Misoprostol is listed in the WHO essential medicines list and can be used for induction of labour, for prevention and treatment of post-partum haemorrhage, and for abortions. The compound is unstable, and substandard misoprostol preparations have been detected in low- and middle-income countries. We now investigated the stability of misoprostol tablets according to the international guidelines for stability testing of pharmaceutical products. Three brands (four batches) of misoprostol tablets were collected in Malawi and Rwanda: the originator product, a WHO-prequalified product, and a generic product without WHO prequalification. A further batch of the originator product was collected in Germany. To investigate the effect of damage to the primary packaging, additional blister strips of one sample were intentionally damaged with a needle and investigated in parallel. Samples were placed in stability chambers for six months at 40°C/75% relative humidity (RH) and at 25°C/60% RH. After 0, 1, 2, 3 and 6 months, misoprostol content was determined according to the International Pharmacopeia. At 40°C/75% RH, all samples showed a decline of misoprostol content, but four of the batches still remained within the pharmacopeial specifications, while one of the two batches of the generic product without WHO-prequalification showed a final content of 86.2% which is out of specifications. Damage to the primary packaging greatly decreased stability, resulting in a final content of only 48.2% of the declared misoprostol amount. At 25°C/60% RH all samples remained in specifications for six months, even the sample with the damaged blister. Dissolution of misoprostol remained in specifications of the pharmacopoeia for six months for all batches, except for the sample with damaged blisters stored at 40°C/75% RH. This study confirms that the stability of misoprostol tablets must be ensured by intact, good-quality primary packaging. Careful supplier qualification is required in the procurement process. Misoprostol , a prostMisoprostol is available in form of tablets which can be administered orally and which do not require refrigerated storage. Therefore, misoprostol tablets appear to offer an attractive alternative to oxytocin injections in the prophylaxis and treatment of PPH in places where no trained staff is available to administer medicines parenterally, or where appropriate storage conditions for oxytocin cannot be ensured , 18, 19.The instability of misoprostol has been known since decades . The mosPure misoprostol shows 50% degradation within two weeks at 55°C . FortunaThe quality of misoprostol preparations circulating in LMICs has received much less attention in the scientific literature than the quality of oxytocin. Besides the above-mentioned study by Hall , only twDuring the registration of medicines, National Medicines Regulatory Authorities (NMRAs) examine the stability data provided by the manufacturers, usually they do not carry out independent experiments to confirm these data. The NMRAs of the EU member states, USA, Japan, Switzerland, Canada, Australia, Iceland, Liechtenstein and Norway are currently considered as “Stringent Regulatory Authorities” (SRAs) , and medRules for stability testing of pharmaceutical products have been devised by the International Council of Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use (ICH), most importantly guidelines Q1A-Q1F . ObviousIn the present study, accelerated stability testing of misoprostol tablets collected in Malawi and Rwanda was carried out according to the ICH guidelines for stability testing of pharmaceutical products .This study was designed observing the recommendations contained in the WHO guidelines on the conduct of surveys of the quality of medicines , the MEDThe brands of misoprostol tablets available at government medical stores and pharmaceutical wholesalers in Malawi and Rwanda during the time of sample collection (February-March 2018) were purchased by local researchers (F.K. and T.B.). If different batches of a certain brand were available, samples from each batch were purchased. From each brand and batch, 250 tablets were purchased. Samples were not collected from individual health facilities or private pharmacies, to minimize the influence of possibly incorrect storage conditions on the results of this study. Also, these facilities usually do not stock sufficient amounts of misoprostol tablets for stability testing. After purchase, samples were stored according to the manufacturers’ specifications. Temperature data loggers were kept with the samples until these were placed into the stability chambers for testing. The investigators hand-carried the samples from Malawi or Rwanda to Germany via airplane; this transport required less than 24 hours. Samples were pre-tested for their misoprostol content according to International Pharmacopeia. Those samples which were within specifications at the time of pretesting were included into the stability study.The samples were stored in their original primary packaging in stability chambers of Alpha-Pharma-Service GmbH, Heilbronn, Germany, for six months (April-October 2018). Conditions were chosen in accordance to ICH and WHO guidelines for stability testing of pharmaceutical products , 31. AccSamples were visually inspected and subsequently tested for identity, assay and dissolution following the procedures described in the monograph for misoprostol tablets of the International Pharmacopeia 2017. Prior to the experiments, the methods were validated according to the International Pharmacopeia. Misoprostol was identified and quantified by high performance liquid chromatography (HPLC), using an Agilent Infinity 1260 II with binary pump and variable wavelength detector , with acetonitrile/water (45:55 V/V) as mobile phase, flow rate 1.5 ml/min, column ReproSil-XR 120 C18, 5 μm, 150 mm x 4.6 mm , and UV detection at 200 nm. The injection volume was 100 μl for identity and assay, or 250 μl for dissolution. For identity and assay, five tablets were dissolved in 50 ml mobile phase; two independent experiments were carried out, and of the resulting solutions two aliquots each were analysed by HPLC, yielding four measurements for each sample. The HPLC autosampler was set to 4 °C to avoid misoprostol degradation. Samples showing additional peaks were also tested for related substances according to Kahsay et al. , using fDissolution was investigated with a dissolution tester PT-WS 610 . Six tablets per sample were tested separately as described in the monograph for misoprostol tablets in the International Pharmacopeia, with 500 ml of water R as dissolution medium, a temperature of 37°C, and a rotating paddle with 50 revolutions per minute. Samples were withdrawn after 30 min through an in-line filter. Misoprostol reference standard Ph. Eur. (batch N° 3.0) was obtained from the European Directorate for the Quality of Medicines (EDQM), Strasbourg, France.At each time point during the stability testing, 5-point calibration curves were prepared to assure linearity. Intermediate precision  was calcStatistical evaluation was done using JMP 14.2 . Significance levels of differences were calculated using uni- and multivariate analysis of variance (ANOVA) and student´s t-test. Differences were considered significant when p< 0.05.®) was purchased through the pharmacy of the University Hospital Tuebingen. This is the brand most commonly used in Germany. A different batch of this originator brand had also been purchased in Rwanda.In this study, misoprostol samples were investigated which were offered by government medical stores and private wholesalers in Malawi and Rwanda, i.e. preparations which reflect the actual quality and stability of medicines distributed in these two African countries. In February and March 2018, the local investigators (F.K. and T.B.) enquired about the available brands and batches of misoprostol tablets. Samples were then purchased from private wholesalers using a mystery shopper approach (i.e. ordering with the help of a licensed pharmacy shop), and from government medical stores in an overt approach. If different batches of a certain brand were available, samples from each batch were purchased. In Malawi, certain brands of misoprostol tablets had recently been withdrawn from the market , and onlTherefore, as shown in Surprisingly, packaging and leaflet of the originator brand collected in Rwanda did not state any storage requirements, while packaging and leaflet of the same brand collected in Germany stated that storage below 30°C was required. The same storage temperature was stated by the manufacturers of the two other brands. The stated requirements for protection from light and humidity were somewhat different between the brands . All samThe three manufacturers were contacted and were requested to confirm the authenticity of the samples. The two manufacturers from India confirmed that label information and appearance of the samples conformed to their products. The latest response from Pfizer (the marketing authorization holder)/Piramal was that they were still investigating this matter.The declared shelf life of the investigated brands was either two or three years. While four of the five investigated batches remained within their shelf life during the entire duration of the study, the WHO-prequalified product by Acme expired during the stability testing. Nevertheless, it was found to remain within specifications during the entire testing period, even after storage for six months at 40 °C and 75% RH (see below).® which was neither SRA-approved nor WHO-prequalified, no excipients were declared at all.According to the current ICH/WHO guidelines for stability testing of finished pharmaceutical products , 31, accepi-misoprostol . The International Pharmacopoeia requires that at least 80% of the declared amount of misoprostol dissolves under the defined conditions see . As showThe sample with the punctured blisters clearly failed dissolution specifications after six months at 40°C and 75% RH, showing 42.4% dissolution of the declared API amount. Assay testing had shown a final API content of 48.2% of the declared amount. Therefore, storage in damaged blisters at these conditions strongly reduced the API content, and to some extent also reduced the dissolved percentage of the remaining API .After storage at 25°C and 60% RH, all tested samples, including the one with the punctured blisters, still complied with the specifications of the pharmacopeia for dissolution of misoprostol .The results of the present study confirm that the limited stability of misoprostol represents a problem for safeguarding the quality of misoprostol tablets especially in countries with hot and humid climates. This problem can be reasonably managed by appropriate packaging and by professional formulation of the tablets. The two investigated batches of the originator product, as well as the WHO-prequalified product, were found to remain within the specifications of the International Pharmacopoeia over six months of accelerated stability testing at 40°C and 75% RH, despite the fact that already 11, 16 and 19 months had elapsed since their date of manufacture when they entered stability testing, respectively. The WHO-prequalified product even exceeded its expiry date 2 months before the end of the stability test, and still conformed to specifications both in assay and in dissolution at the end of the six-months testing period. This may be seen as a further example that WHO-prequalification is a reliable assurance of the good quality of medicines. However, of the two tested batches of a product without WHO-prequalification and produced in a country without stringent regulatory authority (SRA), one batch was clearly out of specifications at the end of the accelerated stability testing. The other batch of this manufacturer showed much less degradation during accelerated stability testing. The clearly different results between the two batches may raise doubts about the batch-to-batch consistency in the manufacturing of this product.In this study, misoprostol tablets were included in the stability test only if they were within specifications at the beginning of the test. One brand of misoprostol tablets collected in Rwanda  had to be excluded due to extreme deviations from specifications in the assay. Furthermore, Hall , AnyakorThe observation that several batches investigated in this study remained in specifications during accelerated stability testing does not prove the absence of stability problems. The current ICH/WHO guidelines , 31 requIn the present study, a decrease of the misoprostol content was observed both at 40°C and 75% RH and at 25°C and 60% RH . An inveManufacturers are aware of the stability problems of misoprostol. They frequently manufacture tablets with an initial content of more than 100% of the stated amount, to ensure that the misoprostol content of their preparations does not fall below the pharmacopeial limit of 90% of the declared amount within their shelf-life. This was observed in the present study: the content of all preparations at the beginning of the stability testing exceeded 100% of the stated amount, despite the time which had elapsed since manufacture. The data reported by Hall  showed tIt is encouraging that all brands of misoprostol tablets distributed by wholesalers and government stores in Malawi and Rwanda at the time of sample collection were packaged in double-sided aluminium blisters. This may be a consequence of the report by Hall  that plaBoth the originator product , 41 and For misoprostol tablets packaged in double-sided aluminium blisters, the intactness of the blister is of principal importance for the stability. This was reported by Berard et al.  and has The results of our study show that unprotected storage of misoprostol tablets for extended periods at high temperature and humidity is extremely detrimental, while unprotected storage for one month at 25°C and 60% RH only had a small effect on the misoprostol content. Nevertheless, in view of the instability of misoprostol, storage outside of intact blisters should be avoided, even for short periods.The key stability problem of misoprostol tablets is the degradation of the API, especially in the absence of proper protection from humidity. In contrast, dissolution of misoprostol was hardly affected during accelerated stability testing in this study, and even unprotected storage did not affect dissolution of the remaining amount of misoprostol very strongly see .This study investigated only those brands and batches of misoprostol tablets which were available at government medical stores and pharmaceutical wholesalers in Malawi and Rwanda during the time of sample collection, and may not be representative for other brands or batches. Furthermore, while this approach to sample collection provides a realistic picture of the stability of the products which were distributed in these countries at that time, it does no give a precise picture of the condition of the samples at the time when they left the manufacturing companies. Differences in the results obtained for different brands and batches may be influenced by the different age of samples at time of analysis, and by their different storage condition prior to collection.The stability of misoprostol tablets is problematic and must be ensured by intact, good-quality double-sided aluminium blisters and by appropriate manufacturing using HPMC as stabilizing agent. Misoprostol tablets of very different quality and stability are on the market, and careful supplier qualification is required in the procurement process. In doubt, procurement should be restricted to WHO-prequalified products, and/or products manufactured under supervision of a stringent regulatory authority. In the prevention and treatment of PPH, misoprostol tablets offer the advantage over oxytocin injections that they do not need parenteral administration. However, the notion that misoprostol tablets present fewer stability problems than oxytocin injections may be misleading, especially in hot and humid climates.S1 FigNeedle punctures are highlighted by arrows.(DOCX)Click here for additional data file.S1 Table(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file.


Adverse cardiac remodeling with a myocardial fibrosis as a key pathophysiologic component may be associated to worse survival in aortic stenosis (AS) patients. Therefore, with the application of advanced cardiac imaging we aim to investigate left ventricular myocardial fibrosis in severe AS patients undergoing aortic valve replacement (AVR) and determine its impact with post-intervention clinical outcomes.In a prospective, observational, cohort study patients with severe AS scheduled either for surgical or transcatheter AVR will be recruited from two tertiary heart centers in Denmark and Lithuania. All patients will receive standard of care in accordance with the current guidelines and will undergo additional imaging testing before and after AVR: echocardiography with deformation analysis and cardiovascular magnetic resonance (CMR) with T1 parametric mapping. Those undergoing surgical AVR will also have a myocardial biopsy sampled at the time of a surgery for histological validation. Patients will be recruited over a 2-year period and followed up to 2 years to ascertain clinical outcomes. Follow-up CMR will be performed 12 months following AVR, and echocardiography with deformation analysis will be performed 3, 12, and 24 months following AVR. The study primary outcome is a composite of all-cause mortality and major adverse cardiovascular events.Despite continuous effort of research community there is still a lack of early predictors of left ventricular decompensation in AS, which could improve patient risk stratification and guide the optimal timing for aortic valve intervention, before irreversible left ventricular damage occurs. Advanced cardiac imaging and CMR derived markers of diffuse myocardial fibrosis could be utilized for this purpose. FIB-AS study is intended to invasively and non-invasively assess diffuse myocardial fibrosis in AS patients and investigate its prognostic significance in post-interventional outcomes. The results of the study will expand the current knowledge of cardiac remodeling in AS and will bring additional data on myocardial fibrosis and its clinical implications following AVR.The study has full ethical approval and is actively recruiting patients. The results will be disseminated through scientific journals and conference presentations.ClinicalTrials.govNCT03585933. Registered on 02 July 2018. Since increased afterload is the main pathophysiology of aortic stenosis (AS) leading to the progressive development of left ventricular (LV) structural and functional alterations, AS is regarded a disease of the aortic valve, as well as of the myocardium . The incFurthermore, because the subendocardial layer is the first to be affected in AS, the longitudinal alignment of myocardial fibers in this layer causes decreased longitudinal contraction, an early sign of LV dysfunction. There are data showing, that global longitudinal strain (GLS) correlate with LV myocardial fibrosis and is a predictor of adverse events in patients with severe AS , 17.Novel diagnostic strategies and more accurate evaluation of the disease severity and consequences of AS are needed in the assessment of subclinical myocardial dysfunction and the detection of myocardial fibrosis to challenge current recommendations for the timing of AVR. To date there are limited data on simultaneous assessment of diffuse myocardial fibrosis by noninvasive multimodality imaging and histological confirmation in severe AS. Optimal T1 mapping application also remains unclear, and data assessing its prognostic value in severe AS are still lacking. Our multicenter prospective trial aims to: (i) non-invasively assess myocardial fibrosis and validate it against histological data in patients undergoing surgical AVR and (ii) assess the impact of myocardial fibrosis on clinical outcomes following AVR in both surgical and TAVR cohorts.Primary hypothesis: diffuse and focal myocardial fibrosis in patients with severe AS is associated with worse immediate  and long-term clinical outcomes, all-cause mortality, and major adverse cardiovascular events (MACEs).The presence and extent of myocardial fibrosis are associated with markers of LV decompensation.The presence and extent of myocardial fibrosis have a negative effect on LV reverse remodeling and patient functional recovery following AVR.Secondary hypotheses:To evaluate the prognostic significance of myocardial fibrosis in patients with severe AS undergoing AVR.Primary objective:To identify parameters of multimodality imaging  predictive of LV decompensation.To quantify LV reverse remodeling 12 months following AVR through CMR and echocardiographic measurements.Secondary objectives:. Both sites have received approval from local ethics committees : 158200–18/9–1014-558; for Aalborg University Hospital (AAUH): N-20180081. All patients will give written informed consent.FIB-AS is a prospective, observational, cohort study with clinical endpoints, conducted in two participating sites (in Lithuania and Denmark) from tertiary care hospitals. A total of 110 patients with severe AS undergoing AVR will be recruited from both institutions, and their data will be collected in a dedicated online database, REDCap (Research Electronic Data Capture) . The cho1. The recruitment period will be 24 months, from May 2019 to May 2021. We expect to enroll a total of 110 patients: 60 patients undergoing surgical AVR and 50 patients undergoing TAVR.We will recruit patients with severe AS requiring AVR according to current treatment recommendations . We will1Surgical AVR will be performed by standard midline sternotomy or J mini sternotomy with cardiopulmonary bypass and mild hypothermia. The St. Jude Medical Trifecta aortic bioprosthesis  or mechanical CarboMedics Standard Aortic Valve  prostheses of varying sizes will be used according to the surgical team’s or patient’s preference. TAVR will be performed under conscious sedation. A balloon-expandable Edwards Sapien-3 Ultra Valve system, size 23, 26, or 29 mm , will be deployed. Preferably a transfemoral route of access will be used, when vascular access will be judged suitable.http://www.euroscore.org/calc.html [http://riskcalc.sts.org/stswebriskcalc [The risk of interventional mortality will be assessed by calculating European System for Cardiac Operative Risk Evaluation II (EuroSCORE II) risk scores and Society of Thoracic Surgeons (STS) risk scores using the online calculators available at alc.html  and at hriskcalc . Althougriskcalc . New Yorriskcalc . The Minriskcalc  will be riskcalc .Blood samples will be collected and levels of cardiac biomarkers (B-type natriuretic peptide and troponin I) will be tested at baseline and at each follow-up visit. In addition, blood samples will be taken on the day of CMR scanning to determine hematocrit and creatinine concentration.Comprehensive transthoracic 2D echocardiography will be performed using a commercially available Vivid ultrasound  systems  and stored on a dedicated workstation for subsequent off-line analysis. AS severity, LV systolic and diastolic functions will be evaluated in accordance with the echocardiographic guidelines –28. SimpFrom the 2D grey-scale images of the apical two-, three-, and four-chamber views, left ventricular global longitudinal strain (GLS) will be measured and processed offline using commercially available software  . The fra2 (using the DuBois and DuBois formula).For each patient, baseline and 1-year post-interventional CMR scans will be performed using standard protocols on a 1.5 T Siemens Aera scanner (at VUH) or a 1.5 T GE Discovery MR 450 scanner (at AAUH) with surface coils and prospective ECG triggering , 9. BothFor detection of delayed hyperenhancement, images will be acquired 10–15 min after intravenous administration of Gadobutrol (0.2 mmol/kg)  or Gadoteridol (0.2 mmol/kg)  using a breath-hold segmented inversion recovery fast gradient echo sequence in the short-axis and long-axis planes of the LV, with an 8-mm slice thickness and 20% distance factor. The image parameters (VUH) are: repetition time of 700 ms, echo time 1.42 ms, flip angle 45°, matrix 256 × 184, and field of view 360 mm. For AAUH, the typical imaging parameters are as follows: repetition time 5.79 ms, echo time 2.72 ms, flip angle 15°, matrix 256 × 256, and field of view 380 mm. The optimal inversion time will be optimized for each patient to null normal myocardial signal, ranging from 220 to 320 ms . The region of myocardial fibrosis will be defined as the sum of pixels with signal intensity above 5 standard deviations of the normal remote myocardium in each short-axis slice. The presence of myocardial fibrosis will be qualitatively determined by two independent readers, blinded to clinical data .The present study describes image acquisition parameters by focusing on T1 mapping images, which will be acquired in four-chamber long-axis and short-axis  images before and at 15 min after contrast administration. All T1 mapping images have been acquired using modified Look-Locker inversion-recovery (MOLLI) , 31 usin.The T1 maps will be generated from the CMR workstation after in-line motion correction just after image acquisition (Siemens scanner) or in post-processing software (GE scanner). Two blinded reviewers will independently review all T1 mapping sequences. Regions of interest will be drawn manually in the blood and septum at the midventricular level on the short-axis image, excluding the myocardium with LGE for T1 measurements , 32. RegThe ECV of the myocardium will be calculated as follows: ECV% = (ΔR1m/ΔR1b) x (1 - hematocrit level) × 100, where R1 is 1/T1, R1m is R1 in the myocardium, R1b is R1 in the blood, and ΔR1 is the change in relaxivity. The change in relaxivity (ΔR1) was determined using the following equation: ΔR1 = R1post - R1pre, where R1post and R1pre are the R1 after and before gadolinium chelate administration, respectively . Blood s.In patients undergoing surgical AVR biopsy specimens will be obtained under direct vision by the surgical team using a surgical scalpel from the basal anteroseptum just after removal of the diseased aortic valve. All myocardium tissue samples will be fixed in 10% neutral buffered formalin and embedded in paraffin. Sections of 3-μm thickness will be made on a Leica RM2145 microtome and stained by hematoxylin and eosin, Masson‘s trichrome, and Congo red methods. Digital images will be captured by using the Aperio Scan-Scope XT Slide Scanner  under 20× objective magnification (0.5-μm resolution). Histological slides  will be processed for evaluation of myocardial fibrosis. An experienced (> 25 years of experience) histologist (E.Ž.) blinded to the clinical and CMR data will investigate all biopsy specimens. The fraction of myocardial volume occupied by collagen tissue (collagen volume fraction) will be determined by quantitative morphometry with an automated image analysis system (HALO™) in sections stained with Masson’s trichrome. Quantification of the myocardial fibrosis area will be performed using HALO™ Classifier Module and HALO™ Area Quantification v1.0 algorithms  within manually selected regions of interest enclosing the tissue section and excluding endocardium from the analysis. All study assessments are summarized in Table 2Primary outcome measure: a composite of all-cause mortality and MACEs (time frame: from 30 days up to 24 months following AVR).In-hospital and 30-day all-cause mortality (time frame: 30 days)Length of hospital stayTime to the event (death or MACE) (time frame: 24 months)Cardiovascular mortality (from 30 days up to 24 months following AVR)Secondary outcomes measures:Primary outcome endpoints will be defined according to current guidelines: Valve Academic Research Consortium-2 (VARC-2) definitions  and guid.Patients will be routinely followed and managed according to available guidelines. Clinical outcome data will be collected from patient clinical visits at 3, 12, and 24 months following AVR. Serum biomarker analysis and 2D echocardiography with STE analysis will be performed at each follow-up visit. A second CMR scan will be performed 1 year after AVR. The 6MWT will be performed, and the MLHF questionnaire will be administered at the 12-month follow-up visit. FIB-AS study follows the standard protocol items: recommendation for interventional trials (SPIRIT) guidelines . The stu4Linear regression analysis to model the relationship between the extent of myocardial fibrosis and patient outcomes.Univariate and multivariate regression analysis of CMR-derived predictors of adverse clinical events.Comparison of clinical, serum, and imaging (CMR and echocardiography) biomarker data between patients with different extents of fibrosis.p-value of < 0.05 will be considered statistically significant. The statistical analysis will be performed using the R language and environment for statistical computing (version 3.5.1) [The statistical analysis will be performed under the supervision of V.S. at the Institute of Applied Mathematics, Vilnius University Faculty of Mathematics and Informatics. Descriptive statistics will be used to summarize the demographic, clinical, and imaging data characteristics of each patient. Categorical variables will be presented as the frequency and percentage, and continuous variables will be expressed as the mean and standard deviation or median and interquartile range. The 95% confidence interval will be reported for primary outcomes. Patients will be grouped, depending on the presence and extent of histologically measured myocardial fibrosis, and their clinical and imaging data will be compared for significant differences. Bivariate correlation analysis of the native T1, ECV, and GLS values with the degree of histologically measured myocardial fibrosis will be performed. Pearson or Spearman correlation coefficient will be used as the measure of correlation. To identify independent predictors of the degree of myocardial fibrosis, multivariate stepwise regression analysis will be performed separately for native T1, ECV, and GLS values, along with demographic and clinical data. The composite event will be assessed from a simple Cox model for survival analysis using ECV, native T1, and GLS, and their c-indexes will be compared for how well each marker predicts the clinical outcome. In addition, the Kaplan–Meier method will be used for the cumulative survival analysis with log-rank test to assess statistical differences between the curves of patients with different degree of myocardial fibrosis. A two-tailed n 3.5.1) .A sample size calculation was performed for the primary outcome: a composite of all-cause mortality and MACEs following AVR. To estimate the sample size required for the detection of a hazard ratio for ECV% of 1.32, R package powerSurvEpi was used . CalculaThe ethics application was approved in March 2018. Study enrollment started in May 2019, and recruitment is expected to be completed in May 2021, with a further 24 months for follow-up, post-processing, and close-out of the study. The main study paper will be submitted within 6 months of the study close-out.The study results will be presented to the participating physicians and the general medical community. Following the complete data collection, the manuscript(s) based on the trial results will be submitted to peer-reviewed journals. Authorship criteria as defined by the International Committee of Medical Journal Editors will be followed.FIB-AS is a study intended to investigate the role of multimodality imaging in the assessment of myocardial fibrosis in severe aortic stenosis patients and its prognostic significance in post-interventional outcomes. We sought to assess diffuse myocardial fibrosis by measuring the ECV and native T1 values obtained from CMR with T1 mapping and GLS from STE and to validate these results against the degree of myocardial fibrosis found from the histological examination. In addition, we will explore LV reverse remodeling following aortic valve intervention and its associations with patient clinical recovery. The results of the study will expand the current knowledge of cardiac remodeling in AS and will bring additional data on myocardial fibrosis and its clinical implications following AVR.The present prospective multicenter study is designed to explore associations between left ventricular myocardial fibrosis and clinical outcomes in severe AS patients undergoing AVR. It has clearly established aims, inclusion and exclusion criteria, as well as defined methods and endpoints.The non-invasive measurement of myocardial fibrosis by CMR with T1 mapping is validated against invasive histological assessment.The trial is restricted to isolated AS, excluding cardiac pathologies which could possibly contribute to myocardial fibrosis burden, such as obstructive coronary heart disease and other significant valvular heart diseases.Inclusion of both surgical and TAVR cohorts will allow the investigation of patients with different risk profiles.The selected sample size may be inadequate to allow a subgroup analysis.Limitations of CMR to establish diffuse and focal fibrosis: no reference regions of normal myocardium due to diffuse fibrosis, arbitrary selection of threshold of signal intensity, overlap of T1 values between normal and diseased myocardium.The FIB-AS study excludes patients with comorbidities, such as obstructive coronary artery disease, a history of myocardial infarction, renal failure, and persistent atrial arrhythmias; therefore, our results should not be overgeneralized to a broader AS patient population.


Cardiac function and morphology are known to differ between men and women. Sex differences seen with echocardiography have not been studied systematically in masters athletes.To evaluate sex differences in cardiac structure, function and left ventricular (LV) systolic global longitudinal strain among masters athletes.M = 109, 60 ± 12 years; F = 55, 57 ± 12 years, range 36–91 years) who participated at the 23rd World Masters Athletics Championship held in Málaga, Spain. All athletes underwent state-of-the-art echocardiography including cardiac function, morphology, strain and hemodynamic assessment.This cross-sectional study comprises of 163 masters athletes  due to greater end-diastolic intraventricular septal, LV posterior wall and LV basal diameter. However, LV mass index did not differ between the groups. End-diastolic LV volume and right ventricular area, both indexed to body-surface-area, were greater in men than in women . In contrast, women had higher LV systolic global longitudinal strain  and LV outflow tract flow velocity . Systolic and diastolic blood pressure, LV ejection fraction, and stroke volume index were not different between sexes.Left ventricular mass was higher in male than in female athletes (174 ± 44 vs. 141 ± 36 Cardiac sex differences are present even among masters athletes. Lifelong exercise training does not appear to exasperate morphological difference to a point of cardiac risk or dysfunction in both male and female athletes. Aging is associated with marked changes in heart function and morphology, also known as cardiac remodeling . Left veEmerging evidence from epidemiological and observational studies primarily in male athletes suggest that individuals who participated in lifelong strenuous exercise demonstrate an increased prevalence of atrial fibrillation, myocardial fibrosis, and coronary artery calcification . AdverseCardiac function and morphology as well as cardiovascular risks are known to differ between men and women . Even afCurrent literature dealing with sex differences in cardiac structure and function has focused primarily on young and middle-aged competitive athletes . CurrentData collection took place at the 23rd World Masters Athletics Championships (WMAC) held in Málaga, Spain. A total of 163 masters athletes participated in the present study . Mean age was 58.8 years ranging from 36 to 91 years of age . To be iOn-site recruitment of the participants, study organization and data acquisition was supported by an adapted version of the REDCap electronic data capture tool , 2019. SFollowing a 10-min supine rest, participants underwent standard echocardiography procedures using a Vivid IQ ultrasound machine equipped with a MSC5 cardiac sector probe  on an ultrasound stretcher with thoracic window. Clinical standard views were recorded including parasternal long and short axis , 2-, 3-, 4-, and 5-chamber views. Morphological measurements included volumes and cardiac dimensions. Functional analysis was performed for systolic function (automated detection of ejection fraction), LV systolic global longitudinal peak strain by speckle tracking and stroke volume. For diastolic function, tissue Doppler spectra of the mitral annulus and pulsed wave Doppler spectra of transmitral filling were analyzed for early and atrial filling peak velocities as well as tissue velocities. Right ventricular function was measured by tricuspid annulus plane systolic excursion (TAPSE) and fractional area change (FAC). All echocardiography and hemodynamic examinations and analyses were performed by the same experienced cardiologist .LV Mass (g) = 0.8 × 1.04 × [(LVEDD + IVSd + PWd)^3 – LVEDD^3] + 0.6 (LVOT-SV (ml) = Π × (LVOT-Diameter/2)^2 × LVOT-VTI . Measurements were repeated three times and averaged.MAP (mmHg) = (systolic blood pressure + diastolic blood pressure × 2)/3 and SVR (Wood Units) = /cardiac output, whereas central venous pressure is derived from echocardiography.Moreover, systemic vascular resistance (SVR) and mean arterial pressure (MAP) were calculated using the following equations, respectively, Test of normality was used to check for normal distribution of data. Sex and age differences were compared using multivariate ANOVA . Pearson’s correlation analyses were used to identify relationship between age and cardiac function and morphology within sex. In the case that the normality assumption was violated, Spearman rank correlation was used for correlation analyses. Data are expressed as means ± standard deviation (SD).Power was calculated  retrospep = 0.14). Male athletes had higher body mass index and body surface area than women . Systolic and diastolic blood pressure were not significantly different between men and women . Heart rate at rest was slightly higher in female athletes than in male counterparts (p = 0.06).As shown in p < 0.01), LV diameter (p < 0.01) and posterior wall thickness (p = 0.049) when measured in end-diastole, compared with women. This resulted in a significantly greater LV mass in men . However, when indexing to body surface area, LV mass index did not differ between sexes  than men. Peak dispersion was defined as the standard deviation of the time until peak strain is reached within all myocardial segments and therefore, is a surrogate for the left ventricle’s economic state of systolic contraction. Stroke volume index was similar in both groups (p = 0.42). This was most likely due to significantly higher blood flow velocities in the LV outflow tract in women despite smaller hearts (p = 0.04). For the assessment of systolic right ventricular function, no differences were observed regarding TAPSE or FAC between sexes .Left ventricular systolic function assessed by biplanar ejection fraction did not differ between sexes . Furthermore, E prime  was not influenced by sex (p = 0.19) which resulted in an increased E to E prime ratio in women (p = 0.01).Female athletes expressed higher intracardiac blood flow peak velocities for the early and atrial wave compared with men . Septal and posterior wall diameters were associated with age for both men  and women . This may explain the finding of a strong positive correlation between age and LV mass index for both groups . Moreover, LV contraction became more desynchronized with age, as measured by peak dispersion.As shown in r = −0.29, p < 0.01 and r = −0.29, p = 0.03) and an increase in E to E prime ratio . Atrial filling wave was associated with age in male athletes , but not in female athletes.Diastolic function was inversely associated with age for both male and female athletes, as seen by a decrease in the early filling wave  for both sexes, we observed similar trends as seen in previous correlations analysis. We observed a number of asymptomatic cardiac abnormalities during echocardiography. This includes LV-hypertrophy, atrial dilation, and different valve deficiencies. Furthermore, we found 1 case with dilated and 3 cases with hypertrophic cardiomyopathy as well as one case with suspected cardiac storage disease. Details of observed pathologies can be found within This is the first and largest study to investigate sex differences in cardiac function and morphology via echocardiography in male and female masters athletes on an international scale. We found that female masters athletes exhibited smaller left ventricles but compensated for reduced cardiac volume by increased LV filling pressures resulting in higher intracardiac blood flow velocities and stronger contractions displayed by global strain analysis. Additionally, deterioration of systolic and/or diastolic cardiac function in male and female athletes was found to be absent with aging. This effect is even more pronounced as we could observe morphological adaptation trending toward LV hypertrophy.Similar to previous studies in sedentary adults , increasLeft and right ventricular systolic function and stroke volume index were not different between men and women. However, mean velocity in the LV outflow tract was elevated in women, which resulted in a significantly higher LV systolic global longitudinal peak strain when compared with men. In 119 healthy subjects  Shi et al. reported a significantly higher global longitudinal strain for the endo-, mid-, and epicardial layer of the left ventricle in women between the age of 22–60 years. Beyond 60 years the effect vanishes . This phNone of the quantified LV systolic function metrics were significantly associated with age in female athletes. However, LV peak dispersion, independently associated with LV mechanical dyssynchrony and mortality , was posA comprehensive analysis of 494 subjects ranging in age from 13 to 87 years found a significant positive correlation of LV ejection fraction with age but not sex in absence of cardiovascular disease. In our study masters athletes’ ejection fraction also did not differ between sexes. However, for masters athletes we could not observe a correlation of ejection fraction with age. Furthermore, mean ejection fraction in our study was lower than compared to the corresponding age group  and informed consent was provided by all Human Participants. The patients/participants provided their written informed consent to participate in this study.All authors contributed to the study conception and design. Material preparation and data collection were performed by SM, JT, and FH. Statistical analysis was done by SW and FH. The first draft of the manuscript was written by SW, HT, and FH. All authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Fetal anemia is associated with a hyperdynamic circulation and cardiac remodeling. Rapid intrauterine transfusion (IUT) of blood with high hematocrit and viscosity into the umbilical vein used to treat this condition can temporarily further affect fetal heart function. The aim of this study was to evaluate the short-term changes in fetal myocardial function caused by IUT using automated analysis of cine-loops of the fetal heart obtained by color tissue Doppler imaging (cTDI).Fetal echocardiography was performed before and after IUT. cTDI recordings were obtained in a four-chamber view and regions of interest were placed at the atrioventricular plane in the left ventricular (LV), right ventricular (RV) and septal walls. Myocardial velocities were analyzed by an automated analysis software to obtain peak myocardial velocities during atrial contraction (Am), ventricular ejection (Sm), rapid ventricular filling (Em) and Em/Am ratio was calculated.Myocardial velocities were converted to z-scores using published reference ranges. Delta z-scores (after minus before IUT) were calculated. Correlations were assessed between variables and hemoglobin before IUT.p = 0.036) and LV Em/Am  z-scores and hemoglobin before IUT. The peak myocardial velocities that were increased before IUT decreased, whereas LV Em/Am increased significantly after IUT.Thirty-two fetuses underwent 70 IUTs. Fourteen were first time transfusions. In the LV and septal walls, all myocardial velocities were significantly increased compared to normal values, whereas in the RV only Sm was increased before IUT (z-scores 0.26–0.52). In first time IUTs, there was a negative correlation between LV Em (rho = − 0.61, This study showed that peak myocardial velocities assessed by cTDI are increased in fetuses before IUT reflecting the physiology of hyperdynamic circulation. In these fetuses, the fetal heart is able to adapt and efficiently handle the volume load caused by IUT by altering its myocardial function. Chronic anemia increases the demands on the fetal heart to increase cardiac output and maintain adequate tissue oxygenation. This hyperdynamic cardiovascular adaptation is characterized by cardiomegaly, increased blood flow velocities, decreased vascular resistance and, if left untreated, cardiac dysfunction, hydrops and eventually death –4. To prCurrent knowledge about cardiac function in anemic fetuses undergoing IUT is mainly derived from assessment of blood flow, whereas surprisingly little is known about the performance of the myocardium. Decreased cardiac output has been demonstrated following IUT when assessing blood flow, but data also indicate that the decrease might be transient –9. Only Color tissue Doppler imaging (cTDI) is an echocardiographic technique evaluating myocardial motion that can be used to derive information on variables describing fetal cardiac function such as longitudinal myocardial velocities and cardiac cycle time intervals –14. DespThus, the aim of this study was to evaluate myocardial function before and after IUT using cTDI with an automated analysis software to identify physiological changes caused by short-term volume load in fetuses under-going transfusion.Pregnant women undergoing IUTs were recruited from the Center for Fetal Medicine at Karolinska University Hospital between 2009 and 2016. Exclusion criteria were multiple pregnancy, major fetal structural/chromosomal abnormality or intraperitoneal transfusion. Gestational age was based on measurement of crown rump length or biparietal diameter in the first trimester or biparietal diameter in the second trimester according to local guidelines. The study was approved by the Stockholm Regional Ethics Committee (DNr 2012/895–31/4 and 2009/1617–31/2) and all women gave their informed consent.Maternal characteristics, medical history and perinatal outcome were recorded. A transabdominal ultrasound examination was performed for the estimation of fetal weight and measurement of peak systolic velocity in the middle cerebral artery (MCA PSV) using a Voluson E8 ultrasound machine with a 2–5 MHz curved array transducer . MCA PSV was recorded and adjusted to multiple of median (MoM) for each gestational age to non-invasively assess the likelihood of fetal anemia as described by Mari et al. , 19. FetIUT was performed according to standard techniques and the local protocol for intravascular transfusions has been described previously . Brieflyo). Off-line analysis was performed in EchoPAC version 201 . Regions of interest (ROIs) were placed at the atrioventricular plane (AV-plane) in the left (LV) and right ventricular (RV) walls and the interventricular septum (IVS)  by two experienced operators (KFW and LH) using a GE Vivid-i ultrasound imaging system with a 3S-RS (1.9–3.8 MHz) phased array transducer  or a Vivid S6 ultrasound imaging system with a M4S-RS (1.9–4.1 MHz) phased array transducer . All cTDI recordings were obtained in an apical or basal four-chamber view and cine-loops of consecutive cardiac cycles were stored for off-line analysis. Acquisition was optimized to obtain a high frame rate and the incident angle was kept as close to the long-axis of the heart as possible , as a text file covering all available velocity data and further analyzed by an automated algorithm also developed into an automated analysis software. This is a fully automated procedure and no manual marking of the traces is needed once the velocity traces are transferred to MATLAB. Moreover, the algorithm compares all identified heart beats and uses that information in identification of cardiac time events. It can also compare different segments, i.e. the IVS, LV and RV walls, over time for more exact definitions –25.The peak myocardial velocity during atrial contraction (Am), ventricular ejection/systole (Sm) and rapid ventricular filling/early diastole (Em) were automatically obtained Fig. . The Em/Data analysis was performed using IBM SPSS Statistics for Windows, version 23.0  and MATLAB. Continuous variables were presented as mean ± standard deviation (SD) or median (interquartile range) as appropriate. Categorical variables were presented as absolute values and percentage [n (%)].All cTDI variables were converted to z-scores [(measurement – predicted mean)/ predicted SD] by using published gestational age specific reference data obtained from normal pregnancies by the same technique . Delta  were presented on measurements before IUT in all IUTs to assess differences between values compared to reference data. Spearman’s correlation (rho) was calculated between the Hb and MCA PSV MoMs before IUT, on one hand, and cTDI z-scores on the other hand. Changes in variables before and after IUT were assessed using Wilcoxon signed rank test for related samples for first and all IUTs. Statistical significance was set to This study included 32 singleton fetuses that underwent a total of 70 IUTs with a mean of 2.2 ± 1.7  transfusions per fetus. cTDI recordings were obtained both before and after IUT in 66, and in four only before IUT. Fourteen of the 70 IUTs were first time transfusions, whereof one was not examined after IUT. In one of these first transfusions there was no hemoglobin value available before IUT. For details see Fig. n = 22) of which nine in combination with another antibody. Five fetuses underwent seven transfusions due to other causes. One fetus had a suspected Diamond-Blackfan anemia that was postnatally confirmed. Postnatal investigation showed suspicion of Chronic Infantile Neurological and Auricular syndrome in one baby but this was not genetically verified [Demographic details and clinical characteristics of the study population, transfusions and pregnancy outcome are shown in Table verified . In threThere were 28 preterm deliveries but only two women delivered before week 34 + 0. Fetal hydrops was present at three (4.3%) IUTs in three fetuses. Isolated ascites was present at eight (11.4%) IUTs in six fetuses.Of the 136 ultrasound examinations performed before or after IUT, the position of the heart was apical in 59 (43%) scans and basal in 77 (57%) scans. The frame rate was 207.7 ± 13.1 frames/sec with a minimum of 173 frames/sec. When extreme outliers in cTDI variables were identified the individual cTDI recordings and velocity traces were visually inspected. Values were omitted if the trace was of insufficient quality for the functioning of the algorithm. This together with missing values due to fusion of Em and Am waves resulted in a mean of 68 observations per variable before IUT . An average of 9.1 ± 1.7 cardiac cycles were analyzed per myocardial velocity trace.Myocardial velocities before all 70 IUTs are presented in Table p = 0.036) as well as LV Em/Am .When analyzing observations made before first IUTs in the 12 cases with available Hb values and cTDI measurements, we observed a negative correlation between Hb and LV Em  as well as LV Em/Am , the median Hb MoM was 0.60 . Three fetuses had mild anemia, six fetuses had severe anemia and three fetuses had no anemia. In the sub-group of anemic fetuses (n = 9) the only variables that showed a significant change after transfusion was Sm which decreased from 1.84 cm/s to 1.53 cm/s in the LV (p = 0.05) and 2.50 cm/s to 1.74 cm/s in the IVS (p = 0.017).In the group of fetuses receiving their first time transfusion that had Hb values as well as pre- and post-transfusion cTDI measurements recorded (p = 0.001) following transfusion. All peak myocardial velocities except Am and Em in the RV decreased significantly after IUT . The LV Em/Am ratio increased after IUT  whereas the other ratios did not change. Table When assessing all IUTs with available data before and after IUT, we found a significant decrease in heart rate from 140.4 ± 9.1 bpm before to 132.7 ± 14.4 bpm (This study showed that all peak myocardial velocities in the LV and IVS and Sm in the RV wall are increased before IUT compared to normal values, reflecting the hyperdynamic circulation. The same myocardial velocities, as well as the heart rate, also decreased significantly after IUT. We also demonstrated a negative correlation between Hb concentration and LV Em and LV Em/Am before transfusion in all IUTs and more prominently when only first time IUTs were assessed. This study also showed that the effect of IUT on the fetal myocardium can be assessed using cTDI with automated analysis.The increase in myocardial velocities before IUT, is in accordance with the hyperdynamic circulation associated with anemia that has been described by other investigators recording fetal blood flow velocities , 27, 28.We observed a decrease of systolic and diastolic myocardial velocities and a significant increase in LV Em/Am ratio after IUT in accordance with the findings reported using fetal cardiac blood flow velocity variables . A decreThe strength of this study is that it describes fetal cardiac physiology in fetuses before and after IUT with a focus on myocardial function that has not been well studied. It also demonstrates the potential of automation in facilitating the use of cTDI in the evaluation of fetal myocardial function that can be applied as a tool for the assessment of fetal cardiac physiology under different conditions.Our study does have some limitations. Multiple measurements were performed in the same fetus when evaluating all IUTs, which may introduce some bias when assessing the correlation between myocardial velocities and degree of fetal anemia. Therefore, we evaluated first IUTs separately even though the numbers were few. The study group was also heterogeneous with varied etiologies and varying degrees of anemia that might influence the results. It is also important to acknowledge that evaluation of cardiac function has been performed close after, and with varying intervals from, the IUT and that several investigators did not demonstrate changes in cardiac variables when longer time had elapsed , 9, 34. However, the current study demonstrated that the rapid physiological adaptation at fetal transfusions well can be monitored by the automated method we utilize in the present study. It remains to be established how this promising method will perform in more chronic disorders such as intrauterine growth restriction. Preliminary studies suggest that the method is sensitive enough to evaluate physiological adaptation at this condition.This study showed that peak myocardial velocities assessed by cTDI are increased in fetuses before IUT reflecting the physiology of hyperdynamic circulation. It also showed that the fetal heart is able to adapt and efficiently handle the volume load caused by IUT by altering its myocardial function.With advances in automation, cTDI could be more readily applicable to study changes in fetal cardiac physiology.


Deposition of amyloid in the heart can lead to cardiac dilation and impair its pumping ability. This ultimately leads to heart failure with worsening symptoms of breathlessness and fatigue due to the progressive loss of elasticity of the myocardium. Biomarkers linked to the clinical deterioration can be crucial in developing effective treatments. However, to date the progression of cardiac amyloidosis is poorly characterized. There is an urgent need to identify key predictors for disease progression and cardiac tissue function. In this proof of concept study, we estimate a group of new markers based on mathematical models of the left ventricle derived from routine clinical magnetic resonance imaging and follow-up scans from the National Amyloidosis Center at the Royal Free in London. Using mechanical modeling and statistical classification, we show that it is possible to predict disease progression. Our predictions agree with clinical assessments in a double-blind test in six out of the seven sample cases studied. Importantly, we find that multiple factors need to be used in the classification, which includes mechanical, geometrical and shape features. No single marker can yield reliable prediction given the complexity of the growth and remodeling process of diseased hearts undergoing high-dimensional shape changes. Our approach is promising in terms of clinical translation but the results presented should be interpreted with caution due to the small sample size. Amyloidosis occurs when proteins that take abnormal forms known as amyloid deposits build up in the tissues. These deposits are composed of abnormal protein fibers that accumulate more quickly than they are cleared away, and thus interfere with the structure and function of affected organs throughout the body. These include the heart, liver, skin, lungs, kidneys, and nervous system  with amyloid becomes firm, rubbery and stiff, similar to hypertrophic cardiomyopathy  imaging.Using Doppler echocardiography, Koyama et al. found that the early impairment in systolic function of a cardiac amyloidosis heart can be reflected by changed longitudinal strain and strain rate  and at 6 or 9 months after the treatment (follow-up). The information of all patients is in dt at time t. Following this, the most-basal short-axis image is moved toward the annuls ring along the long axis with a distance of dt − min(di), . Note that the long axis is defined by connecting the center of the LV base and the apex. In order to align the LV geometries circumferentially, the angles of right ventricular insertion points are defined in the basal plane , v1 near the inferior segment, and v2 near the anterior segment, as shown in v1, and the same number of elements are assigned to the septum circumferentially when generating the layered hexhedron mesh. A prolate spherical coordinate system is used to reconstruct the LV geometry following the steps in Liu et al. . Short-ao at the epicardium to 60o at the endocardium, and the sheet angle from −45o at the epicardium to 45o at the endocardium  locally with respect to the long axis, and the transmural direction (r) pointing from endocardium to epicardium, and then the local longitudinal direction l = r × c, which follows local geometrical curvature  can be readily calculated as We use the Holzapfel-Ogden strain energy function to describe myocardial passive properties  are invariants of the right Cauchy-Green tensor. A more detailed description of the model (1) can be found in Holzapfel and Ogden  and PED at end-diastole (t = 1 s), following  are inferred by using an optimization algorithm through minimizing an objective function . Sensitivity analysis in our previous study :This allows us to divide the parameters in Equation (1) into two groups, the first group includes o et al.  for healnlayer is the number of layers considered, nlayer = 3 indexed by k; ntime is the number of time steps, ntime = 13 indexed by j; nreg is the number of regions in each layer considered, nreg = 20 indexed by i; wv, wE are the weights. Note we do not include the longitudinal strains in our objective function because the uncertainty of estimating longitudinal strains is much larger than the circumferential strains. This is because the simplified long-axial alignment we use does not account for the out-of-plane motion along the long axis. We are unable to quantify the out-of-plane motion due to the lack of necessary features in cine images. 3D strain imaging such as tagging allows one to include longitudinal strains  shows that bsbfs. Let ξ = bf/bfs, we writeIn the second equation of (6), two unknowns remain, FE in Equation (5).It is easy to see that ξ ∈  from data in Gao et al.  and PaliThe flowchart of the two-step optimization method and the inferred parameters are given in For feature comparison between the amyloidosis patients and control, we make use of the LV geometries of 26 healthy subjects from our previous study  relies on, successively, finding the principal directions of variation in the data where the amount of variation explained by the eigenvectors  can be quantified using the corresponding eigenvalues Bishop, . If we b Shlens, . Similar Shlens,  providesFor the purpose of classification, we can first consider a supervised method similar to PCA, namely linear discriminant analysis (LDA) Bishop, . WhereasTable 5, and The optimization procedure is given in PED, the p-V curves estimated from the FE models are shown in Based on the wall-thickness scaled The myocardium stress-stretch response along the myofiber direction for each patient can now be obtained from a pseudo uni-axial test of the myocardium using the material parameters estimated with perfectly aligned myofibers in one direction. The results are plotted in p < 0.05 for a test of a difference between median wall thickness of healthy volunteers and amyloidosis patients using Mood's median test (these results are consistent with a t-test and a rank sum test).PCA permits an intuitive representation of the variations in the data via the modes of variation See , obtaine1 score:Before studying amyloidosis progression prediction, we perform classification of the seven amyloidosis patients at baseline compared with a set of 26 healthy volunteers based on geometries alone, assessing the performance of LDA and a kernel SVM. Sensitivity  and specificity  scores for these methods, obtained using a leave-one-out cross validation (LOOCV) procedure, are given in TP is number of true positives, FP is number of false positives and FN is number of false negatives. The F1 score is the harmonic mean of the precision  and the recall . We use the harmonic rather than the arithmetic mean to penalize the improvement of one of the scores at the expense of the other.where Accuracy of both the linear and non-linear methods is encouraging, suggesting the discriminative properties of the LV geometry with regards to amyloidosis.The analysis above shows that no single marker provides a clear indication for disease progression. A combination of multiple features must be considered. To this end, we summarize the representative markers below that may contribute to the growth and remodeling of myocardium.Vwall/VLV, which reflects the wall thickness change relative to the ventricular volume. A lower value of Vwall/VLV means the wall becomes thinner (recovery);cc, the circumferential strain at end-diastole averaged from the three planes. A larger value of Ēcc indicates that the LV is more compliant in the circumferential direction (recovery);Ēll, the longitudinal strain at end-diastole averaged from the three planes. A larger value of Ēll indicates that the LV is more compliant in the longitudinal direction (recovery);ĒW, work done by pressure during diastolic filling, because it is not straightforward to compare different p-V curves, we compare the area-under-the-curve, which is defined asW in the follow-up means more work is required to maintain the heart function, corresponding to worsening.A higher value of The average slope Shape features, obvious shape changes compared to control indicate worsening.The markers used for follow-up cardiac amyloidosis status prediction are divided into three classes: (1) geometrical markers (features 1–3), (2) biomechanical markers (features 4–6), and (3) LV shape markers (feature 7). Geometrical markers include normalized LV wall thickness, which has been applied clinically. Biomechanical markers are discussed in section 3.2.We first study the amyloidosis patient recovery using shape features. As we do not know what is a “healthier” shape for LV, we make use of the control data from our previous study , that matters in our evaluations. Clearly, Other modeling limitations should also be mentioned. In this paper, amyloidosis LV is regarded as homogeneous material. The loaded early diastolic configuration is used as the reference configuration which excludes the effect of residual stresses. Our alignment of LV geometries at different times is based on a simplified linear registration approach. Nonlinear methods, such as deformable registration approaches . All research-related procedures were performed in accordance with local guidelines and regulations. The patients/participants provided their written informed consent to participate in this study.XL, HG, and DH designed the study. WL and AL developed the mechanical and statistical models. AM-N, MF, and PH acquired the CMR scans. SB, RJ, and JC initiated the research idea and helped with image analysis. CB provided clinical insight. All participated in writing the manuscript.RJ and JC were employed by the company GlaxoSmithKline, plc, UK. The University of Glasgow holds research and/or consultancy agreements with Siemens Healthcare and GlaxoSmithKline plc for work done by XL, CB, and others.


Pulmonary arterial hypertension (PAH) is a rare, progressive disease with a poor prognosis. The pathophysiologic model is mainly characterized by an afterload mismatch in which an increased right ventricle afterload, driven by increased pulmonary vascular resistance (PVR), leads to right heart failure. International guidelines recommend optimization of treatment based on regular risk assessments to achieve or maintain a low-risk status. Current risk scores are based on a multi-modality approach, including demographic, clinical, functional, exercise, laboratory, and hemodynamic parameters, which lack significant echocardiographic parameters. The originality of echocardiography relies on the opportunity to assess in a non-invasive way a physiologically meaningful combination of easy to measure variables tightly related to right ventricle adaptation/maladaptation to increased afterload, the main determinant of a patient’s prognosis. Echo-derived morphological and functional parameters have been investigated in PAH, proving to have prognostic relevance. Different therapeutic strategies proved to have different effects in reducing PVR. An upfront combination of drugs, including a parenteral prostacyclin, has shown to be associated with right heart reverse remodeling in a greater proportion of patients than other treatment strategies as a function of PVR reduction. Adding echocardiographic data to current risk scores would allow better identification of right ventricle (RV) adaptation in PAH patients’ follow-up. This additional information would allow better stratification of the patient, leading to optimized and personalized therapeutic management. Pulmonary arterial hypertension (PAH) is a rare, progressive disease with a poor prognosis. It is characterized by the remodeling of small pulmonary arterial vessels with a subsequent increase in pulmonary vascular resistance (PVR). This increase leads to right ventricular (RV) failure and ultimately to death . InternaAn accurate and regular risk assessment allows clinicians to follow their patients over time, update treatment goals, and identify the correct timing for lung transplantation.REVEAL was derived from a cohort of 2716 PAH patients with all subgroups and then validated internally and externally in registries and clinical trials datasets, providing the probability of survival at 12 months in the incident and prevalent IPAH patients and in patients with associated PAH [It was subsequently upgraded to REVEAL 2.0 with the modification of cut-off and relative risk points of several variables and the introduction of two further variables, including overall 13 parameters , blood pressure, heart rate, all-cause hospitalization within the previous 6 months, 6 min walking distance (6MWD), brain natriuretic peptide (BNP) or N-terminal fragment of pro-BNP (NT-proBNP), pericardial effusion, % predicted diffusing capacity of the lungs for carbon monoxide, right atrial pressure (RAP), PVR). The sum of the various parameters’ risk points determines the final score that identifies the patient’s risk class.On the other hand, European registries based their scoring system on the parameters and relative thresholds included in the European Society of Cardiology and European Respiratory Society (ESC/ ERS) guidelines.COMPERA and SPAHR estimate the overall risk category by computing the mean of the risk grades from available variables for each patient and rounding to the nearest integer. The variables considered in the latter score are WHO FC, 6MWD, BNP or NT-proBNP, RAP, cardiac index, and mixed venous oxygen saturation. A different approach characterized the FPHR, which considered the number of low-risk variables achieved among 6MWD, WHO FC, RAP, and cardiac index, BNP or NT-proBNP.REVEAL 1.0 and 2.0 are broadly applicable to the general PAH population, as they predict the risk in both incident and prevalent patients with various PAH etiologies. SPAHR and COMPERA were derived from incident patients with only idiopathic, heritable, drug-induced PAH or connective tissue disease-PAH. In FPHR, risk assessment was performed in incident IPAH patients.All these scores have the advantage of being multiparametric and based on simple parameters that are easy to obtain, allowing a patient’s risk stratification into three classes (low-intermediate-high).On the other hand, these scores have significant limitations, such as the relatively short mortality prediction period, the large number of variables needed for the REVEAL score, and the “a posteriori” nature of the validating analyses in the case of the European risk scores. Furthermore, standardization of data collection was not guaranteed in all published registries, with a significant loss of data.All these scores are based on survival and do not consider clinical worsening and disease progression, which do not match with clinical goals to be achieved according to current guidelines .Moreover, most patients end up in the intermediate-risk category, characterized by a heterogeneous population with different outcomes, representing a missed opportunity for treatment optimization and personalized medicine .The PAH pathophysiologic model is mainly characterized by an afterload mismatch, where increased RV afterload, driven by increased PVR, leads to right heart failure.At an early stage, the RV adapts to the increased afterload to preserve stroke volume (homeometric adaptation), followed by an heterometric adaptation when the latter gets exhausted, characterized by increased filling pressures, progressive RV dilation, and hypertrophy in order to reduce wall tension (Laplace’s law), leading to eccentric hypertrophy and right heart failure . Adapted ventricles usually have preserved ventriculo-arterial coupling (a condition that occurs when the right ventricular function is adapted to the pulmonary vascular load so that energy transfer is most efficient). In contrast, maladapted ventricles usually are characterized by uncoupling of the RV to the pulmonary circulation ,9,10.The RV remodeling process in PAH is complex, as it results from the interaction of several factors, and the precise mechanisms determining the transition from adaptation to maladaptation and finally failure are not yet fully understood. However, what contributes most likely to these changes is the interaction between genetic factors, neurohormonal activation, etiology and duration of pulmonary hypertension over time, RV ischemia, and the amount of afterload increase ,12,13,14It is now clear that patients’ clinical status and prognosis depend on the RV’s ability to adapt to the increased afterload, maintaining its function and preserving cardiac output ,16.The originality of echocardiography relies on the opportunity to assess a physiologically meaningful combination of easy to measure variables tightly related to RV adaptation/maladaptation to increased afterload. Echo-derived systolic functional parameters are robust though load-dependent measures of contractility, while right heart dimensional parameters reflect increased RV and RA pressure and dimensions. Thus, the RV Starling’s heterometric adaptation that comes into play in the advanced stage of the disease is easily detected by echocardiographic evaluation. Echo-derived systolic functional parameters reflect contractility adaptation and increased right heart, and inferior vena cava dimensions its failure . MagnetiEchocardiography is the most user-friendly and widely used tool for assessing RV structure and function in clinical practice. It is a rapid, low-cost exam with an extremely favorable cost/benefit ratio. It may help in the non-invasive assessment of the hemodynamic RV burden in PAH, allowing a more detailed and precise analysis of the patient’s condition compared to the common clinical examination ,20.Considering the close correlation between ventricular morphology and function and the pathophysiological basis of PAH, it is clear that the introduction of echocardiographic parameters within the current risk scores would provide a greater opportunity to improve prognostication ,22.However, the initial risk scores in PAH were based on an expert clinician experience-derived multi-modality approach, in which echocardiographic parameters were limited only to RA surface area and pericardial effusion , or in oDue to the complex and irregular right heart shape, RV and RA dimensions and their derived functional parameters—ejection fraction (EF), stroke volume, and RV fractional area change (FAC)—are all elaborated according to geometric models that approximate the RV’s shape. Nevertheless, all measurements that act as surrogates of true right heart volumes are widely used in everyday clinical practice. They are representative of the RV’s afterload mismatch condition and can help the clinician in routine patient management due to their ability to monitor disease progression and response to therapy. Studies showed that stable patients with reassuring WHO FC and 6MWD at follow-up develop a progressive deterioration of RV EF and dimensions . Indeed,Dimensional RV assessment is followed by RV dysfunction evaluation defined by the echocardiographic measurement of FAC, tricuspid annular plane systolic excursion (TAPSE), tissue Doppler and myocardial strain recording of RV free wall motion, all of them representing isovolumetric and ejection phase indices of loading-induced RV pump failure. Due to its simplicity of acquisition, TAPSE, despite being a load-dependent and angle-dependent parameter reflecting mainly the lateral wall longitudinal motion, is the most popular measure used as a surrogate for RV longitudinal systolic function, with a high proven prognostic significance ,32,33,34The ratio of TAPSE to pulmonary artery systolic pressure (TAPSE/PASP) has been validated against the ratio of end-systolic to arterial elastances (Ees/Ea) , which dFAC is a load-dependent parameter, highly correlated with RVEF assessed on an MRI, representing the transversal component of RV contraction . HoweverRV isovolumic contraction peak velocity (IVCv) and dP/dt (max) are isovolumetric phase parameters used to assess RV contractility. As all isovolumetric phase parameters, they have the advantage of being less affected by afterload. On the other hand, because of the increased RV contractility , these pAnother prognostic relevance parameter is the RV myocardial performance index, which analyzes both RV systolic and diastolic function. However, this parameter has a high degree of inter, and intraobserver variability is load-dependent and unreliable when R-R intervals are irregular ,41,45.RV filling pressure can be estimated using inferior vena cava diameter and collapsibility, the presence of pericardial effusion, and the ratio between the early diastolic myocardial velocity at the lateral tricuspid annulus (E) and early diastolic tricuspid inflow (E’). All these measurements have proven to correlate with hemodynamic values, showing prognostic relevance in PAH patients ,46,47,48Speckle tracking and myocardial deformation (strain and strain rate) are emerging techniques that have shown promising RV contractility evaluation results. Although regional and global 2D strains are load and angle-dependent measurements , severalInhomogeneity in time to peak myocardial strain, known as RV dyssynchrony, also influences RV function, potentially to a significant extent independent from systolic function itself and showed a prognostic role in predicting clinical worsening ,55.The evaluation of right heart reverse remodeling (RHRR) defined in two-dimensional echocardiography by combining decreased right-sided atrial and ventricle areas and left ventricle eccentricity index, has recently emerged as a key parameter in evaluating patients’ response to therapy. Indeed, a dramatic reduction in PVR in response to therapy has been shown to be associated with RHRR, which in turn is strongly associated with improved outcomes ,57.However, while clinicians are becoming used to collecting echocardiographic variables for clinical practice, the most useful RV indices have been a source of debate for several years. Most echo-derived parameters lack validation in prognostic relevance, needing further investigation with dedicated multicentric prospective studies to establish solid and clinically relevant cut-off values. According to RV failure’s pathophysiologic basis in PAH, parameters like TAPSE/PASP, FAC, and RHRR represent promising parameters to improve the accuracy of current risk tools in the multi-modality assessment. Understanding the complex interactions among RV size, systolic function, and clinical parameters will require accurate and reproducible research methods that reasonably encourage collaboration between centers.The 6th World Symposium on PH recommends that most low- and intermediate-risk patients be treated with two oral drugs targeting NO signaling and endothelin pathways, while triple upfront therapy with parenteral prostacyclins is reserved for high-risk patients .Indeed, the impact of oral monotherapies on RV morphology and function have been proven to be trivial. The BREATHE-1 echo-substudy, the only randomized trial on oral monotherapy, showed trivial beneficial effects of bosentan on echo-derived RV dimensional or systolic functional parameters . SimilarDouble oral therapy showed an additional effect on PVR reduction compared with oral monotherapy, with a reduction ranging between 30 and 60% ,62,63,64In Sitbon et al.’s study,  triple uSuch RHRR is associated with excellent long-term survival and quality of life. However, studies showed that monotherapy or stepwise additions to initial monotherapy allow these results in a small percentage of patients (18%) ,61,63,67Notably, in the study of D’Alto et al. , no sign


The acquisition of the ventricular short axis cine stack forms the backbone of the routine cardiovascular magnetic resonance scan . From thThe purpose of this study was to assess the practicality of using a two-slice per breath-hold ventricular short axis cine sequence in routine daily practice.From the beginning of March 2008 to September 2008, we used a two-slice per breath-hold steady state free precession sequence to acquire the ventricular short axis stack of cines in a total of 478 patients. All patients were scanned with a 1.5 T Siemens scanner  using anterior phased-array coils and ECG gating. Sequence parameters for the SSFP cine were as follows: 2 slices (8 mm slice thickness), 25% distance factor (2 mm gap), TR 40.2 ms, TE 1.13 ms, flip angle 80°, base resolution 192, number of signal averages 1, parallel imaging , bandwidth 930 Hx/Px, echo spacing 2.7 ms. Ten patients had both two-slice and single slice per breath-hold cine stack acquired. The parameters for the single slice acquisition were identical to those given above apart from using a 7 mm slice thickness with 3 mm gap.Since the beginning of the study, there have been no problems with the acquisition of the short axis stack using this technique. The images have all been suitable for analysis and calculation of ventricular volumes despite slightly increased partial volume effects towards the apex as a result of using an 8 mm slice thickness. In patients who had undergone both techniques, there was a low coefficient of variation between the two with no significant difference in volume or ventricular function calculations (see Figure In conclusion, the use of a two-slice per breath-hold cine acquisition is a practical method for use in daily practice. This shortens the time required for the whole ventricular short axis cine stack and allows a streamlining of workflow with overall reduction in the time taken for each CMR study. There is no significant difference in the ventricular volumes, mass or ejection fraction calculated.


Multiple logistic regression analysis revealed that LVGFI was an independent predictor of CA . In conclusion, LVGFI is a novel and clinically useful parameters with excellent ability in determining myocardial function and differentiating cardiac amyloidosis from hypertrophic cardiomyopathy.Differentiating cardiac amyloidosis (CA) from hypertrophic cardiomyopathy (HCM) remains a clinical challenge, particularly in those with preserved left ventricular ejection fraction (LVEF) and similar hypertrophy. This study aimed to use left ventricular global function index (LVGFI) and myocardial contraction fraction (MCF) to discriminate CA from HCM without using contrast agents on cardiovascular magnetic resonance imaging (CMR). In total, we included 68 CA patients, 90 HCM patients, and 35 healthy controls. We found that LVGFI had excellent diagnostic performance in differentiating CA from HCM (area under the curve (AUC) = 0.91, 95% CI [0.86–0.95]), even in the challenging conditions of similar hypertrophy  and preserved LVEF . LVGFI also had significant correlations with LGE extent, NT-proBNP and troponin T (all  Differentiating CA from LVH caused by other etiologies such as hypertrophic cardiomyopathy (HCM) remains a clinical challenge, particularly in those patients with preserved left ventricular ejection fraction (LVEF) and similar hypertrophy. Early diagnosis of CA is critical for the selection of appropriate therapy and improvement of prognosis3.Cardiac amyloidosis (CA) is a progressive and infiltrative cardiomyopathy with poor prognosis. Left ventricular hypertrophy (LVH) is a typical sign of CA5. However, CA patients frequently suffer from renal failure, either due to amyloid deposition in the kidneys6 or due to reduced cardiac output from heart failure (HF), which makes them contraindicated to contrast agents7. In addition, early-stage CA may not present with typical LGE patterns. Thus, using a non-contrast method for diagnosing CA and avoiding the contrast-induced renal injury seem extremely important.At present, late gadolinium enhancement (LGE) of cardiovascular magnetic resonance imaging (CMR) is of great diagnostic value in diagnosing CA in clinical settings8; therefore, it is not an optimal marker for the assessment and discrimination of CA from other types of hypertrophy9. Although LVEF may be remarkably high in some hypertrophic patients, they may have smaller LV volumes result from thickened walls and their higher LVEF does not necessarily produce sufficient cardiac output10.Whereas conventional non-contrast index LVEF is often preserved or ‘normal’ in patients with myocardial hypertrophy and HF symptomset al. and King et al., are two indices that combine global systolic performance with anatomic information12. They are derived from non-contrast cine CMR images and do not require additional post-processing software. In the present study, it was hypothesized that compared with conventional indices, LVGFI and MCF provide better evaluation of myocardial dysfunction and are better at discriminating CA from the other patients with thickened ventricular walls.Left ventricular global index (LVGFI) and myocardial contraction fraction (MCF), initially introduced by Mewton The purpose of this study was to evaluate LVGFI and MCF in their ability to discriminate CA from HCM without using contrast agents on CMR. To our knowledge, this is the first study that used LVGFI to aid in the diagnosis of thickened hearts.p < 0.001 for both). Nine (11.7%) and seven (7.8%) patients in the CA and the hypertrophic control groups had a history of atrial fibrillation, respectively. Healthy subjects were slightly younger than the two groups of patients .Demographic and clinical characteristics of 158 included patients and 35 normal controls are presented in Table p = 0.64), whereas mWT was higher in the HCM group than in CA (p = 0.01). LVGFI and MCF were significantly higher in healthy controls . And 58.8% CA patients had preserved LVEF (LVEF ≥ 50%). CA patients displayed remarkably higher LGE extent than HCM (p < 0.001). LVGFI and MCF were correlated with LGE extent . Furthermore, Correlation analyses revealed that LVGFI and MCF also had strong associations with troponin T  and NT-proBNP levels .LV morphological and functional parameters are presented in Table A p = 0.0. LVGFI ap = 0.011; MCF vs LVEF, p = 0.256; LVGFI vs quantified LGE, p = 0.68). Multiple logistic regression analysis revealed that LVGFI , LGE , log NT-proBNP  and log Troponin  were independent predictors of CA.ROC analyses for discriminating the two patient groups from the normal subjects is presented in supplementary Table Diagnostic values of LVGFI and MCF in patients with similar hypertrophy and preserved LVEF (≥50%) are presented in Table Twenty subjects were reanalyzed by a second investigator who was blinded to clinical information to assess the interobserver agreement. The results of all the included parameters are shown in Table The major findings of the present study were as follows: (1) LVGFI and MCF are two novel parameters that could determine myocardial function in ventricular hypertrophy, (2) LVGFI was better than LVEF, and comparable to quantified LGE at differentiating CA from HCM; (3) LVGFI still demonstrated outperformance under difficult diagnostic conditions of similar hypertrophy or preserved LVEF.et al. as a three-dimensional, volumetric measure of myocardial shortening and was used to distinguish between patients with hypertensive heart disease (HHD) and athletes with physiologic hypertrophy12. From a mathematic perspective, SV is a measure of the difference between end-diastolic and end-systolic volumes that determine the output of blood from each myocardial contraction. Because it is the myocardium that contracts during systole, by dividing SV by myocardial volume, MCF represents an index of myocardial contraction that is not influenced by volume. Arenja et al.13 assessed MCF in patients with heart failure and LVH and showed that MCF outperformed LVEF and LVMI in discriminating CA from other forms of LVH. In the present study, MCF was significantly reduced in CA and HCM groups for different reasons. Decreased MCF in patients with CA is a result of decrease in SV and increase in myocardial volume. In early stage HCM patients, SV remains normal or even larger, while myocardial volume increases in a larger scale, thus resulting a decreased MCF. However, MCF was not statistically superior to LVEF at differentiating CA from HCM in our study group.MCF was initially described by King et al., LVGFI represents the relationship between SV and total heart size, including myocardial volume and mean LV cavity volume (mean of end-diastolic and end-systolic volumes)11, which carries additional information about cardiac remodeling. As it was reported by previous studies, LVGFI could provide incremental prognostic value in the prediction of major adverse cardiac events in patients after ST-segment elevation myocardial infarction15. In the present study, LVGFI decreased in both CA and HCM groups. And LVGFI had strong correlation with myocardial fibrosis assessed by LGE. In the HCM group, LV cavity volume is reduced because of increased wall thickness, and to preserve SV, the end-diastolic volume is enlarged through remodeling, which is consistent with findings of the present study. EDV was not significantly increased in CA, probably because amyloid deposition in the myocardium has caused more pronounced diastolic dysfunction. Different pathophysiologic mechanisms that have led to alterations in LV function and structure are comprehensively expressed by LVGFI. And this might be the foremost reason that LVGFI could outperformed conventional functional parameter in differentiating CA from HCM. And we had also found that LVGFI had comparable diagnostic capacity to LGE, which had been widely used in clinical settings.The present study showed that LVGFI was superior to conventional LVEF in assessing and discriminating CA patients, and comparable to quantified LGE in the differential diagnosis. As described by Mewton In patients with similar hypertrophy, LVGFI continued to be an accurate parameter with high sensitivity and specificity. In this condition, myocardial volume of the two group had no significant difference. But amyloid deposition could cause a thickened and stiffened ventricular wall, restraining EDV to dilate, leading to a dramatically decreased LVGFI. On the contrary, early stage HCM could preserve cardiac output by enlarging EDV. So decreased LVGFI in HCM would not be as reduced as CA. In the presence of preserved LVEF, LVGFI still demonstrated excellent AUC with high sensitivity and moderate specificity. Differentiating between patients with similar hypertrophy and preserved LVEF are difficult issues that clinicians may frequently encounter. Subgroup analyses in the present study indicated that LVGFI can be powerful even in these challenging clinical settings.The formulas for calculating LVGFI and MCF are similar. They both account for the relationship between LV mass and LV dimensions. However, LVGFI accounts for alterations in LV mass and overall LV cavity size that reflect remodeling in the heart, which demonstrates intrinsic pathophysiologic characteristics of the amyloid deposits-hypertrophy. We also found that the superiority of LVGFI was not a result of a simple combination of basic LV geometric measurements, such as ESV, EDV and LV mass. When we use a linear combination of LVEF and LV mass to differentiate CA from HCM, the diagnostic value was the same as LVEF. And this is because there was no significant difference of LV mass between the two groups (p = 0.65).17. The present study also demonstrated that LVGFI and MCF are strongly associated with troponin T and NT-proBNP. The CA group had significant higher levels of troponin T, NT-proBNP, and NYHA than HCM in this study mainly due to the following reasons. For one thing, cardiac amyloidosis is a more aggressive disease with poor prognosis. Amyloid deposits can cause myocardial microvascular dysfunction, decrease in LV systolic function, and cardiac troponin release18. For another, circulating amyloid light chain seems to have a direct cardiotoxic effect, which could impair the cardiac function. Last but not least, the atrial chamber would be dilated as a result of the more pronounced diastolic dysfunction in CA patients, which could lead to a great release of natriuretic peptide.Troponin T level was markedly elevated in hypertrophic patients in this study, which is consistent with previous studiesThe retrospective nature of this study carries its major limitations. The lack of cardiac biopsy and genetic testing in these patients made it impossible to precisely differentiate the subtypes  of cardiac amyloidosis. In addition, sub-analysis of patients with preserved LVEF was limited by small sample size. Further well-designed prospective studies are needed to determine the utility of these two parameters for a more general application.LVGFI is a novel and clinically useful parameters with excellent ability in determining myocardial function and differentiating cardiac amyloidosis from hypertrophic cardiomyopathy.All the patients with increased ventricular wall thickness who underwent CMR between 2015 and 2018 at West China Hospital were retrospectively reviewed.19. The HCM group was included according to the 2014 ESC Guidelines on diagnosis and management of hypertrophic cardiomyopathy20.CA was diagnosed on the basis of myocardial biopsy, if performed. If not, cardiac involvement of amyloidosis was considered if mean LV wall thickness ≥12 mm on echocardiography (EEG) or CMR in conjunction with a positive extra-cardiac biopsy and conditions such as plasma cell dyscrasia, and multiple myelomaExclusion criteria included patients with valvular heart disease (greater than mild stenosis or greater than moderate regurgitation), significant coronary artery disease, and other confirmed systemic diseases. Hypertensive patients with concentric hypertrophy were also excluded.We also included a normal control group, which was confirmed by no evidence of structural or functional heart diseases on CMR, no clinical heart disease history and no electrocardiogram abnormalities. The New York Heart Association Classification (NYHA) was performed by the patient’s referring cardiologist. Baseline characteristics and laboratory biomarkers were assessed within one week of the CMR examinations. This study was approved by the Institutional Ethics Review Board of West China Hospital and in accordance with the Declaration of Helsinki. All participants signed written informed consent before the CMR scans.2; matrix size = 176 × 256).CMR scans were acquired using standard institutional CMR protocols with a 3.0 Tesla MR system . Cine images were obtained using balanced steady-state free-precession sequences in consecutive short axis covering the LV  and in the long axis . Scanning parameters were as follows: repetition time (TR), 39.34 ms and echo time (TE) 1.22 ms; flip angle, 38°; field of view, 284 × 399 mm; matrix size 139 × 208 mm; and slice thickness, 8 mm. 25 cardiac phases were retrospectively reconstructed for each slice level. LV volumes, and wall thickness were all assessed from these continuous cine sequences. Late gadolinium enhancement (LGE) imaging was acquired 10–15 min after contrast injection by using an inversion recovery TrueFISP sequence . Endocardial and epicardial contours were manually delineated by two experienced radiologists in consecutive short-axis slices at end-diastolic and end-systolic phases. The radiologists were blinded to the patients’ clinical profiles. Trabecular tissue and papillary muscles were excluded from LV mass. LGE extent was quantified by 5 standard deviations (SD) above the signal intensity of remote normal myocardium21.CMR analysis was conducted using a commercially available software .11. LV global volume was defined as the sum of the mean LV cavity volume [(LVEDV + LVESV)/2] and the myocardial volume. LV myocardial volume was calculated as LV myocardial mass divided by myocardial density, which is specified as 1.05 g/mL22.LVGFI was calculated according to equation (1) and expressed as a percentage12.MCF was calculated by dividing LVSV by LV myocardial volumeThe formula of left ventricular ejection fraction:P < 0.05 was considered statistically significant. Statistical analyses were conducted using SPSS  and GraphPad Prism 7 . Significance of the difference between the areas under ROC curves was verified using Stata .Data were expressed as mean ± standard deviation or median (interquartile range). Continuous variables were tested for normal distribution using Kolmogorov–Smirnov test. CMR measures were compared between CA and the control group using independent samples t-test or Mann–Whitney rank sum test, as appropriate. Comparisons between multiple groups were conducted using one-way analysis of variance or Kruskal–Wallis test with post hoc Bonferroni correction. Receiver operating characteristic (ROC) curve analysis was conducted to delineate the diagnostic accuracy of LVGFI and MCF. Areas under the curves (AUC) were statistically compared. NT-proBNP and Troponin T were log-transformed because of their skewed distribution. Correlations were assessed using Pearson’s correlation coefficient. To assess whether LVEF, LVGFI, MCF, LGE, log NT-proBNP, log Troponin T, and age had additional predictive values for the detection of amyloid myocardial infiltration, a forward and backward logistic regression with multiple covariates was performed using the diagnosis of amyloidosis as the outcome variable. Supplementary information