cardiorenal response to volume load (VL) which may contribute to the progression to clinical heart failure with preserved ejection fraction (HFpEF). The objective was to evaluate if phosphodiesterase V inhibition (PDEVI) alone or combination PDEVI plus B‐type natriuretic peptide (BNP) administration will correct the impaired cardiorenal response to VL in PDD. A randomized double‐blinded placebo‐controlled cross‐over study was conducted in 20 subjects with PDD, defined as left ventricular ejection fraction (LVEF) >50% with moderate or severe diastolic dysfunction by Doppler echocardiography and without HF diagnosis or symptoms. Effects of PDEVI with oral tadalafil alone and tadalafil plus subcutaneous (SC) BNP, administered prior to acute volume loading, were assessed. Tadalafil alone did not result in improvement in cardiac response to VL, as measured by LVEF, LV end diastolic volume, left atrial volume (LAV), or right ventricular systolic pressure (RVSP). Tadalafil plus SC BNP resulted in improved cardiac response to VL, with increased LVEF (4.1 vs. 1.8%, ClinicalTrials.gov NCT01544998. We hypothesize that the combination therapy of tadalafil plus BNP will result in a greater improvement in cardiorenal adaptation to VL as compared to tadalafil alone.22.1The study is a double‐blinded, placebo‐controlled, cross‐over study to compare tadalafil alone versus tadalafil plus subcutaneous (SC) BNP on cardiorenal responses to saline VL in subjects with PDD. This study was approved by the Mayo Foundation Institutional Review Board and was performed at the Clinical Research Unit at Saint Mary's Hospital, Mayo Clinic Rochester Minnesota. Written informed consent was obtained from all participants.2.2Twenty patients with PDD (AHA Stage B HF) were enrolled Figure  and rand2.3Inclusion criteria include the following: ejection fraction greater than 50%; moderate or severe diastolic dysfunction as assessed by Doppler echocardiography; no current or previous diagnosis of HF; not on loop diuretics; and New York Heart Association Class 1 with minimal distance >450 meters on 6‐minute walk test in the absence of mechanical limitations  for 1 week prior to study initiation and were compliant by self‐reporting. Twenty‐four‐hour urine collection was obtained 1 day prior to the active renal clearance study day for assessment of sodium excretion, creatinine clearance, and microalbuminuria. If the 24‐h urine collection revealed that subject has been non‐compliant with diet, diet instructions were given again and the subject returned after at least 1 week with repeat 24‐h urine collection. Hematology and biochemistry labs, a 6‐min walking test, and a physical examination with vital signs were obtained. One day prior to the study date, subjects who met inclusion criteria were recruited and admitted to the Clinic Research Unit (CRU) at St. Mary's Hospital, Mayo Clinic CTSA, Rochester, Minnesota  were administered. Venous blood samples and renal clearances were obtained at 60 min. Measurements of the urine and blood were made: urinary flow (ml); urinary sodium excretion (UNaV) (mEq/min); urinary cGMP excretion (UcGMP) (pmol/min); and blood sodium mEQ/L and cGMP pmol/ml. Continuous monitoring by ECG and blood pressures were obtained in subjects. An echocardiogram was obtained for left atrial (LA) and LV volumes and systolic and diastolic functions.Subjects were then randomized to receive oral tadalafil 5 mg and SC placebo or oral tadalafil 5 mg and SC BNP 10 µg/kg . Based on prior studies and to minimize hypotension, a tadalafil dose of 5 mg was used , BNP, aldosterone, angiotensin II, and urine cGMP were obtained and determined by radioimmunoassay as previously described  were made using a paired t‐test. For all analyses, statistical significance was accepted as p < 0.05. Statistical analyses were completed with SAS 9.4 .Continuous variables are presented as mean ± SD and discrete variables as frequency (percentage). Comparisons between the two treatment groups  were made using the Student's 32). Comorbidities such as diabetes mellitus (25%), coronary artery disease (45%), and history of hypertension (75%) were common. The average left ventricular ejection fraction (LVEF) was 58.4%, left ventricular end diastolic volume (LVEDV) was 152.6 ml, right ventricular systolic pressure  was 31.8 mmHg, E/e’ was 14.3, and left atrial volume (LAV) was 80.8 ml. The average ANP and BNP values were low, 43.5 pg/ml and 79.9 pg/ml, respectively. Similarly, both mean plasma aldosterone and angiotensin II levels were within the normal range. Despite subjects having diastolic dysfunction and LA enlargement, both ANP and BNP levels are low with no activation of the angiotensin aldosterone system, consistent with asymptomatic diastolic dysfunction. The characteristics of the study population are thus similar to that previously reported  (Table p = 0.23), LVEDV , RVSP , or LAV .With tadalafil alone, there was a decrease in systolic and diastolic blood pressures and trend for an increase in heart rate with intravenous saline VL  (Table p < 0.01), a decrease in LVEDV , a decrease in LV end systolic volume (LVESV) , a decrease in RVSP , and a trend for decrease in LAV  in response to VL.Tadalafil plus SC BNP combination, there was also a decrease in systolic and diastolic blood pressures and increase in heart rate with VL  and RVSP   and heart rate   and sodium excretion ; however, there was no significant increase in GFR, renal plasma flow, or urinary cGMP excretion in response to VL  and urinary cGMP excretion , but no significant increase in urine flow, GFR, or renal plasma flow in response to VL   Figure .3.3p = 0.69) and a trend for increase in plasma cGMP  in response to VL  and an increase in cGMP  and BNP  in response to VL    Figure  and a gr) Figure .3.4No subjects receiving tadalafil and SC placebo had adverse events. With administration of tadalafil and SC BNP, two subjects (10%) experienced hypotension that subsequently resolved with saline infusion, and one subject (5%) had diarrhea that self‐resolved.4This study characterizes the acute cardiac and renal adaptations of PDEVI alone and in combination with SC BNP in response to VL in subjects with PDD. This is the first‐in‐human study to define the effects of such cGMP modulation in PDD. Our goal was to determine if tadalafil alone or in combination with SC BNP would result in improved cardiac and renal adaptations to VL in PDD subjects. Our findings showed that PDEVI alone did not improve cardiac adaptation to VL. However, the combination of PDEVI and exogenous SC BNP resulted in improved cardiac adaptation to VL. With regard to renal response, there was no difference in either group.p = 0.06) may suggest a decrease in cardiac filling pressures with PDEVI and BNP. In the PDEVI alone group, there was no change in ANP with VL .Preclinical diastolic dysfunction (PDD), which resides in ACC/AHA Stage B heart failure, and is defined as ejection fraction greater than 50%, with moderate/severe diastolic dysfunction and no diagnosis or signs and symptoms of heart failure, is a common entity that disproportionately affects older patients with cardiovascular comorbidities, and results in increased risk of progression to symptomatic HFpEF  is currently approved for the treatment of heart failure with reduced ejection fraction (HFrEF) (Yancy et al., These findings may be attributed to the fact that endogenous NP levels are lower in HFpEF compared to HFrEF (Maisel et al., We have previously reported that there is impaired cardiorenal response to VL in patients with PDD as compared to normal subjects (McKie et al., Given that this was a small proof of concept randomized double blinded placebo controlled cross over study with 20 subjects, and the focus is on investigating the effects of PDEV inhibition, either alone or added to BNP, we wished to maximize enrollment in groups receiving PDEV inhibition and thus did not have a BNP alone study arm. Prior studies have demonstrated that chronic administration of BNP alone in PDD results in significant activation of cGMP, natriuresis, and improvement in diastolic function (McKie et al., While there was no cardiac function improvement in response to acute VL with tadalafil alone, there was a significant improvement in cardiac function with combination tadalafil and BNP administration in subjects with PDD. However, there was no change in renal function which may be due to the decrease in blood pressure observed with tadalafil alone or in combination with BNP. This results in reduced renal perfusion pressure and distal sodium delivery, with worse sodium excretion (Haas et al., There are several potential mechanisms for the benefits in tadalafil, as a regulator of nitric oxide levels, when combined with exogenous BNP administration. PDEV hydrolyzes cGMP coupled to nitric oxide, but not cGMP associated with natriuretic peptides in cardiomyocytes (Lee et al., In summary, the results demonstrated that in human subjects with PDD, tadalafil and BNP combination compared to tadalafil alone resulted in cardiac function improvement, but no change in renal function with acute VL.4.1A limitation of this study is that only one dose of tadalafil and SC BNP was tested. Future studies are needed to further evaluate the effects of lower, non‐hypotensive doses of tadalafil and SC BNP. This is also a study assessing acute physiological derangements and hence, may not reflect the long‐term effects of tadalafil and SC BNP on cardiorenal function and neurohormonal effects in PDD. This study was limited by a shorter duration of investigation. Additionally, PDD is heterogenous, and future studies require selection of more specific patient groups from this diverse population.5In human subjects with PDD, PDEV inhibition alone with tadalafil resulted in no improvement in cardiac adaptation to VL. However, the combination of PDEV inhibition and exogenous SC BNP resulted in improved cardiac adaptation to VL.Dr. Chen and Dr. Burnett have patented and licensed designer natriuretic peptides.The authors have no conflict of interest to report.All authors have read and approved the manuscript, and have contributed to the design, collection of data, analyses, writing, and/or review of the manuscript.


Left ventricular geometry changed with an increase of end-diastolic volume (LVEDV) while end-systolic volume (LVESV) remained unchanged. Parameters concerning myocardial deformation , strain rate (SR)) decreased whereas the RR interval-corrected GLS (GLSc) remained unchanged. In a simple linear regression model, GLS correlated with 2D- and 3D-LVEF not only before (pre) ECV, but also after (post) ECV. We demonstrate that the increase of LVEF and GLS (ratios pre/post) correlates with the change of HR , whereas ratios of GLSc and SR do not significantly correlate with HR . Conclusion: In patients with tachyarrhythmia and reduced ejection fraction, ECV leads to immediate improvement in EF and GLS while HR-corrected LV contractility remains unchanged. This suggests that the immediate effects of rhythm restoration are mostly related to changes in left ventricular volume, but not to an acute improvement of heart-rate independent contractility.Aims: Tachyarrhythmia due to atrial fibrillation (AF) is often associated with reduced left ventricular (LV) function and has been proposed to cause arrhythmia-induced cardiomyopathy (AIC). However, the precise diagnostics of AIC and reversibility after rhythm restoration are poorly understood. Our aim was to investigate systolic LV function in tachycardic AF and to evaluate the direct effect of rhythm restoration. Methods: We prospectively studied 24 patients  with tachycardic AF and newly diagnosed reduced left ventricular ejection fraction (LVEF). Just before and immediately after electrical cardioversion (ECV), transthoracic echocardiography was performed. Geometric as well as functional data were assessed. Results: Patients presented with a heart rate (HR) of 117.4 ± 21.6/min and a 2D-/3D-LVEF of 32 ± 9/31 ± 8%. ECV to sinus rhythm normalized HR to 77 ± 11/min with an increase of 2D-/3D-LVEF to 37 ± 9/37 ± 10% ( Atrial fibrillation (AF) is the most frequent rhythm disorder and is often associated with heart failure (HF). These two conditions themselves share common risk factors . In addiBesides LV ejection fraction (LVEF), which remains the most commonly used conventional parameter, we employed speckle-tracking echocardiography (STE) and global longitudinal strain (GLS) analysis, as they are of high resolution. In addition to that, they have been validated in comparison to magnetic resonance and sonomicrometry ,11,12. TWe hypothesized that ECV leads to immediate improvements in LVEF by a combination of volume-dependent effects  and direct effects on left ventricular contractility. Therefore, geometric as well as functional parameters were comprehensively recorded using echocardiography directly before and after energy application in order to only detect the remodeling-independent effects of AF.This prospective observational single center study was reviewed by the independent ethics committee of the University of Regensburg  and all patients provided written informed consent. We included patients with tachycardic AF and newly diagnosed reduced LVEF < 50%, who were scheduled for ECV. If the intake of oral anticoagulation (OAC) in the past three weeks was uncertain, a transesophageal echocardiography was performed to rule out intracardiac thrombus. Patients with known left ventricular systolic dysfunction, more than moderate valvular heart disease and patients with acute coronary syndrome within in the last three months were excluded. All patients underwent echocardiography just before and after ECV. Patients with unstable clinical conditions, suboptimal image quality for analysis  or arrhythmias other than AF were excluded see .Patients scheduled for ECV were screened for HR > 100/min and LVSD (EF < 50%). Thirty five patients met the inclusion criteria. Four of them could not be cardioverted, echocardiographic image quality in seven patients was not satisfying . Twenty four patients were analyzed. Echocardiographic images were obtained using a Philips Epiq CVx ultrasound scanner  with a transthoracic matrix phased-array transducer . Standard 2D views in the parasternal and LV apical sound field were used, frame rate > 50 frames/s. Two-beat 3D full LV volumes were acquired for 3–5 cycles in AF and 2–3 cycles in sinus rhythm. Image acquisition occurred at a frame rate of 26.5 ± 3.4 30 frames/s. Careful attention was paid to adjusting the sector width and depth to include the entire LV myocardium, including the epicardial surface within the pyramidal scan volume. All original echocardiographic images were captured with the same machine, probe and investigator. Acquired 2D and 3D data were digitally transferred to a separate workstation for offline analysis of LV volumes, LVEF and strain using the IntellisSpace Cardiovascular Software  by a blinded user. Image loops  with the best image quality were used for analysis. Seven patients with suboptimal image quality, defined as more than three  until loss of consciousness. Blood pressure, heart rate and oxygen saturation were monitored. Synchronized shocks to a maximum of three shocks  were delivered until sinus rhythm was restored . All shop values < 0.05 were considered statistically significant. Data were analyzed using standard statistical software .Continuous variables are expressed as mean ± SD, categorical variables as percentages. Due to the nature of the study, pre vs. post ECV comparisons were made by paired Student’s t-testing. Relationship analysis between ΔHR and strain measurements as well as LVEF were assessed using a simple linear regression analysis and using Pearson’s correlation. Comparison of the ratios  was done by one-way ANOVA Tukey’s multiple comparison test. 2DS2-VASc-Score 3.1 ± 1.4). Patients who were not on OAC at presentation received initiation of OAC, and every patient whose intake of OAC within the last three weeks was uncertain received a transesophageal echocardiography to rule out intracardiac thrombus. LVEF was severely reduced (32.5 ± 9.1%) ranging from 13.8% to 46% and HRpre ranged from 63/min to 165/min. After ECV, systolic blood pressure increased from 134 ± 18 mmHg to 138 ± 16 mmHg without reaching the significance level (p = 0.07), diastolic blood pressure remained in a similar range of 81 ± 10 mmHg (pre) and 78 ± 11 mmHg .Thirty five patients were screened, but 11 patients had to be excluded from analysis  with a goodness of fit (R2) of 0.52 for 2D- and 0.54 for 3D-LVEF, thus there is a significant correlation between 2D/3D-LVEF and GLS . Post cardioversion slopes did also deviate significantly from zero (each p < 0.001) with an R2 of 0.57 and 0.51 for 2D- and 3D-LVEF (p = 0.99), a pooled slope of −1.91 can be specified. Moreover, slopes for 2D- (pre vs. post) and 3D-LVEF (pre vs. post) did not differ. These results indicate that the measurement of 2D- and 3D-LVEF is reliable in these patients and GLS correlates with LVEF, which has been shown in normofrequent patients before [LV function measured by Simpson’s method (2D) and by 3D volumetry correlates with global longitudinal strain before (pre) and after (post) cardioversion in a model for simple linear regression. More negative values of GLS represent more strain of the LV . Slopes  and GLS a. Among  3D-LVEF b. Since s before . ECV did−1  showing a positive correlation , the pooled slope equals 0.003 (p = 0.10 for GLSc and p = 0.59 for SR). R2 for the ratios of GLSc and SR are 0.03 and 0.01; Pearson’s was −0.17 for ratio-GLSc vs. ΔHR and 0.11 for ratio-SR vs. ΔHR. Moreover, ratio-GLSc- and ratio-SR-slopes differ significantly from ratio-GLS . There was no significant difference in ratio-GLSc- and ratio-SR-slopes . This means that GLS, but not GLSc and SR, correlate with ΔHR, and rhythm control via ECV did not impact these parameters of frequency corrected contractility.The means of 2D- as well as 3D-LVEF were higher after (post) than before (pre) ECV . To detels 0.003 a. For beHF and AF very often coexist. The recognition concerning arrhythmia-induced or arrhythmia-mediated cardiomyopathy is growing. However, the detection of LV function remains a difficult issue in patients with AF especially when associated with tachycardia. It is possible that the extent of contractile dysfunction is sometimes overestimated in the presence of atrial tachyarrhythmia. We sought to separate acute adaptive LV contractile changes due to tachyarrhythmia from distinct LV remodeling processes. The latter ones may recover if sinus rhythm persists. The discrimination of adaptive LV function vs. LV remodeling plays a relevant role for daily practice but also for further clinical trials. This study was designed as a single center study, thus the sample size is rather small, which is a limitation of the study.pre 117.4 ± 21.6/min). However, this study did not report LV geometry before and after ECV within the same modality. In the former study, an increase of LVEDV after ECV could be detected; at that time, there was no relining by 3D-data. In AF-ablation studies, end-diastolic LV size declined after intervention: end-systolic diameters decreased after 1–12 months, while LVEF increased after 1–6 months post procedure [Our data indicate that 2D-, 3D-LVEF, as well as LV strain, improved immediately after ECV. This improvement was dependent on the degree of HR-reduction. Remarkably, measures more specific for contractility  than LVEF did not display this frequency dependence. These findings are important for the interpretation of data in the literature and for the design of clinical trials in the field of AIC. Raymond et al. showed that LVEF  increased by 10% up to 61% two to 24 h after cardioversion. Heart rates were 77/min before and 59/min after ECV [rocedure . Likewisrocedure ,21. Baserocedure  and 79/mOur data show an acute increase of LVEDD and LVEDV, whereas end-systolic parameters remained unchanged. This might be due to prolonged diastolic filling after rhythm control and HR reduction by ECV. A study using cardiac scintigraphy with technetium-99m-labeled red blood cells demonstrated a strong correlation of RR interval with LVEF in patients with AF . An incrHere we show that the restoration of normofrequent sinus rhythm immediately improves LVEF and global longitudinal strain. Both parameters were dependent on the extent of HR reduction, but not for the HR independent SR or the heart rate corrected GLSc parameter.In summary, tachycardic AF is a condition in which systolic left ventricular function is reduced, which is—dependent on the study design—more or less visible. The increased end-diastolic volume can be explained by a prolonged diastolic filling period as a direct consequence of rhythm restoration. Moreover, an increased preload following rhythm restoration may potentially improve contractility via the Frank–Starling mechanism, leading to the improvement of parameters measuring LV deformation and ejection.In conclusion, we recommend deploying parameters of systolic LV function other than LVEF  in daily clinical practice but also in clinical trials if neither rhythm nor frequency control can be achieved.


Cardiac MRI in rhesus macaques, a species of major relevance for preclinical studies on biological therapies, requires artificial ventilation to realize breath holding. To overcome this limitation of standard cine MRI, the feasibility of Real-Time (RT) cardiac MRI has been tested in a cohort of ten adult rhesus macaques using a clinical MR-system. In spite of lower tissue contrast and sharpness of RT-MRI, cardiac functions were similarly well assessed by RT-MRI compared to cine MRI (similar intra-subject repeatability). However, systematic underestimation of the end-diastolic volume (31 ± 9%), end-systolic volume (20 ± 11%), stroke volume (40 ± 12%) and ejection fraction (13 ± 9%) hamper the comparability of RT-MRI results with those of other cardiac MRI methods. Yet, the underestimations were very consistent (< 5% variability) for repetitive measurements, making RT-MRI an appropriate alternative to cine MRI for longitudinal studies. In addition, RT-MRI enabled the analysis of cardio-respiratory coupling. All functional parameters showed lower values during expiration compared to inspiration, most likely due to the pressure-controlled artificial ventilation. In conclusion, despite systematic underestimation of the functional parameters, RT-MRI allowed the assessment of left ventricular function in macaques with significantly less experimental effort, measurement time, risk and burden for the animals compared to cine MRI. Translation of promising novel treatment approaches into clinical application often requires animal models very closely resembling humans. In this context, non-human primates (NHPs) have become of great interest for cardiac research2. For instance, studies on treatment of post-infarction heart failure by stem cell-based approaches have utilized the close immunological and genetic similarities between humans and NHPs7.Cardiovascular diseases, in particular coronary artery disease and heart failure, are the leading causes of death in Europe10. The required conscious breath-holding during data acquisition, however, can be impossible in many situations. Thus, artificial ventilation with anesthesia has been used for instance for cardiac MRI in neonates, infants and small children12.Monitoring treatment effects and disease progression requires a robust and reliable assessment of left ventricular function. The current gold standard in human cardiac MRI is cine MRI, an ECG-gated gradient echo sequence16. The currently proposed methods of RT-MRI differ regarding their steady state of magnetization , acquisition scheme , and image reconstruction algorithms. Commonly used are parallel imaging techniques such as GRAPPA13, SENSE14, and regularized nonlinear inversion (NLINV)15.In contrast to the cine MRI, Real-Time (RT) MRI can be performed without breath holding. By reducing the number of acquired data points, an image can be obtained within about 20 ms12, the current standard of cardiac MRI in NHPs is to perform “forced” breath holding during artificial ventilation20. Therefore, RT-MRI may be an alternative and, together with the significant reduction of measurement time, this method may reduce additional risks and burdens for the animal. Rather fast spontaneous breathing and high heart rates  are, however, particular challenges of RT-MRI in these small animals and the potential bias in quantitative analyses by substantial under sampling needs to be evaluated.The feasibility of RT-MRI for cardiac MRI in NHPs is, however, largely unknown. As in infants and young children25. Since cine MRI requires breath holding, comparative studies in children were limited to an age not younger than seven years26, but reported likewise similar results.In adult humans, the few studies comparing cine MRI and RT-MRI have mostly reported similar results of left ventricular function obtained with both methodsThe aim of this study was to evaluate the capability of RT-MRI for assessing the cardiac function in NHPs and to compare the information obtained by this method with that of cine MRI. This was achieved by measuring each animal twice within few weeks and parameters of left ventricular function were independently extracted for both cine MRI and RT-MRI by two experienced observers. Intra-subject repeatability, inter-observer agreement and systematic differences between the two acquisition methods were evaluated. Moreover, since RT-MRI allowed for the analysis of every single heartbeat, left ventricular function parameters were monitored over at least six heartbeats while the animal was continuously breathing.Parameters of the left ventricular function and mass obtained by cine MRI (short-axis ECG-gated segmented Cartesian FLASH acquired during expiratory breath holds) at the first (test) and the second study time point (retest) are summarized in Table Left ventricular wall mass , end-diastolic volume , end-systolic volume  and stroke volume  varied widely between the animals, but showed an excellent intra-subject repeatability  which corresponded to about 5.5% of the total LVM (Range: 2.1–8.1%). Including the papillary muscle from the myocardium, led, as expected, to a decrease in the calculated EDV, ESV and SV of about 6% (± 1.4), 10% (± 2.9) and 3% (± 2.3), respectively. In contrast, the EF increased by about 3% (± 1.9).p < 0.01). The lower tissue contrast hampered in particular the definition of the most basal and the most apical slice of the heart. Moreover, the lower image sharpness led to a more challenging segmentation of the papillary muscles and the trabeculae . As shown exemplarily for a mid-ventricular slice , the signal profile of RT-MRI appeared smoother and Gaussian shaped compared to the signal profile of cine MRI.RT-MRI allowed for mostly artifact-free data acquisition while the animals were continuously breathing. Figure The time needed for the myocardial segmentation of RT-MRI data sets was on average five times longer compared to cine MRI, mainly due to the larger number of separately segmented heart cycles. In addition, the lower tissue contrast and image sharpness as well as the lack of appropriate segmentation software prolonged the segmentation time.Table To compare cine MRI and RT-MRI, only real-time data acquired during expiration was chosen and papillary muscles were excluded from the myocardium. Absolute and percentage differences between parameters acquired by cine MRI and by RT-MRI are summarized in Table Using RT-MRI we were able to analyze the cardiac function for every heartbeat and respiratory phase separately. Figure By analyzing the entire left ventricle, EDV and ESV exhibited about 10–20% higher values in inspiration compared to expiration while the SV increased by about 40%. In contrast, EF showed only a small increase of less than 10% while LVWM remained almost constant. Beside the difference in absolute values, EDV, ESV, SV, and EF showed a strong relationship between the two respiration phases as illustrated in Fig. 28. The almost similar LVWM values suggest that physiological differences may dominate the difference observed between the two study time points. Moreover, the only two non-human primate studies having so far addressed the repeatability of cardiac function measured by cine MRI presented very similar results17.This study compares cine MRI and RT-MRI for assessing left cardiac ventricular function in healthy rhesus macaques, a non-human primate species of significant importance in preclinical research on biological therapies. In addition to demonstrating the feasibility of RT-MRI in animals with higher heart rate (~ 105 bpm) and much smaller heart size , we showed a similar data quality of RT-MRI and cine MRI. Particular advantages of RT-MRI were the possibility of beat-by-beat image analysis and markedly shorter acquisition time. Another clear advantage of RT-MRI is that it does not require breath holding alleviating the need for artificial ventilation. This significantly reduces the preparation and measurement time, the duration of anesthesia and therefore the potential risk and burden of the animal. With the chosen protocols, we could reduce the acquisition time, in comparison to cine MRI, by a factor of more than two. Taking the recovery time between two breath holds into account, which may be particularly relevant in cardiac disease models, the acceleration factor increases even more. In fact, it was possible to limit the total scan time including localizer and functional data set to about five minutes per animal. However, these benefits were achieved at the expense of a slightly reduced sharpness of the image and a lower tissue contrast. This fact made it more challenging to identify the most basal and the most apical slice of the heart. Moreover, the achieved image quality hindered an automatic segmentation of myocardial contours with the current available software tools. The lower image quality of RT-MRI, however, did not lead to a reduced repeatability of the method. In fact, cine MRI and RT-MRI revealed very similar intra-subject repeatability and an excellent inter-observer agreement. In particular, EDV was best repeatable, but even the lower repeatability of ESV, SV and EF was very much in line with those reported in human studies24. Although exhibiting the strongest correlation for the two methods, volumetric parameters  were systematically underestimated by approximately 20 to 40% when using RT-MRI. This is in contrast to previous human studies29 and most likely caused by the required regularization during image reconstruction, the intrinsic characteristics of the radial point-spread-function which may be more prominent in the smaller and faster beating heart of a rhesus macaque and inaccuracies in the delineation of endocardial contours of the myocardium. This underestimation was, however, rather consistent. The two methods revealed almost identical differences between the two study time points  the required high spatiotemporal resolution of RT-MRI restricted the use of bSSFP due to the higher specific absorption rates (SAR) of the sequence.A limitation of this study is the exclusive use of spoiled segmented FLASH instead of balanced steady-state free precession (bSSFP), a MR sequence commonly used in cardiac MRI of adult humans. However, bSSFP sequences are, especially at higher magnetic fields and spatial resolution used here, particularly sensitive to differences in magnetic susceptibility and to magnetic field inhomogeneities which often lead to disturbing banding artifacts36 and respiratory maneuvers35 can significantly influence the cardiac flow, volumes and function. However, the exact underlying mechanism is not clear yet. RT-MRI offers the unique opportunity to investigate these effects during free and artificial breathing. In our experiments  higher volumetric values were observed during inspiration, while in case of free breathing EDV, ESV and SV revealed the highest values during expiration . This is most likely due to the fact that in normal breathing the inspiration goes along with a reduction of chest pressure, whereas in case of artificial ventilation the breathing air is actively pressed into the airways leading to a pressure increase on the vascular system.One of the advantages of RT-MRI is the possibility to analyze individual heartbeats and therefore to study the interaction between respiration and cardiac function. It has been shown that normal breathingDue to the continuously changing of EDV and ESV during breathing, we strongly recommend the inclusion of more than one heart cycle in the quantitative analysis of RT-MRI. Based on this study we suggest acquiring cardiac Real-Time data continuously over at least two breathing periods. The respective heart cycles can then be selected based on their position in the breathing cycle  which will allow a more robust comparison between different animals.Another, and for quantitative group analyses probably even more suitable approach, could be the use of self-gating algorithms utilizing, for instance, signal and phase information of the central data point of each k-space line. Unfortunately, the best sampling schema for RT-MRI is not also the best for self-gating. Thus, this would exceed the scope of the manuscript and will be the subject of another study.In conclusion, Real-Time cardiac MRI allows for the assessment of left ventricular cardiac function of NHPs. Compared with cine MRI, it significantly reduces the potential burden of the animal. The absolute values of cardiac parameters differed between cine MRI and RT-MRI, which hampers a direct comparison across sites and studies. However, comparisons within each of the two methods will most likely reveal very similar results.In addition, further optimization of tissue contrast and sharpness, as well as development of automatic segmentation tools are required to speed up the currently rather time-consuming data analysis. The lack of appropriate software to automatically or semi-automatically quantify the cardiac function using RT-MRI is currently probably the biggest barrier in bringing it to a broader application.macaca mulatta, mean age: 7.5 ± 2.2 year, including 5 males) was included in this study. All monkeys were purpose-bred, raised and housed according to the standards for macaques of the German Primate Center . All animal experiments were in full accordance with the ARRIVE guidelines, the German animal welfare law (TierSchG), the regulations for research animals (TierSchVerV) and approved by local authorities . Initially, animals were sedated with an intramuscular injection of 7 mg/kg ketamine  and 0.04 mg/kg medetomidine . Anesthesia was maintained via continuous infusion of 10–40 mg/kg/h Propofol-Lipuro 2% . The intubated monkeys were pressure-controlled ventilated  with at least 60% oxygen in the exhalation air in order to keep the O2 saturation between 98–100% . The CO2 concentration in the exhalation air  was maintained below 30% to avoid spontaneous breathing.A cohort of 10 adult, healthy rhesus macaques . While the heart rate under anesthesia  varied widely between the two study time points (mean difference of 3 ± 11 bpm), the body weights  remained stable (mean difference of 0 ± 0.2 kg).2 were continuously monitored.Measurements were performed on a 3T MRI-system  using a 16-channel multipurpose coil  for signal detection. The animals were positioned in supine position. Physiological parameters such as heart rate, breathing rate and end-tidal COCine MRI: For slice positioning, low-resolution (1.55 × 1.55 × 8 mm3) ECG-gated T1-weighted images were conducted during free breathing in 2-chamber, 4-chamber, and short-axis orientation. Subsequently, short-axis cine multi-phase segmented FLASH data sets were acquired using the following parameters: repetition time (TR) = 6.48 ms, echo time (TE) = 2.84 ms, flip angle = 12°, bandwidth = 215 Hz/Px, field of view (FOV) = 132 × 149 mm2, acquisition matrix = 128 × 176 (reconstructed to 156 × 176), 5 segments, readout oversampling = 2, spatial resolution = 0.85 × 0.85 mm2, slice thickness = 3 mm, slice gap = 0.3—0.6 mm, and 12–14 slices covering the heart from the apex to the base. No parallel imaging was used. Data was continuously recorded over 26 heartbeats and then retrospectively reconstructed to 25–40 phases per cardiac cycle. The number of phases per cardiac cycle (MR data was obtained during breath holds (15–20 s at average) in the expiratory phase. Including resting time of at least two minutes between two breath-holds the total measurement time of a complete cine dataset was around 30 min. Noteworthy to mention, the pure time of data acquisition for a single slice was on average 15 s.RT-MRI: Similar to the localizing measurements for cine MRI, data sets were acquired in 2-chamber, 4-chamber, and short-axis orientation. This time, however, a low-resolution version of the final RT-MRI protocol was used  without any cardiac triggering and breath holding. Real-Time data sets for quantification of left ventricular function were acquired using spoiled radial FLASH with an interleaved ordering scheme: TR = 2.89 ms, TE = 1.92 ms, flip angle = 8°, bandwidth = 1305 Hz/Px, FOV = 128 × 128 mm2, base resolution = 128, 13 spokes per frame, nyquist undersampling factor = 15.5, spatial resolution = 0.9 × 0.9 mm2, slice thickness 3 mm, slice gap = 0.3–0.6 mm, acquisition time per frame = 37 ms, 210 frames per slice and 12–14 consecutively acquired short-axis slices covering the heart from the apex to the base. The scan time for a single slice was 8.2 s. The number of heart cycles covered by the 210 frames was depended on the individual heart rate and were in the range of about 10 to 15 consecutive cycles per slice. Online reconstruction of Real-Time data sets was carried out using the iterative non-linear approach described by Uecker et al.38 and was performed in real time on a graphical processing unit by bypassing the MRI host.39.The left ventricle function of short-axis images was independently evaluated in a blind fashion by two observes with at least two years of experiences in cardiovascular MRI of primates using the freely available software package Segment 40. The most apical slice was defined as the last slice, which includes blood volume. The resulting number of analyzed slices per animals was 10–12.For cine MRI end-diastolic and the end-systolic phase were selected as the phases with the highest and lowest blood volume of a mid-ventricular slice, respectively. Endocardial and epicardial contours were manually drawn from base to apex of the heart in both cardiac phases Fig. . AccordiIn case of RT-MRI data sets were firstly sorted regarding their cardiorespiratory phase using an in-house Matlab script . Therefore, the signal-time curve of manually drawn regions of interest in the left ventricle and at the lung-liver interface were extracted for each slice. The diastolic and systolic phases were selected automatically using a peak detection algorithm. Inspiration and expiration were separated by signal thresholding. Heart cycles in the transition phase of inspiration and expiration were neglected. This procedure is exemplary shown for a mid-ventricular slice in Fig. 41. The stroke volume (SV), the ejection fraction (EF) and the cardiac output (CO) were defined as the difference between EDV and ESV, the relation of SV and EDV and the product of SV and heart rate (HR), respectively. Additionally, for cine data sets left ventricular wall mass (LVWM) and the papillary mass (PM) were calculated as the average of the corresponding mass in the diastolic and systolic phase. The left ventricular mass (LVM) was determined as the sum of LVWM and PM.Based on the segmentation of endocardial and epicardial contours on the cine and Real-Time data sets the end-diastolic volume (EDV) and the end-systolic volume (ESV) were calculated using Simpson’s method of disks42. All statistics were carried out in Matlab .Intra-subject repeatability and inter-observer agreement were calculated using the intraclass correlation coefficient (ICC). Additionally, regression analysis and Bland–Altman plots were used to visualize possible systematic trends. Limits of agreement for repetitive experiments and multi observer were calculated as described by Bland and AltmanSupplementary Information.


All HIT-positive patients were switched from heparin to argatroban. HIT-associated thrombotic events were observed in two cases resulting in Impella dysfunction due to pump thrombosis (n = 1) and left ventricular (LV) thrombus formation (n = 1). Under large Impella support, the prevalence of HIT was relatively high. Further, thrombocytopenia does not deliver a high specificity in the setting of Impella 5+ support. Considering HIT manifestation, a routine HIT test may be considered to avoid critical thrombotic adverse events.Despite the critical feature of heparin-induced thrombocytopenia (HIT) for patients on mechanical circulatory support, reports on its incidence and outcome are still scarce. Thus, we report on clinical features of HIT in patients under Impella 5.0 or 5.5  (Impella 5+) support for acute cardiogenic shock (CS) by focusing on observed thrombotic events. Between November 2018 and December 2020, a total of 56 consecutive patients were enrolled in a single-center retrospective study. A total of 21 patients (37.5%) were tested for HIT, and 6 (10.7%) proved positive for HIT at 10.5 ± 2.89 days after the first heparin administration during current admission. Interestingly, thrombocyte counts dropped under Impella support in all groups (all cases, no HIT test, and HIT negative group:  The use of an extracorporeal circulatory system, e.g., veno-arterial extracorporeal membrane oxygenation therapy (va-ECMO), is widely adopted in many countries as a solid and reliable tool for mechanical circulatory support (MCS) in patients with severe heart failure (HF) or cardiogenic shock (CS). In this patient cohort, anticoagulation is indispensable for preventing thrombotic adverse events. However, bleeding disorders, as well as thromboembolic events, are considered as relevant complications of patients under MCS by va-ECMO. Recently, microaxial pumps, i.e., Impella , have emerged as an alternative tool for temporary MCS. Despite the substantially different mode of action of Impella omitting extracorporeal blood circulation and the need for an oxygenator, MCS by Impella yet requires anticoagulation. As recommended by the manufacturer, a purge fluid containing anticoagulant is constantly administered to wash the Impella pump and prevent patient blood from entering the motor. In such a setting, anticoagulation management is crucial to avoid pump thrombosis, and unfractionated heparin (UFH) is utilized as the first choice for anticoagulation.However, heparin-induced thrombocytopenia (HIT) is a well-described and severe complication in patients with continuous intravenous heparin administration. Antibodies to the platelet factor 4 and heparin complex play a central role in HIT development, resulting in a fatal hypercoagulation state without causing bleeding . In fronEven though HIT is a crucial issue in utilizing the Impella system, only a few cases are reported in the literature ,4,5,6,7.Thus, in this report, we documented our clinical experiences of Impella 5+ and analyzed HIT-positive patients to review the strategy for early HIT diagnosis and early modification of Impella management.n = 67 for Impella 5.0; n = 3 for Impella 5.5) implantations in 60 episodes of care  in our institution. Four patients who underwent off-pump coronary artery bypass grafting under Impella 5.0 support in the setting of ischemic cardiomyopathy without CS were excluded. Thus, a total of 56 acute CS cases were enrolled in this study  with UFH (50 U/mL) was used as standard. Moreover, systemic UFH was also administrated to achieve target activated partial thromboplastin time (aPTT) above 45 s and below 72 s. aPTT was monitored every 8 h until aPTT values became stable, from there on in 24-h intervals. However, in patients with clinical bleeding tendency, i.e., coagulopathy, or in postoperative cases, anticoagulation was adopted to the clinical situation in a case-by-case manner, in the extreme scenario of intractable postoperative bleeding also considering temporary pause of anticoagulation therapy. In such a case, we restarted the anticoagulation once the bleeding situation had improved. HIT test was performed in suspected cases with a marked drop in platelet counts. In general, some scoring systems, e.g., the 4Ts scoring system  or the HIn the setting of HIT, argatroban (2 μg/kg/min) was initially administrated intravenously and modified according to aPTT for systemic anticoagulation. As far as the addition of anticoagulation agents to purge solution is concerned, no proper guidelines existed at the time of the reported cases. Likewise, a case-by-case decision was made in an interdisciplinary team consisting of a cardiac surgeon, an intensive care specialist, an haemostaseology specialist, and in compliance with the recommendations of the clinical specialists of the manufacturer.® (SPSS) 25.0 . Using this program, descriptive and comparative statistics were performed. Unless otherwise indicated, continuous variables were compared using the Mann–Whitney U test for two groups’ analysis. For three group analysis, the Kruskal–Wallis test was conducted. A Chi-Quadrat-Test was conducted for nominal scaled variables. However, Fisher’s exact test was adapted instead of the Chi-Quadrat-Test for a minimum expected value of less than 5. p-values less than 0.05 were considered statistically significant. The statistical analysis was conducted in a patient-based manner in this study. All analyses were performed with the Statistical Package for Social Sciencesn = 35, 62.5%), HIT-positive group , and HIT negative group  (p < 0.05). Further, the C-reactive protein value was also statistically higher in the no HIT test group than the HIT negative group (<0.05). As far as the in-hospital mortality is concerned, 33 patients of the total cohort (57.9%) did not survive, whereas, in HIT-positive group, 2 patients (33.3%) died, which suggested no statistically significant difference.Among 56 patients, we performed HIT tests in a total of 21 patients (37.5%), 6 of whom  were confirmed to be HIT positive. For the descriptive analysis, we assigned all patients in three groups; no HIT test group  . Most ofp = 0.15). In each case, the first heparin administration was in the context of either veno-arterial extracorporeal membrane oxygenation (va-ECMO) implantation, open-heart surgery, or Impella implantation, with no statistical difference between the study sub-cohorts . HIT-associated thrombotic events under Impella 5+ support were observed in two cases, presenting as Impella dysfunction due to pump thrombosis (n = 1) and as left ventricular (LV) thrombus formation (n = 1)  .p < 0.001, HIT-positive group: p = 0.001).The survey of platelet counts using Impella 5+ depending on HIT diagnostic is shown in p = 0.02)  .2 as determined by right-heart catheterization and LV ejection fraction (EF) of 24% as determined by echocardiography. Due to fulminant myocarditis with high catecholamine levels, va-ECMO became necessary, immediately followed by axillary Impella 5.0 implantation on admission day. A HIT test was repeatedly performed because of a decrease in circulating thrombocyte counts. However, the HIT test initially proved to be negative, although OD became tendentially higher. Thus, according to the standard protocol, UFH was further applied for systemic anticoagulation and supplemented to the purge solution. On the eighth postoperative day (POD), repeated HIT tests resulted positive, so anticoagulation therapy was switched from UFH to argatroban. On the ninth POD, purge flow rate decreased, and Impella flow also dropped to levels below 2 L/min despite choosing P7 Impella setting. Finally, Impella flow decreased to less than 1 L/min, and purge pressure increased to over 1000 mmHg. In an analysis of all available parameters and compliance with recommendations given by clinical specialists of the manufacturer, system dysfunction due to pump thrombosis was diagnosed. Therefore, we explanted the dysfunctional Impella 5.0 on the 10th POD. Fortunately, LV function had meanwhile markedly improved so that the patient did not require a continuation of MCS. Transthoracic echocardiography (TTE) revealed 47% LVEF after va-ECMO explantation on the 18th POD. After a steady recovery course, the patient was finally on the 27th POD transferred to a peripheral hospital in a satisfying condition for further re-convalescence.The clinical course of a patient with pump thrombosis associated with HIT is shown in The clinical course of another patient with LV thrombus associated with HIT is shown in The main results of this study are: (1) the incidence of HIT under Impella 5+ lies at 10.7% and is diagnosed at 10.5 ± 2.89 days after initial heparin exposure; (2) a marked drop in thrombocyte counts was present in all Impella patients. However, with the most substantial relative drop observed in HIT-positive patients (average 76.1%), and (3) in the small sub-cohort of Impella patients developing HIT a high rate of 33.3% was observed for thrombotic clinical events.Generally, the prevalence of HIT in critically ill patients is reported to range under 1%, whereas the incidence of HIT post cardiovascular surgery is reported at 1–2% . This stWe performed HIT tests in one-third of our Impella 5+ patients (21/56), who had relevant thrombocytopenia suspicious for HIT. An early HIT diagnosis is indispensable for favorable therapy outcomes because of its crucial life-threatening condition, i.e., hypercoagulation. There may not be a constant high level of attention present in routine clinical practice towards the potential HIT development. Occasionally, due to the absence of suspicion about HIT, HIT seems to remain “hidden.” In other words, HIT positive cases might probably be just “the tip of the iceberg.” This puts a timely and efficient diagnosis finding process into the center of therapy efforts, warranting a constantly high awareness for HIT, especially in patients with temporary MCS. For the proper differential HIT diagnosis, the 4Ts scoring system  was established . In a prArising from this dilemma, we conducted this retrospective observational study to examine HIT incidence, to determine how HIT may impact on clinical outcomes, and to elucidate the clue for HIT strategy in CS patients with Impella 5+. Then we observed that the decrease in thrombocytes was manifested in the HIT-positive group and in all other groups. Thus, we suggest that the HIT test be performed routinely, and anticoagulation management be modified if the thrombocytes are reduced, until HIT diagnosis is validated.Regarding risk factors of HIT, no risk factors were determined in this study, although female gender and major surgery were reported as risk factors in previous reports ,14. ThisConcerning anticoagulation in HIT, anti-thrombin inhibitors, e.g., argatroban ,6,7 or bOn the other hand, another report documents the successful approach with no anticoagulant in purge solution under therapeutic systemic anticoagulation with bivalirudin . In the There are several limitations to this study. First, this manuscript deals with a retrospective observational study on a limited cohort size of non-randomized patients at a single center. Potential systematic measurement errors and also unidentified bias can affect the outcomes. Secondly, our analysis does not cover data on long-term outcomes. Third, regarding HIT diagnostics, we additionally performed a HIPA test only in the case of positive first ELISA and negative high-dose heparin confirmatory ELISA. For platelet function, the HIPA test should be performed to confirm if HIT-antibody occurred thrombocytes aggregation. Further studies are warranted for confirmation of our therapy strategy in HIT under Impella.Under large Impella support, the prevalence of HIT was relatively high (10.7%), with the onset time at 10.5 ± 2.89 days after the first heparin administration during current admission. 33.3% of all HIT patients suffered from critical hypercoagulation events. In the light of these findings, routine HIT tests and a liberal change of anticoagulation regimen upon the drop of thrombocyte counts appear favorable for patients under Impella support. Anti-thrombin inhibitors, argatroban, or bivalirudin will be an alternative anticoagulant instead of UFH. However, ideal dosage and administration pathways remain the subject of future studies. Further, the findings of this study are hypothesis-generating, which need to be confirmed in future studies involving larger patient cohorts, ideally in the setting of a randomized controlled trial.