The ATTRv patient underwent heart transplantation because of progressive heart failure. Within the next two years, progressive myopathic symptoms and extracardiac tracer uptake on 99mTc-DPD planar scintigraphy were documented, attributable to ATTR amyloid myopathy. Interstitial amyloid deposits were confirmed by muscle biopsy in both patients, with a particularly high amyloid burden in the adipose tissue. This case report highlights the frequent concomitant presence of cardiac ATTR amyloidosis and ATTR amyloid myopathy. ATTR amyloid myopathy may precede cardiac manifestation in ATTRwt or occur after heart transplantation in ATTRv. Due to the high diagnostic accuracy of 99mTc-DPD scintigraphy for detecting ATTR amyloid myopathy and the emergence of novel therapeutics, it is important to increase the awareness of its presence.We identified two patients with transthyretin (ATTR) amyloid myopathy . Myopathy was the initial manifestation in ATTRwt, whereas it followed neuropathy and cardiomyopathy in ATTRv. The ATTRwt patient showed muscular tracer uptake on  This was particularly pronounced in ATTRwt and ATTR-Val122Ile and could be confirmed histologically as ATTR amyloid myopathy in a small subset of patients with Perugini grade 3 on 99mTc-DPD scans.7ATTR amyloid myopathy, which is characterized by intramuscular interstitial amyloid deposits leading to weakness in the upper and/or lower extremities may contribute significantly to morbidity.We herein describe two cases of ATTR amyloidosis (ATTRv and ATTRwt) with clinical, imaging, and histological evidence of ATTR amyloid myopathy, which is probably often underappreciated and underrecognized in routine clinical practice Table .Table 1C99mTc-DPD whole-body planar imaging . There was no family history of amyloidosis. The patient’s medical records were remarkable for bilateral carpal tunnel syndrome ten years and decompression for lumbar vertebral stenosis two years before diagnosis of ATTRv amyloidosis. Neurological examination at the time of diagnosis of ATTRv amyloidosis in 2015 revealed fatigue, paresthesia of the feet, and loss of Achilles tendon reflexes. Dyspnea, bilateral lower leg edema, muscle cramps, mild proximal muscle weakness, upper extremity paresthesia, and numbness in the upper and lower extremities developed over the following 1.5 years. In addition to heart failure treatment, the ATTR tetramer stabilizer tafamidis was started in 2017 when this substance was first available in Austria. Three years after the initial diagnosis, the patient underwent heart transplantation due to progressive heart failure. Tafamidis, which was paused perioperatively, was restarted seven weeks thereafter. On routine follow-up 12 and 19 months after successful heart transplantation, no evidence for recurrent cardiac amyloidosis was found on endomyocardial biopsy and 99mTc-DPD planar scintigraphy  showed minor abnormalities with a mixed pattern of mild neurogenic and myogenic changes.Serum creatine kinase (CK) levels were within the normal range.Figure Reticular deposits of homogeneous eosinophilic material, representing amyloid, were mainly observed in the perimysial adipose tissue. Interstitial deposition of amyloid was also found in endomysium of skeletal muscle. Interestingly, the ATTR amyloid load was greater in the surrounding adipose tissue than in skeletal muscle. Immunohistochemistry showed amyloid deposits immunoreactive for transthyretin Figure E-G.99mTc-DPD scintigraphy and cardiac magnetic resonance imaging. No pathogenic mutation could be detected and treatment with tafamidis was initiated in 2019. 99mTc-DPD whole-body planar imaging  after diagnosis of ATTRwt amyloidosis showing significant ATTR amyloid deposition  initially presented with bilateral lower leg edema, vertigo, numbness in the pretibial area, lower extremity muscle weakness and muscle cramps. The latter had already started 1-2 years before and was initially considered as side effect of statin use. In the absence of monoclonal proteins, diagnosis of cardiac ATTR amyloidosis was obtained by A low-grade sensorimotor axonal polyneuropathy of lower extremities was revealed in 2019. Myopathic changes in the left gluteal muscle could be detected one year later by EMG.CK levels were within the normal range.Figure Hematoxylin and eosin staining showed slight variation of fiber size and increase of endomysial connective tissue. Necrotic fibers were not seen, although few regenerating fibers were present. Single atrophic fibers were vacuolated and contained amorphous sarcoplasmic deposits suspicious of amyloid. These amorphous deposits were clearly identified as green birefringent amyloid deposits on Congo red staining and were immunoreactive for transthyretin Figure A-D.TTR gene have been described to date, characterized by a wide phenotypic heterogeneity.2 The ATTR-Val40Ile variant, which was diagnosed in patient 8 A similar course was seen in patient 99mTc-DPD planar scintigraphy 19 months after heart transplantation revealed pronounced muscular tracer uptake in the shoulder and gluteal regions in the absence of cardiac tracer uptake in the transplanted heart. At the same time the patient complained of progressive muscle weakness and muscle cramps. Biopsy from the deltoid muscle confirmed significant ATTR amyloid deposition. As there was no evidence of progression of the previously diagnosed neuropathy in the follow-up examinations, it seems likely that the patient's complaints were due to increased amyloid deposits in the peripheral muscles after heart transplantation.In ATTRv more than 140 mutations in the 9One possible explanation for the development and clinical presentation of amyloid myopathy could be that ATTRv amyloid binds to pre-existing ATTRv amyloid deposits with high affinity. Therefore, after heart transplantation and release of ATTR amyloid cardiomyopathy, ATTRv amyloid may preferably deposit on pre-existing, formerly subclinical, ATTRv amyloid deposits in skeletal muscle, causing posttransplant amyloid myopathy in our patient. Similarly, it was reported that amyloid cardiomyopathy may develop after liver transplantation in ATTRv patients, due to binding of wild-type transthyretins, produced by the transplanted liver, on pre-existing ATTRv amyloid deposits in the myocardium.7 In this study bone uptake on 99mTc-DPD scintigraphy increased over the 3 h imaging period, whereas cardiac uptake and soft-tissue uptake (mainly muscle) decreased over time. This seems to be contradictory to the Perugini grading system, which is based on the reciprocal and relative decrease in bone uptake on planar imaging. The authors postulated that bone uptake particularly in patients with Perugini grade 3 on 99mTc-DPD scans may be obscured by tracer uptake in overlying skeletal muscles. Extensive soft-tissue uptake may even obscure visualization of cardiac tracer uptake on planar imaging.7 Conversely, it is conceivable that extensive cardiac tracer uptake may obscure visualization of muscular tracer uptake. It must therefore be considered that in our patient amyloid myopathy was already present before heart transplantation but was not noticed due to this phenomenon. However, increasing muscular complaints suggest an increase in amyloid load after heart transplantation.Hutt et al reported that most patients with cardiac ATTR amyloidosis, especially ATTRwt and ATTR-Val122Ile variant showed muscular tracer uptake on bone scintigraphy.Despite clinical improvement of muscular complaints under gene silencing therapy, a scintigraphic follow-up examination in November 2021 (40 months after heart transplantation) still showed a marked increase in muscular tracer uptake in shoulder and gluteal regions. Whether prolonging the duration of therapy will ultimately result in regression of muscular amyloid deposits remains to be seen.6Patient 2 (ATTRwt) complained about lower extremity muscle weakness and muscle cramps, which already started 1-2 years before cardiac manifestation. This was primarily attributed to a side effect of statin therapy. However, it is more likely that these complaints were already due to the subsequently confirmed amyloid myopathy. This is in concurrence with previous reports indicating that amyloid myopathy may precede cardiac manifestation in ATTRwt.It must be noted that ATTR amyloid myopathy is often difficult to distinguish from neuropathy in routine clinical practice. This is particularly because symptoms attributed to amyloid myopathy, such as weakness and cramps, are similar to symptoms of neuropathy, CK levels are mostly within the normal range and changes in EMG are often not very pronounced. This is due to interstitial amyloid localisation, whereas muscle fiber necrosis and muscle fiber alterations, causing changes in CK levels and EMG, are rarely present. Consequently, ATTR amyloid myopathy is likely to be frequently overlooked or mistaken as neuropathy.99mTc-DPD scintigraphy may ultimately be the more important diagnostic screening tool for diagnosis of ATTR amyloid myopathy than biopsy in future, given the current literature.7Histologically remarkable in this case report is the pronounced amyloid deposition in the adipose tissue in both patients, especially in patient 99mTc-labeled-pyrophosphat (99mTc-PYP) was minimal when assessed by qualitative and quantitative metrics. Thus, they conclude that the properties of 99mTc-PYP may be different from 99mTc-DPD in terms of non-cardiac uptake and that 99mTc-PYP cannot be used to image extracardiac ATTR deposition.10In a previous retrospective analysis of 57 patients with cardiac ATTR amyloidosis, Sperry et al noted that skeletal muscle uptake of 99mTc-DPD scintigraphy with high accuracy. As already shown in previous studies, myopathy can precede cardiac manifestation in ATTRwt amyloidosis.6 To date, amyloid myopathy has not been reported in the ATTR-Val40Ile variant and must be recognized as an important contributor to morbidity in these patients. In concurrence with previous reports, our findings from patient 1 (ATTRv) show that neuropathy and cardiomyopathy may precede the initial manifestation of myopathy.6 Whether this also applies to other patients with the ATTR-Val40Ile variant will be subject of future studies.In summary, muscular ATTR deposits can be visualized by 99mTc-DPD scintigraphy enables non-invasive diagnosis of ATTR amyloid myopathy with high accuracy, while invasive muscular biopsy is no longer obligatory. Due to the broad availability of 99mTc-DPD scintigraphy and the emergence of novel therapeutics it is of utmost importance to increase the awareness for the frequent concomitant occurrence of ATTR amyloid cardiomyopathy and myopathy, expanding the clinical spectrum of ATTR amyloidosis.Conclusively, we think that ATTR amyloid myopathy is still underappreciated and underrecognized, because accurate diagnosis in the past seemed difficult to achieve in routine clinical practice. As shown in this case report,


The aim of the study was to determine whether left ventricular electrical potential measured by electromechanical mapping with the NOGA XP system has predictive value for response to CRT.Approximately 30% of patients who undergo cardiac resynchronization therapy do not see the expected effects.The group of 38 patients qualified for CRT implantation were included in the study, of which 33 patients were analyzed. A 15% reduction in ESV after 6 months of pacing was used as a criterion for a positive response to CRT. The mean value and sum of unipolar and bipolar potentials obtained by mapping with the NOGA XP system and their predictive value in relation to the effect of CRT were analyzed using a bulls-eye projection at three levels: 1) the global value of the left ventricular (LV) potentials, 2) the potentials of the individual LV walls and 3) the mean value of the potentials of the individual segments  of the individual LV walls.24 patients met the criterion of a positive response to CRT vs. 9 non-responders. At the global analysis stage, the independent predictors of favorable response to CRT were the sum of the unipolar potential and bipolar mean potential. In the analysis of individual left ventricular walls, the mean bipolar potential of the anterior and posterior wall and in the unipolar system, mean septal potential was found to be an independent predictor of favorable response to CRT. In the detailed segmental analysis, the independent predictors were the bipolar potential of the mid-posterior wall segment and the basal anterior wall segment.Measurement of bipolar and unipolar electrical potentials with the NOGA XP system is a valuable method for predicting a favorable response to CRT. It is estimated that about 2% of global population is affected by symptomatic heart failure, and this percentage increases with age. Symptoms of the disease appear in 10% of people over 70. The 12-month total mortality rate for heart failure among inpatients and stable/outpatients is 17% and 7%, respectively, and the 12-month incidence of hospitalization is 44% and 32%, respectively. The pathomechanism of heart failure is complex. ECG changes are observed in some patients with significantly reduced ejection fraction (EF < 35%) in the form of atrioventricular conduction abnormalities and widened QRS complex, about 35% of whom manifest left bundle branch block. Conduction disturbances underlie cardiac systolic dyssynchrony and cause hemodynamic consequences that lead to adverse remodeling. Cardiac resynchronization therapy (CRT) restores the normal heart contraction sequence, improves the depolarization pattern, has a beneficial effect on quality of life, reduces the severity of symptoms, morbidity and mortality rates. Cardiac function is based on a complex electro-mechanical coupling mechanism. Systolic function deterioration is not only a result of the impaired propagation of the electrical impulse in the heart, but also of the loss of active cardiomyocytes capable of automatic resting depolarization and generating electrical potential. This raises the question of whether the value of this potential can predict the effect of resynchronization. The present study aimed to determine whether left ventricular (LV) electrical potential, as examined by electromechanical mapping with the NOGA XP system, has predictive value in responding to resynchronization therapy.2.Between April 2014 and July 2017, the study enrolled 38 patients, including 9 women (24%) and 29 men (76%), aged 49 to 76 years (mean age of 65.6 ± SD 5.7), with sinus rhythm, left bundle branch block and ischemic cardiomyopathy, with ESC class I and II indications for resynchronization system implantation (CRTD). Exclusion criteria were defined as follows: acute coronary syndrome for less than 3 months prior to the study inclusion, coronary artery disease requiring revascularization, previously implanted pacemaker or cardioverter-defibrillator, aortic valve calcification or LV thrombus, chronic kidney disease, pregnancy or lactation, active malignancy, viral infection, bleeding diathesis, allergy to contrast agent, current participation in another study, life expectancy less than 6 months. The project was prospective, single cohort study compliant with the Declaration of Helsinki. It was approved by the Bioethics Committee of the Silesian Medical University in Katowice KNW/0022/KB1/17/15. All patients signed an informed consent form. The characteristics of the patient group is shown in Thirty-three patients were analyzed; one patient could not be mapped with the NOGA system, and four withdrew before the 6-month follow-up examination (6MFU) .All patients qualified for implantation of the resynchronization system underwent echocardiography using the Phillips iE 33 and Cx 50 devices. End-systolic volume (ESV), left ventricular ejection fraction (LVEF), end-diastolic volume (EDV), and the degree of mitral regurgitation were assessed. A minimum 15% reduction in ESV after 6 months of follow-up was used as a criterion for evaluating the response to resynchronization therapy as positive. A clinical evaluation was also conducted, including the NYHA cardiovascular fitness classification, 6MWT. Electromechanical mapping was performed once in each patient using the NOGA XP AEMM system . Left ventricular electrical potentials were recorded in unipolar and bipolar systems (273 ± 47 recorded and mapped points per patient). The analysis of the averaged potentials and their sum was performed in the LV bull-eye projection divided by basal segments , analogous intermediate segments  and in one apical segment. The analysis was performed for the entire left ventricle, individual walls  and individual basal and intermediate segments. The resynchronization system implantation procedure was performed in all patients. The right ventricular lead was located in the RV apex. Implantation of the left ventricular lead in all patients was performed in a standardised standard manner, with the selection of the target lateral or posterolateral vein avoiding the location in the apical region . Changes in echocardiographic and clinical parameters after 6 months concerning baseline values are shown in An example of bipolar (1) and unipolar (2) potential maps in a non-responder (A) and a responder (B) is shown in 5.The values of mean potentials and the sum of mean potentials measured in 9 segments in the responder and non-responder groups were compared, obtaining a significant difference between compared groups except for the mean value of the unipolar potential. The values of LV electrical potential in the responders and non-responders groups are shown in r = 0.45, p = 0.009), as well as their sum  and the percentage of change in the end-systolic volume after 6 months of resynchronization pacing relative to baseline. Such a relationship was also found for the mean values of potentials  and the sum of potentials measured in the 9 segments in the unipolar configuration . Logistic regression analysis and ROC analysis found clear predictive value of mean unipolar  and bipolar  and summed unipolar  and bipolar  LV electrical potentials of positive response to resynchronization therapy.Regression analysis showed a clear correlation between the mean values of the potentials measured in the 9 segments in the bipolar configuration (p = 0.001), as well as bipolar mean potential  could be an independent predictor of a positive response to the resynchronization therapy.Based on the results, the prediction models were developed for the individual parameters of LV electrical potentials in unipolar and bipolar configurations. The analysis revealed that the sum of recorded unipolar potentials—unipolar sum  and the septum . These parameters tended to be significant in the anterior  and lateral  walls. The mean potential in the unipolar configuration of the anterior wall , lateral wall , posterior wall  and septal wall  also show predictive value. A predictive model was developed for the mean potential values of each left ventricular wall in bipolar and unipolar configurations. Mean bipolar anterior and posterior wall potential, as well as mean septal potential in unipolar configuration, appeared to be independent predictors of favorable response to the resynchronization therapy, noted in In logistic regression analysis confirmed by C-statistic analysis, the predictive factors of positive response to resynchronization therapy include mean bipolar potentials of the posterior wall  and mean posterior wall potential  while for the unipolar system, the mean septal potential .7.In the next step, the relationships between the values of potentials in unipolar and bipolar configurations of individual basal and medial segments in cross sections of the anterior, lateral, posterior and septal walls were analyzed .Differences between the values of potentials in responders (R) and non-responders (nR) in individual transverse segments in the study group were observed only during measurements in the bipolar configuration. The exceptions were the middle segment of the anterior wall and the middle segment of the lateral wall, in which no difference was observed .p = 0.010), as well as the mean value of the bipolar potential of all middle segments . In the analysis of individual segments, the bipolar potentials of the mid-segment of the posterior wall  and the basal segment of this wall show a predictive relationship with respect to a favorable response to CRT , as well as the basal segment of the anterior wall  and lateral wall . The bipolar signal of the medial septal segment also shows predictive value in this regard , while the bipolar signal of the inferior part of the septum tends to be significant in this regard . No predictive value was found in response to resynchronization therapy for the anterior and lateral medial segments.In the regression analysis confirmed by the results of the C-statistic, the predictive factors of a positive response to the resynchronization therapy in the analysis of transverse segment potentials include the mean value of the bipolar potential of all basal segments  in global, individual left ventricular wall and individual segment analysis has predictive value for a positive response to resynchronization therapy.The mean bipolar potential and the sum of mean bipolar and unipolar potential are the predictive factors for a positive resynchronization response. The mean bipolar and the sum of unipolar potential obtained during the electromechanical mapping of the AEMM were the independent predictors of a positive response to resynchronization therapy.In the analysis of the bipolar electrical potential of individual heart walls, the mean potential of the anterior and posterior walls was an independent predictor of favorable response to resynchronization therapy. In the unipolar signal analysis of individual walls, the mean interventricular septal potential was an independent predictor of favorable response to CRT.In a detailed segmental cross-sectional analysis, independent predictive value for favorable response to the resynchronization therapy was shown by the bipolar potential of the basal part of the anterior wall and the medial part of the posterior wall.9.Although cardiac resynchronization therapy (CRT) is an established and effective treatment for systolic heart failure , it doesIn recent years, in addition to standard resynchronization therapy, the frequency of using various techniques of pacing the conduction system has been increasing. Pacing of the His bundle has been reported in many studies to have similar electroradiographic and hemodynamic effects compared to traditional biventricular pacing , 5, althEach of the methods mentioned above has its limitations and new pacing modalities have yet to be evaluated in randomized clinical trials. Therefore, the reasons for the lack of response to conventional resynchronization therapy are constantly being sought and efforts are being made define predictive parameters to identify those patients in whom we can expect a beneficial effect of biventricular pacing. One factor undoubtedly limiting the effectiveness of resynchronization is atrial fibrillation, which reduces cardiac output through loss of the hemodynamic effect of atrial contraction on the one hand, and a reduction in the percentage of biventricular pacing on the other hand. It is worth noting that seven patients (2.94%) in the study group had a history of atrial fibrillation. According to the inclusion criterion of sinus rhythm in the study group, atrial fibrillation in these patients was paroxysmal and did not constitute a clinical problem during the 6-month follow-up. Therefore, the study group did not consider advanced treatment strategies using node ablation, which are effective in the case of permanent form of this arrhythmia .In our study, we focused on electrophysiological data obtained during electro-mechanical mapping (AEMM).The potential recorded in the AEMM is a direct consequence of the electrophysiological properties of the myocardial tissue. It has not yet been used in predicting a response to resynchronization therapy. Electro-mechanical coupling underlies the heart's systolic function. The ability to depolarize and induce effective contraction is a property of all individual cardiomyocytes joined into structures that form a three-dimensional the myocardium model . The synThe myocardial viability assessment is widely used in diagnosing primary and secondary heart diseases and evaluating the adverse cardiotoxic effects of certain agents and exogenous substances, including drugs used in oncology and others. Viability assessments include methods using ultrasound, multi-row computed tomography, single photon emission tomography (SPECT), positron emission tomography (PET), and magnetic resonance imaging (MRI), and have been the subject of many studies –17. VitaIn the analysis of our patient group, the predictive value of a favorable response to resynchronization therapy was demonstrated by the mean unipolar and bipolar potentials, as well as the sum of the mean unipolar and bipolar potentials of the individual, analyzed segments. In our study, we attempted to assess the predictive value of unipolar and bipolar electrical potentials demonstrated by mean unipolar and bipolar potentials, as well as the sum of mean unipolar and bipolar potentials of the individual, analyzed segments and their configurations in relation to a favourable response to resynchronization therapy. Considering the mechanics of left ventricular contraction under physiological conditions , 23, theThese parameters reflect global systolic potential and global left ventricular myocardial viability.When analyzing the values of potentials for individual walls analyzed in the long axis of the heart, the value of the mean bipolar potential of the anterior and posterior walls demonstrated the predictive value of a favorable response to resynchronization therapy. In a detailed segmental cross-sectional analysis, the independent predictive value for favorable response to the resynchronization therapy was shown by the bipolar potential of the basal part of the anterior wall and the medial part of the posterior wall. Higher mean values and total potentials are associated with a smaller mass of scar tissue within the left ventricle, which is not able to generate electrical potentials. Scar tissue does not determine the effect of cardiac resynchronization therapy (CRT) solely by the size of its mass. According to the results of the target study , the locIn previous studies, the values of unipolar and bipolar potentials characterized the healthy tissue, the scar area, and the transition zone differ in their values. According to Pavo et al. , the potA meta-analysis of viability assessment based on the measurement of potentials recorded during electromechanical mapping against various reference methods was presented by Gyongyosi . AssessmIn the study group, we found that a mean unipolar potential higher than 8.2 mV and a bipolar potential higher than 2.15 mV were predictors of a favorable response to resynchronization therapy. The electrical potential that produces a mechanical effect in the form of contraction and the perfusion of the analyzed area are two most important factors that guarantee the normal function of the myocardium. Identifying these mechanisms, their disorders, as well as their reserves seems crucial for identifying potential responders to resynchronization therapy. Nevertheless, it seems that further research using various methods to study these phenomena, including the evaluation of generated electrical potentials in the key areas of the myocardium, will help increase the percentage of patients benefiting from resynchronization therapy.10.The value of this study was the use of the highly accurate NOGA XP electromechanical mapping system, innovative in the evaluation of CRT response. It should be noted that electromechanical mapping was performed exclusively for this project. The main limitation of the study is the relatively small group of patients studied. However, due to the invasive nature of the mapping, recruitment to the study was halted when statistically significant correlations occurred.Currently, new pharmacological strategies are being used in the treatment of chronic heart failure, i.e., phlazines  and sacu


P < .05), except for the ADC CST-ratio (P > .05). Regarding the DTT parameters of the CRT-ratio, the patient group FN value was significantly lower than that of the control group (P < .05), whereas the FA and ADC CRT-ratios did not show significant differences between the patient and control groups (P > .05). MAS scores showed a strong positive correlation with the ADC CRT-ratio (P < .05) and a moderate negative correlation with the FN CRT-ratio (P < .05). We observed that the injury severities of the CST and CRT were related to spasticity severity in chronic stroke patients; moreover, compared to the CST, CRT status was more closely related to spasticity severity.We investigated the correlation between spasticity and the states of the corticospinal tract (CST) and corticoreticular tract (CRT) in stroke patients after early stage. Thirty-eight stroke patients and 26 healthy control subjects were recruited. The modified Ashworth scale (MAS) scale after the early stage (more than 1 month after onset) was used to determine the spasticity state of the stroke patients. Fractional anisotropy (FA), apparent diffusion coefficient (ADC), fiber number (FN), and ipsilesional/contra-lesional ratios for diffusion tensor tractography (DTT) parameters of the CST and CRT after the early stage were measured in both ipsi- and contra-lesional hemispheres. This study was conducted retrospectively. The FA and FN CST-ratios in the patient group were significantly lower than those of the control group ( Among the above-mentioned neural tracts, previous studies have reported that the VST has a minor role in spasticity.–13 In this study, we hypothesized that spasticity in stroke patients could be related to CST and CRT injuries.The introduction of diffusion tensor tractography (DTT), which is derived from diffusion tensor imaging (DTI), has enabled 3-dimensional reconstruction and estimation of various neural tracts, including the CST, CRT, and VST.In the current study, by using DTT, we investigated the correlation between spasticity and the state of the CST and CRT in stroke patients after early stage.,33); no history of neurologic/psychiatric disease or head trauma. MAS scores, used to determine the spasticity state of the stroke patients, were obtained at the time of DTI scanning.,33 This study was conducted retrospectively, and all patients and control subjects provided written informed consent. The study protocol was approved by the institutional review board of a university hospital (IRB number: YUMC 2021-03-014).In this study, the stroke patients were recruited according to the following inclusion criteria: first-ever stroke; age at the onset of stroke: 20 to 69 years; spontaneous intracerebral hemorrhage or cerebral infarction confined to a unilateral hemisphere, as confirmed by a neuroradiologist; DTI scan performed after the early stage (more than 1 month after onset); spasticity in the contra-lesional extremity . Each DTI replication was intra-registered to baseline “b0” images to correct for residual eddy-current image distortions and head motion effects by using a diffusion registration package . The CST was reconstructed using fibers passing through 2 regions of interest (ROIs) on the DTI color map. The seed ROI was placed at the upper pons, and the target ROI was placed at the mid pons. For analysis of the CRT, the seed ROI was placed on reticular formation of medulla, and the target ROI was placed on the midbrain tegmentum. Termination criteria used for fiber tracking were fractional anisotropy (FA) < 0.15 and angle < 27°. The FA, apparent diffusion coefficient (ADC), and fiber number (FN) values for the CST and CRT after the early stage were measured in both the ipsilesional and contra-lesional hemispheres. Subsequently, ipsilesional/contra-lesional ratios of the CST and CRT for each of the DTT parameters  after the early stage were calculated and are presented as CST- and CRT-ratios, respectively  equipped with a Synergy-L Sensitivity Encoding (SENSE) head coil and using a single-shot, spin-echo planar imaging pulse sequence. For each of 32 non-collinear diffusion sensitizing gradients, 60 contiguous slices were acquired parallel to the anterior commissure-posterior commissure line. Imaging parameters were as follows: acquisition matrix = 96 × 96, reconstructed to matrix = 192 × 192, field of view = 240 mm × 240 mm, TR = 10,398 ms, TE = 72 ms, parallel imaging reduction factor (SENSE factor) = 2, EPI factor = 59 and b = 1000 s/mmt tests were used to examine the age distribution difference and the DTT parameter differences in CST- and CRT-ratios between the patient and control groups. The level of statistical significance was set at P < .05. The MAS scale category 1 + was modified to a score of 1.5 for statistical analysis purposes. Spearman rank correlation coefficients were used to examine correlations between MAS score and CST- and CRT-ratios for each of the DTT parameters. A correlation coefficient  was interpreted as strong when >0.50, as moderate when between 0.30 and 0.49, and weak when between 0.10 and 0.29.SPSS software  was used for data analysis. The chi-squared test was used to assess a sex-based difference between the patient and control groups. Independent P > .05).Thirty-eight stroke patients  and 26 age- and sex-matched healthy control subjects  with no history of neurologic/psychiatric disease or head trauma were recruited. Demographic data for the patient and control groups are summarized in Table P < .05). However, there was no significant difference between the 2 groups in the ADC CST-ratio (P > .05). Regarding the CRT, the FN CRT-ratio of the patient group was significantly lower than that of the control group (P < .05), whereas the FA and ADC CRT-ratios were not significantly different between the patient and control groups (P > .05).The CST- and CRT-ratios for each DTT parameter in the patient and control groups are summarized in Table R = 0.574, P < .05) and a moderate negative correlation with the FN CRT-ratio .Correlations between MAS score and the CST- and CRT-ratios for the DTT parameters in the patient group are summarized in Table In the present study, by examining the correlation of MAS score with DTT parameter ratios for the CST and CRT, we investigated the correlation between spasticity and the CST and CRT states in stroke patients after early stage. Our results are summarized as follows. First, our CST-ratio examination showed that the patient group had lower FA and FN values than those of the control group. Second, the CRT-ratio assessment showed that the FN value in the patient group was lower than that in the control group. Third, the MAS score had a strong positive correlation with the ADC CRT-ratio and a moderate negative correlation with the FN CRT-ratio.,36,37 The FA parameter, which indicates the degree of directionality of water diffusion, is used to assess the degree of tract directionality, whereas the ADC parameter indicates the magnitude of the water diffusion.,36,37 Therefore, the FA and ADC parameters may be used to indicate the microstructural integrity status of white matter microstructures, such as axons, myelin, and microtubules.,36,37 In contrast, the FN value indicates the number of voxels in a neural tract, suggestive of the total number of fibers in a neural tract. Therefore, decrements in FA and FN values and an increment in the ADC value of a neural tract indicate that neural tract injury status.,36,37 In addition, DTT parameters presented as ipsi-/contra-lesional ratios of a neural tract reflect the degree of asymmetry between the ipsi- and contra-lesional tracts. Therefore, relatively large decrements in FA and FN values and increment in ADC value in bilateral ratios for a neural tract indicate greater injury in the ipsilesional neural tract than in the contra-lesional tract. Consequently, greater decrements in FA and/or FN CST- and CRT-ratios in the patient group than in the control group indicate the stroke patients exhibit more severe injuries in the ipsilesional CST and CRT than in the contra-lesional tracts.Among the various DTT parameters that can be examined, FA, ADC, and FN values are most commonly used when evaluating the state of neural tracts in patients with brain injury.The relationship between MAS scores and DTT parameter CST- and CRT-ratios in the patient group showed that the DTT parameter CST- ratios had no correlation with the MAS score, whereas, the ADC CRT-ratio was strongly positively correlated with the MAS score, and the FN CRT-ratio was moderately negatively correlated with the MAS score. Compared with the contra-lesional CRT, injury severity of the ipsilesional CRT was significantly related to the severity of spasticity in stroke patients after early stage. Specifically, compared with the ipsilesional CST, DTT results for the ipsilesional CRT were closely associated with spasticity severity.–13,15 In detail, spasticity is maintained by controlled balancing of the stretch reflex via inhibitory influences of the CST and the CRT with the lateral RST and the excitatory influences of the medial RST and VST; however, the VST is reported to have a minor effect on spasticity.–13,15 Therefore, in brain injuries that include the CST and CRT, the inhibitory influences on the medullary brainstem and intraspinal network can be lost, leading to an unopposed excitatory influence by the medial RST; thus, hyperexcitability of the medial RST can occur.–13,15 Hyperexcitability of the RST has been related to spasticity, abnormal motor synergy, and disordered motor control.,18 In addition, previous studies have reported that if a CST injury due to extensive cortical damage occurs, injury recovery can use an alternative motor pathway, such as the RST, for compensation, leading to abnormal motor synergy, dexterity loss, and spasticity.,38–41 Therefore, although various neural tracts are involved in spasticity, given the above results and the difficulty in identifying the individual actions of brainstem nuclei when using DTT, we focused on CST and CRT injuries in stroke patients after early stage with spasticity. Consequently, based on previous and current studies, we suggest that abnormal supraspinal influences on the intraspinal network due to the CRT injury rather than the CST injury could be responsible for spasticity in stroke patients after early stage.Among various pathophysiologic mechanisms of spasticity, abnormal supraspinal factors due to injury of neural tracts are reported to be major causes of spasticity.–44 In 2016, Barlow demonstrated injuries of gray  and white  matters using lesion density plots and voxel-based lesion-symptom mapping in 20 acute patients with spasticity following ischemic stroke. During the same year, Lee et al, using brain magnetic resonance imaging and positron emission tomography, reported on a patient who showed spasticity in the left leg due to injuries of the CST and CRT following infarction in the right cerebral peduncle. In 2019, Plantin et al demonstrated CST injury due to intracerebral hemorrhage or cerebral infarction by examining weighted CST lesion loads and voxel-based lesion-symptom mapping in 61 stroke patients with hand spasticity evaluated by the NeuroFlexor method, which accounts for velocity dependence in the tonic stretch reflex. Taken together, our results support previous studies showing that injuries of the CST and/or CRT are associated with spasticity. To the best of our knowledge, this is the first DTI-based study to report a correlation between spasticity and CST and CRT injuries.A few studies have reported on the correlation between spasticity and CST and/or CRT injuries in stroke patients.Several limitations of this study should be considered. First, the fiber tracking technique applied during DTT reconstruction is operator-dependent. Second, this study recruited a relatively small number of subjects. Third, among the descending motor pathways related to spasticity, we only reconstructed the CST and CRT because the descending motor pathways originating from the brainstem are challenging to reconstruct using DTI. Fourth, only MAS scale was used to evaluate spasticity in this study.–47 However, the reliability and validity of Ashworth scale methods for measuring the severity of muscle spasticity have been controversial.,48 Therefore, further prospective studies that include larger numbers of patients, more advanced spasticity evaluation tools, and enhanced DTI techniques to reconstruct motor pathways originating from brainstem should be encouraged.By contrast, the Modified MAS (MMAS) scale which is an advanced version of MAS that was reported as a more reliable and valid method than the MAS scale by excluding the MAS score category 1 + aspect, could be better for this kind of study.In conclusion, we have demonstrated that CST and CRT injury severities were related to spasticity severity in stroke patients after early stage. In particular, of the 2 tracts assessed, CRT injury status was the most closely related to spasticity severity. Our results suggest that recovery of CST and CRT injuries through neuro-rehabilitation during the stroke recovery phase is important in the prevention of spasticity after the early stage. Thus, DTT-based reconstruction and assessment of the CST and CRT could be helpful when predicting the occurrence of spasticity and planning neuro-rehabilitation treatments that could improve recovery prognosis.This work was supported by the National Research Foundation of Korea (NRF) grant funded by the Korean Government (MSIP) (No. 2021R1A2B5B01001386).MJC and SSY contributed equally to this work.Conceptualization: Min Jye Cho, Sung Jun Lee.Data curation: Min Jye Cho, Sang Seok Yeo, Sung Jun Lee, Sung Ho Jang.Formal analysis: Sung Jun Lee.Investigation: Min Jye Cho, Sang Seok Yeo, Sung Jun Lee.Methodology: Sang Seok Yeo, Sung Ho Jang.Supervision: Sang Seok Yeo, Sung Ho Jang.Visualization: Sung Jun Lee.Writing – original draft: Min Jye Cho, Sung Jun Lee.Writing – review & editing: Sang Seok Yeo, Sung Ho Jang.


Lipedema is a painful subcutaneous adipose tissue (SAT) disease characterized by adipocyte hypertrophy, immune cell recruitment, and fibrosis in the affected areas. These features are thought to contribute to the development and progression of the condition. However, the relationship between lipedema disease stage and the associated adipose tissue changes has not been determined so far.SAT biopsies of 32 lipedema patients, ranging across the pathological stages I to III, and 14 BMI- and age-matched controls were harvested from lipedema-affected thighs and non-symptomatic lower abdominal regions. Histological and immunohistochemical (IHC) staining and expression analysis of markers for adipogenesis, immunomodulation, and fibrosis were performed on the tissue biopsies.Lipedema patients showed increased adipocyte areas and a stage-dependent shift towards larger cell sizes in the thighs. Lipedema SAT was linked with increased interstitial collagen accumulation in the thighs, but not the lower abdominal region when compared to controls. There was a trend toward progressive SAT fibrosis of the affected thighs with increasing lipedema stage. Elevated gene expression levels of macrophage markers were found for thigh SAT biopsies, but not in the abdominal region. IHC staining of lipedema thigh biopsies confirmed a transiently elevated macrophage polarization towards an M2-like (anti-inflammatory) phenotype.In summary, lipedema SAT is associated with stage-dependent adipocyte hypertrophy, stage-progressive interstitial fibrosis and elevated proportion of M2-like macrophages. The character of the inflammatory response differs from primary obesity and may possess an essential role in the development of lipedema. Based on the limited data available, epidemiological estimates indicate an approximate prevalence of 10% in the overall female population. The disease is characterized by a bilaterally symmetrical and disproportional expansion of the subcutaneous adipose tissue (SAT) of the extremities in relation to the torso –4. SurgiRecent studies have highlighted characteristic abnormalities for lipedema-affected adipose tissue, including dysfunctional blood and lymphatic vessels (microangiopathy) , excess Current literature considers lipedema-associated symptoms as largely limited to the extremities. Nevertheless, the SAT of the lower abdomen is frequently claimed to be symptomatic in lipedema . The inv22.1The study protocol was approved by the local ethics committee prior to patient enrollment (Nr. S 6(a)/2020, accepted on 18. February 2020) and the study was conducted according to the principles of the Declaration of Helsinki and its amendments. All subjects provided their informed consent before study participation. Lipedema was diagnosed according to the criteria described in our previous work . A total2.2Adipose tissue biopsies were obtained from the thigh and the lower abdomen regions of female lipedema patients and age- and BMI-matched control subjects during elective liposuction. Tissue samples of the thigh were harvested in the ventral portion of the dominant affected (or right) thigh midway between the anterior superior iliac spine and the cranial margin of the patella. Samples from the caudal abdomen were taken originating from the umbilical approach. The biopsies were harvested before infiltration of the tumescent solution by surgical dissection through the regular surgical incisions for liposuction. One-half of the freshly obtained biopsy samples was stored in phosphate-buffered saline (PBS) with 3.5% BSA  at 4°C immediately after collection and during transport. Subsequently, the specimens were fixed in 4% formaldehyde/PBS before embedding for histology. The remaining biopsy material was immediately transferred into dry 1.5 ml collection tubes and stored without addition of cryopreservant on dry ice. For long-term storage, frozen biopsy specimens and lipoaspirates were transferred to -80°C.2.32] was quantified using Image J software  with the freely available plugins “MRI fibrosis tool” and “Color Deconvolution” , M1-like macrophages , and M2-like macrophages  at 4°C overnight. Secondary antibodies were either Anti-rabbit  or Anti-mouse . 4’,6-Diamidino-2-Phenylindole, Dihydrochloride  was used for nuclei staining. Images were captured at 200x, and labelled cells were counted manually using BZ-II Viewer software .2.4RNA from 30mg of minced and frozen adipose tissue samples Table 2C2.5For the assessment of the metabolic risk profile, blood plasma samples were collected preoperatively after 6-8 h of starvation in an S-monovette . Blood samples were stored on ice until further processing, subsequently centrifuged for 10 min at 6,000 rpm at 4°C, after which plasma was collected and stored at -80°C until analysis. Plasma profiles of 14 controls were analyzed and compared to 8 stage I, 16 stage II and 7 stage III classified lipedema patients Table 2E2.6Analyses were performed using GraphPad Prism, version 8.0.0 for Windows . All data were presented as mean ± standard error of the mean (SEM), or median . Sample sizes as well as the statistical analyses are indicated in the respective figure legends. Welch corrected t-test and multiple t-tests were applied for comparisons between two groups. Statistically significant differences are assumed for p-values < 0.05.33.1After verification of previously published diagnostic criteria  and evalAdipose tissue morphology was assessed in thigh and abdominal biopsies for all patients included in this study  and compared to morphologies of the 14 non-lipedema control subjects Table 2BThe stage dependent analysis revealed no significant differences between lipedema and controls in both biopsies comparing stage I patients to the control group . We therefore conducted a histological analysis of fibrosis levels in controls and lipedema patients. No difference of pericellular fibrosis was detected in the non-symptomatic lower abdominal region when comparing the control group to all lipedema patients while a significant increase of fibrotic areas was detected in the lipedema-affected thigh compared to healthy controls  was assessed as a general marker of mitochondrial function. In abdominal and thigh biopsies comparing control subjects to all lipedema patients, several marker genes of inflammation and angiogenesis were broadly and significantly upregulated, suggesting that this process is a target of lipedema pathology  and fibrosis marker collagen 6a1 (COL6A1) were elevated in abdominal biopsies of stage I patients , and increased expression of angiogenesis marker gene vascular endothelial growth factor C (VEGFC) IL6, tumor necrosis factor-alpha (TNF), and macrophage markers CD86 and mannose receptor c-type 1   macrophages Table 2D3.5As lipedema-associated changes in our cohort, such as adipocyte hypertrophy, interstitial fibrosis, and inflammation, are commonly associated with an altered metabolic profile in patients with primary obesity, we analyzed plasma samples to assess glucose and lipid metabolism in patients and controls Table 3.4The progression of lipedema has been linked to three pathophysiological processes, (1) morphological changes to adipocytes in the affected adipose depots, (2) accumulation of fibrotic areas within adipose depots, and (3) inflammatory processes, which are characterized by altered immune cell distribution and cytokine release patterns. A major challenge to adequate medical care for patients with lipedema is a robust and early diagnosis of the disease. In the present study, we have therefore combined diagnostic staging of disease severity, which requires specialist expertise in the attending physician, with a morphological assessment of adipose tissue and molecular analysis of fibrosis and inflammation. We show that all pathological changes occur stage-dependently and that adipocyte hypertrophy only occurs as a marker of later and more severe disease stages. It is preceded by characteristic changes in tissue fibrosis, inflammatory marker gene expression, and macrophage accumulation. Moreover, our data also show that patient-reported discomfort, which is mainly localized to SAT of the extremities, can be supported by objective measurement readouts. However, it does not serve as a certain predictor of molecular changes, which also occur in SAT of the abdominal region without a clear association to lipedema pathology, albeit with a milder phenotype. In summary, our analysis establishes that biomarkers of disease could help improve the initial diagnosis.Adipocyte hypertrophy and heterogeneity in cell size in lipedema thighs was previously reported by several groups –11, 15. Similar to fat mass expansion in obesity, our study demonstrated increased amounts of collagen as a sign of pericellular fibrosis surrounding adipocytes in lipedema . In contGiven that fibrosis was significantly elevated already in stage I patients, i.e. prior to onset of adipocyte hypertrophy, we believe such alterations could serve as markers for lipedema but might also be linked to the subsequently occurring changes in adipocyte size. While a general shift in adipocyte distribution toward larger cells was observed in the lipedema thigh regions in stage II and III, the mean adipocyte size did not vary between the stages. Some reports suggest that excessive extracellular matrix remodeling in the form of fibrosis inversely correlates with decreased adipocyte size, meaning that increased adipose tissue fibrosis restrains adipocyte hypertrophy , 25–27. CD163, a marker of M2-like macrophages, in SAT biopsies of lipedema patients (TNF and IL6. Unlike in the regions affected by lipedema, these increases were milder and appeared to not be affected by disease stage. In primary obesity, increased expression of genes related to inflammation were also detected in both the abdominal and gluteofemoral SAT, but in contrast to lipedema, these depot-specific associations were significantly weaker in the gluteofemoral SAT (It is generally accepted that modulation of inflammation may play a key role in the pathogenesis of lipedema , 13, 28.oral SAT . It remaLastly, an altered lipid metabolism is closely associated to adipose tissue inflammatory response in states of obesity , 46. SpeObesity-related adipocyte hypertrophy was demonstrated to trigger low-grade inflammation and excessive collagen deposition that would eventually lead to abnormal glucose homeostasis and systemic insulin resistance . In lipeen bloc from both regions. Unlike in punching procedures, the surgical dissection ensures that sufficient amount of tissue is obtained from both compartments. This en bloc harvesting technique provides comparability between all samples and individuals, regardless of local tissue variances, but does not allow differentiation between the two layers.Our present study has some limitations that should be considered. In view of the high inter-individual variability of human SAT samples, the relatively small total number of cases and control subjects requires careful interpretation of the results. Moreover, without an objective biomarker, the diagnosis of lipedema remains subject to a certain bias and relies on the presence of extensive expertise in the diagnosis of the disease. In this study, independent diagnosis validation and the patient’s interest in surgical symptom reduction with liposuction reduced this factor to a large extent. Another limitation might be the location of adipose tissue biopsy harvesting: The human SAT consists of two distinct compartments separated by Scarpa’s fascia into superficial and deep SAT. It was recently demonstrated that these depots are functionally and morphologically different in terms of adipocyte size and inflammatory gene expression levels . Since i5Lipedema SAT is associated with progressive interstitial fibrosis, inflammatory processes and an elevated proportion of M2-like macrophages in early stages of the disease that is followed by adipocyte hypertrophy and a more pro-inflammatory profile in later stages. The observed SAT changes are mainly limited to the clinically affected areas and provide objective measure to assist diagnosis, but are also present in other adipose regions to a more limited extent. The characteristics of the inflammatory response, and in particular the increased presence of anti-inflammatory, M2-like macrophages clearly distinguishes lipedema from primary obesity and may reflect a key role in the pathophysiology and symptoms of lipedema at different stages. Thus, the regulation of M2-like macrophages warrants further studies in relation to the clinical characterization of patients and development of diagnostic tools of the disease. Although altered lipid metabolism is closely related to the inflammatory response of adipose tissue in obesity, key predictors of metabolic risk were not altered in the lipedema cohort studied. In contrast, some parameters of metabolic health were improved in lipedema patients compared to BMI- and age-matched individuals. We conclude that a better understanding of the distinct composition of plasma lipids in conjunction with an assessment of inflammatory processes may help to better characterize the underlying patho-mechanisms of lipedema.The original contributions presented in the study are included in the article/Conceptualization, TS, MG, PK, SG and FG-C. Methodology, TS, MG, PK, SG, GS and FG-C. Formal analysis, PK, SG, KL, and FG-C. Investigation, PK, SG, KL, and FG-C. Resources, TS and MG. Data curation, PK, SG, KL, and FG-C. Writing—original draft preparation, PK and SG. Writing—review and editing, TS, MG, SG and FG-C. Visualization, PK and SG. Supervision, TS and MG. Project administration, TS and MG. All authors have read and agreed to the published version of the manuscript.


Lack of significant advancements in early detection and treatment of heart failure have precipitated the need for discovery of novel biomarkers and therapeutic targets. Over the past decade, circulating sphingolipids have elicited promising results as biomarkers that premonish adverse cardiac events. Additionally, compelling evidence directly ties sphingolipids to these events in patients with incident heart failure. This review aims to summarize the current literature on circulating sphingolipids in both human cohorts and animal models of heart failure. The goal is to provide direction and focus for future mechanistic studies in heart failure, as well as pave the way for the development of new sphingolipid biomarkers. Heart failure (HF) is a debilitating medical condition where the heart is unable to meet the metabolic demands of the body. It is a widespread clinical syndrome affecting at least 64 million people globally, and this number is expected to rise , 2. HF pAdvances in medicine over the past decade have improved the quality of life and outcomes for patients with HFrEF. These include pharmacotherapies such as Aldosterone/Mineralocorticoid Receptor Antagonists , Angiotensin Receptor-Neprilysin Inhibitor , ventricular assist devices (VAD), cardiac resynchronization therapy (CRT), and implantable cardioverter-defibrillator (ICDs) , 7–10. Hde novo in the endoplasmic reticulum via condensation of an amino acid, typically serine, and a fatty acid, catalyzed by serine palmitoyltransferase (SPT)  . This pase (SPT) . A fourtse (SPT) . Differese (SPT) . This gese (SPT) –26. The se (SPT) –30. For se (SPT) .de novo pathway, S1P lyase (S1PL) catabolizes S1P or DHS1P in lysosomes and plasma membranes to yield a fatty aldehyde and phosphoethanolamine , which is then N-acetylated by one of 6 isoforms of (dihydro) ceramide synthase (CerS), with different chain length fatty acids to synthesize dihydroceramides (DHC) . CerS1 tnolamine . The heanolamine . Studiesnolamine . The dognolamine –45. Thernolamine . As suchIn this review, we examine the relationship between sphingolipids and HF in both human study cohorts and animal models. We analyze the current literature to identify potential biomarkers and druggable targets for the detection and treatment of HF.post hoc analyses. This involves mass spectrometry to identify and quantify sphingolipids in blood serum, blood plasma, or myocardial tissue. This method yields measurements for numerous sphingolipids, which can then be correlated to disease and other clinical parameters to gain a better understanding of their relationship. Current literature on sphingolipid associations with HF can broadly be divided into two types: sphingolipid associations in patients with incident HF , have been ongoing for decades and have analyzed traditional risk factors like obesity, smoking status, and LDL-cholesterol from these patient cohorts. However, only in recent years have investigators started to address circulating sphingolipids in subjects from these studies through total Cer in HFrEF and HFpEF patients , the enzyme converting S1P to Sph, and S1PR3 were downregulated in HF patients. Though this should result in decreased S1P in myocardial tissue and not the observed increase in levels  cohort, identified higher levels of SM C16:0, and Cer C16:0 associated with higher risk and SM C20:0, C22:0, C24:0, and Cer C22:0 with lower risk of developing HF, independently. Interestingly, these species had similar associations regardless of whether the patient had HFpEF or HFrEF  studies. These assays were able to show a higher ratio in plasma Cer(C24:0/C16:0) was inversely associated with incident HF . Outcometotal Cer >6.05 ng/ml, and HF patients that survived (at least up to the 4 years during the study) had circulating total Cer <6.05 ng/ml  undergoing placement of VAD , Ventricle Assist Device (VAD) placement or replacement, death, or heart failure admission (DHFA), and HF-related mortality are described in this section. A 2015 study by Yu et al., concluded that plasma from patients with HF who died had 05 ng/ml . There ht of VAD . Interesting Cer . A studyting Cer . In a stting Cer . Though ting Cer .N = 12) , obesity or incident coronary heart disease (CHD), including but not limited to myocardial infarction (MI), atrial fibrillation, coronary insufficiency, adverse cardiac remodeling, and angina pectoris. The SHS (Strong Heart Study) and SHFS (Strong Heart Family Study) fare population-based longitudinal studies addressed cardiovascular disease (CVD) in several American Indian communities in Arizona, North and South Dakota, and Oklahoma. It is interesting to note that increased Cer C16:0, though not significant, correlated with increased CVD risk after diabetes onset, while the inverse was true of increased SM C22:0, C24:1, C24:0, C26:1, and SM C26:0 . A studyN = 12) . AnotherN = 12) . In contth HFpEF . The LUdth HFpEF . Indepenth HFpEF . Anotherth HFpEF . Newly dth HFpEF . A more th HFpEF . A 2020 th HFpEF .total Cer accumulation in HF and HF-related outcomes, holds true. Though these results are from vastly different studies, they consistently show inverse association of the LCFA Cer C16:0, synthesized by CerS5 or CerS6, and the unsaturated VLCFA Cer C24:1, synthesized by CerS2, to HF, regardless of HF classification and population ethnicity. While the saturated VLCFA Cer C24:0 and in some studies the SM C24:0 is positively correlated with HF. Though, ratios of sphingolipid LCFA/VLCFA seem to be better prognostic markers of adverse cardiac events, HF and prediction of HF-mortality up to 5 years.Taken together, these studies suggest the dogma implicating Developing new pharmacotherapies is a costly and time-consuming process, and drug companies are incentivized to speed up the path to market approval due to the time-limits on drug patents. However, testing agents that may augment a specific disease biomarker without fully understanding their potential mechanistic roles has led to failed clinical trials. To increase the likelihood of identifying causal biomarker-disease associations, researchers have started to employ Mendelian randomization (MR). This approach involves a genome-wide association study (GWAS) analysis to identify candidate genetic variants associated with variations in the biomarker, which are then compared to those associated with differences in the population. When a biomarker is identified and a causal effect is confirmed by MR, the probability of clinical trial success for interventions targeting that biomarker increases significantly .Although MR analyses have not been reported in the literature for the association between circulating sphingolipids and HF, a GWAS analysis has been conducted for circulating sphingolipids associated with a reduction in HF events. MR analyses for sphingolipids linked to incident coronary heart disease (CHD) have also been reported, which is relevant since CHD and HF can coexist in the same patient, and patients with CHD are at increased risk for development of HF , 68, 69.VLCFA Cer measured in the plasma, has been shown to be prognostic for incident HF events (relative risk of 0.75 for every 3 unit increase in plasma Cer C24:0 for HF) despite traditionally being implicated in the etiology of T2DM . AnalyseN = 1,094 and N = 4,034), were congruent with the FHS findings. Variants near SPTLC3 were confirmed to be contributing factors in the variations of circulating Cer linked to CVD and T2DM . SM C28:1, one of the biomarkers identified, showed an inverse relationship with incidence of CHD, but was not found to have a causal relationship by MR analysis , 7372, 7HFpEF is a complex condition that only a few animal models have been able to replicate successfully. Animal models of HFpEF are typically pressure overload models as 55%–86% of patients with HFpEF have hypertension. The Dahl salt-sensitive (Dahl/SS) rat is the most popular HFpEF model and is characterized by hypersensitivity to sodium intake. Other popular models include animals with transverse aortic constriction (TAC), and gene knockout (KO), or overexpression (OE) that develop left ventricular hypertrophy (LVH).total Cer, aggravating contractile dysfunction, whereas the LVH mice on HFD showed a similar phenotype as the WT and no accumulation of total Cer, suggesting that impaired fatty acid oxidation in this model is associated with Cer lipotoxicity (β-oxidation. Hearts from heterozygous CPT1b knockout (CPT1b+/−) mice subjected to TAC-induced pressure overload, showed substantially elevated total Cer levels driven by the Cer C16:0, C18:0, and C24:0 species, while sham WT and CPT1b+/− mice showed no differences with respect to Cer content , 1.44× SM , and 1.32× Sph . Anothertoxicity . CPT1b c content . While wDgat in mice led to no known cardiac dysfunction, and these mice express normal levels of circulating Cer. However, half of the cardiomyocyte-specific DGAT1 KO mice died within 9 months and their hearts showed an 85% and 95% increase in total Cer and DAGs, respectively, compared to littermate controls  have a mutation in the Insulin2 gene and are a good non-obese type 1 diabetes mellitus (T1DM) model. At adulthood, the hearts of these mice had accumulated Cer C18:0 and exhibited preserved systolic function, reduced diastolic function, and increased inflammation. This lipotoxic cardiomyopathy phenotype was reversed with insulin replacement therapy -induced HF (cardiac-specific dnPI3K-Mst1 KO) showed significant increases of Cer C16:0 and the most abundant mammalian ganglioside, GM3 C16:0, in atria of AF + HF mice compared to control . These cMost HF animal models are used in research to study HFrEF due to the clear clinical diagnostic criteria, and parameters. The two main animal models for HFrEF are ischemia/infarction and pacing models . Other lvia permanent LAD ligation showed higher Cer C16:0 in the heart tissue compared to sham-operated mice , and partial reversal with ischemic preconditioning. The most significant being that of Cer C16:0, C18:0, C18:1, C18:2, C20:4, C22:5, and Cer C22:6  coronary artery. A study on this model found that stearoyl-SM was reduced by less than 50% in heart tissue 5 days post permanent-LAD ligation in mice compared to mice with sham surgery . Rats wited mice . 8 weekson of HF , 122. Iser C22:6 . These sGreat strides have been made in understanding the sphingolipidome in patients with HF. The Mayo Clinic's CERAM panel's ratios of the VLCFA to MCFA Cer are becoming increasingly popular for assessing the risk of HF development and progression. Evidence suggests these novel Cer biomarkers will continue be more successful than conventional risk factors in predicting adverse cardiac events and the onset of HF, especially when applied on a larger scale. However, there is still no clear evidence to show mechanistically how Cer C24:0 is linked to beneficial pathways or Cer C16:0 to adverse pathways in HF. More studies are needed to determine how Cer (and other sphingolipids) with different acyl chain lengths regulate various signaling pathways. Despite these developments, prognosis for HFrEF and HFpEF patients remain poor. Thus, when considering future studies in humans and animal models of HF some points should be considered.The first point to consider: different results can be obtained from different materials analyzed, and while tissue sphingolipid levels are useful in patients receiving non-pharmacological intervention such as LVAD, it is of more clinical benefit to discover potent biomarkers in circulation, i.e., serum or plasma. This is due to ease of accessibility, and ability to collect samples more frequently in follow-up appointments. Nonetheless, tissue sphingolipid levels are important In fact, tissue from HF patients prior to LVAD showed induction of SPTLC3 at the protein level, a follow-up showed reduced SPTLC3 levels post-VAD .de novo sphingolipid synthesis pathway can make very diverse sphingolipids in turn triggering unique signaling cascades. While d18 backbone sphingolipids are the most abundant and well-studied, there is mounting evidence to suggest substrate availability is the major determinant for the type of sphingolipids synthesized. For example, variable amino acid substitutes for serine, such as alanine or glycine, result in loss of the hydroxyl group at the alpha carbon to produce deoxysphingolipids. Deoxysphingolipids have been shown to play important roles in pathogenesis of both T1 and T2D. SPTLC3 can use isoleucine, a branched chain amino acid, to create methyl-branched long chain based sphingolipids, the implications of these in HF have yet to be elucidated  approaches have advanced yielding thousands of distinct MS peaks representing individual sphingolipid species and their metabolites , 127. Ta


Hypertensive heart disease (HHD) remains a major global public health concern despite the implementation of new approaches for the management of hypertensive patients. The pathological changes occurring during HHD are complex and involve the development of structural and functional cardiac abnormalities. HHD describes a broad spectrum ranging from uncontrolled hypertension and asymptomatic left ventricular hypertrophy (LVH), either a concentric or an eccentric pattern, to the final development of clinical heart failure. Pressure-overload-induced LVH is recognised as the most important predictor of heart failure and sudden death and is associated with an increased risk of cardiac arrhythmias. Cardiac arrhythmias are considered to be one of the most important comorbidities affecting hypertensive patients. This is the second part of a three-part set of review articles. Here, we focus on the macrostructural and functional abnormalities associated with chronic high pressure, their involvement in HHD pathophysiology, and their role in the progression and prognosis of HHD. A systematic analysis for the Global Burden of Disease Study 2019 demonstrated that hypertensive heart disease (HHD) showed decreasing trends in the number of disability-adjusted life-years (DALYs) in different age groups between 1990 and 2019 ,4. The pThis is the second part of a three-part set of review articles. The published Part 1 dealt with pathophysiological and microstructural changes in HHD . Here, wThe methods applied for the preparation of the article were published in Part 1 of this review series . In brieThe physiological function of the LA has conventionally been divided into three parts: (a) as a reservoir: the LA collects pulmonary venous return during LV systole and isovolumetric relaxation, (b) as a conduit: the LA passively transports stored blood into the LV during ventricular diastole, (c) as a contractile pump: the LA actively contracts and increases LV filling during the final phase of diastole, providing 15% to 30% of total LV filling ,9. Thus,The LA is innervated by the autonomic nervous system (ANS), which modulates LA electrical and contractile activity. The atria receive both sympathetic and parasympathetic innervation, and stimulation or imbalance of either component of the ANS may be associated with increased atrial arrhythmogenicity .It is known that any architectural or structural change in the atrial myocardium can cause significant electrophysiological disturbances. Furthermore, atrial cells respond quickly and comprehensively to various pathological stimuli. In connection with this, it is possible that the atrial myocardium is more sensitive to these stimuli than the ventricular myocardium. Depending on conditions, atrial responses include cardiomyocyte hypertrophy and contractile dysfunction, arrhythmogenic changes, atrial fibroblast proliferation, hyperinnervation, and thrombogenic changes . ExperimMicrostructural changes in the LA precede macroscopic ones such that an atrium of normal size may already have lost its normal compliance and contractility . HTN mayIncreased afterload of the LV results in higher LV filling pressures, which, in turn, may increase LA pressure, which causes increased atrial wall stress. Due to their thin walls, atria are particularly sensitive to high pressure, which is a trigger for atrial remodelling . LA remo2+ release, and (c) the release of renin–angiotensin–aldosterone system (RAAS) neurohormones, natriuretic peptides and endothelin-1 (ET-1) ..49].Those with eccentric hypertrophy are more likely to develop HFrEF than those with concentric hypertrophy, and at the same time, there are data that there may be a direct progression to HFrEF without concentric hypertrophy ,49. It iPatients with HFmrEF have, on average, features that are more similar to HFrEF than HFpEF, and are less likely to have AF and non-cardiac comorbidities . HoweverIn hypertensive humans, sustained pressure overload leads to the progression of diastolic dysfunction, decompensation of LV concentric hypertrophy, and hypertensive HFpEF development. In contrast, sustained volume overload leads to LV dilatation progression, decompensation of eccentric remodelled LV, and HFrEF development ,110. In Borlaug and Redfield named conditions predisposing accelerated myocyte dysfunction on the background of hypertension a “second hit” . Such a It is important to remember that a significant portion of HFpEF patients do not have evidence of LVH on echocardiogram or have apparently normal measurements of diastolic function, but conversely, many older adults have LVH and/or LV diastolic dysfunction but do not have clinical evidence of HFpEF . This suOne additional factor that has to be kept in mind is that chronic hypertension and HFpEF are often accompanied by renal dysfunction, and this combination is associated with greater LVH and worse cardiac haemodynamics, leading to RV dysfunction and worse prognosis . Also, mIn advanced HF, systolic BP is usually low, even in previously hypertensive patients. This phenomenon is termed “decapitated hypertension” and is used to describe the decrease in BP resulting from reduced systolic function and fall in cardiac output despite the presence of compensatory mechanisms such as peripheral vasoconstriction, contributing to the progression of HF .Based on the current evidence, the guidelines recommend the treatment of HTN in patients with HF with an initial combination of ACE inhibitors or ARBs, beta-blockers, and a thiazide/thiazide-like diuretic, alternatively a loop-diuretic . If HTN In general, the influence of first-line antihypertensive drugs on the risk of HF development and on the course of HF in hypertensive patients is under great attention in appropriate guidelines and the consensus is that the most important task is to treat blood pressure to the recommended targets by the use of first line antihypertensive treatments ,6,94. HoThe right ventricle (RV) and right atrium (RA) have long been considered unessential in patients with HTN. In recent years, with the arrival of new reliable and reproducible techniques such as speckle tracking imaging, 3D echocardiography, and cardiac magnetic resonance (CMR), the ability to image and assess various aspects of the RV and RA in hypertensive patients has increased significantly.Recent studies have revealed functional and mechanical changes as well as structural remodelling of the right heart in patients with HTN that have not been previously associated with HTN . Thus, RBased on the results of their study, Zhang et al. suggested that regional diastolic dysfunction in patients with primary HTN may occur first in the RV before the LV , which wLiu and colleagues underlined three mechanisms that may be responsible for structural remodelling of the RV in HTN and may explain the pathophysiological processes linking the right heart and the development and progressing of HHD . First, A cross-sectional study investigating the influence of different LV geometric patterns via 2D and 3D echocardiography reported that the RA conduit and reservoir function gradually decreased and that the RA pump function gradually increased from hypertensive patients with normal LV geometry to those with LV dilatation and concentric LVH ,121. It New approaches have shown that RA and RV dilatation and dysfunction provide the background for electrical instability with the onset and recurrence of AF, which is especially frequent among hypertensive patients, and loss of contractile force, leading to the right HF ,126. ThePrevious studies have suggested that subclinical RV dysfunction may be observed early in the course of HTN ,128. TadTaken together, new imaging modalities have revealed significant RV and RA changes in patients with HTN. The causes of right heart adaptions seen in essential HTN are complex and remain largely unclear. At first sight, these adaptions seem subtle and subclinical, but they are associated with adverse outcomes in hypertensive patients and should therefore be included in the clinical assessment protocols of patients with HTN . Also, eThe involvement of the RV and RA in the pathogenesis of HHD and their influence on the clinical pattern and prognosis of HHD have become a point of special interest only in the last decade. This may partly explain the lack of randomised controlled trials designed to evaluate the influence of different groups of antihypertensive drugs on right heart function in HHD patients. A beneficial effect of RAAS inhibitors on RV function has been demonstrated in a few studies . The metEven though it seems to make sense that β-blockade should have a therapeutic benefit in the failing RV, there is no evidence of the benefit of β-blockers in RV failure on the background of HTN. Moreover, the results in adults with RV failure are controversial ,134,135.Further controlled trials are needed to better elucidate the influence and mechanisms of antihypertensive treatment effects on the right heart in HHD patients.HHD is associated with different cardiac supraventricular and ventricular arrhythmias, most commonly AF, especially in those with LVH or HF.LA dilatation, a hallmark of atrial structural remodelling, was non-invasively recorded via ECG over decades until echocardiography took over as the standard tool to evaluate left atrial size and nowadays also function. As early as 1963, G. Ross demonstrated the increase in P-wave duration and amplitude in hypertensive patients and the correlation of P-wave characteristics with different forms of HTN ,136. EarIt is considered that several different mechanisms contribute to the pathophysiology of arrhythmias in HHD.+, Ca2+) and propensity for arrhythmia in HHD [2+ handling is thought to contribute to the progression of HHD and may be a trigger for atrial tachyarrhythmias. [Elevated systemic pressure influences the size and function of the LA, which makes uncontrolled HTN a key risk factor predisposing the development of AF ,139,140.a in HHD ,141. Remythmias. .LVH results in significant electrophysiological changes characterised by an increased electrical vulnerability on both cellular and tissue levels. These changes result in the dispersion of refractoriness and considerably lower ventricular fibrillation threshold. Moreover, an increased myocardial mass and interstitial myocardial fibrosis are associated with reduced coronary flow reserve . MetabolAnimal studies have shown important arrhythmogenic effects of angiotensin II on the cardiac conduction system . RAAS acRecently published studies suggested that atrial epicardial adipose tissue could upregulate the expression of some pro-inflammatory and pro-fibrotic adipokines while downregulating the expression of anti-fibrotic cytokines and predispose AF . However2+ handling may serve as the trigger initiating the first episodes of AF [As the most often encountered arrhythmia, AF is associated with an increased risk of death, stroke/systemic embolism, and bleeding ,149. LVHes of AF ,147. Froes of AF . Moreovees of AF ,151.Generally, an increased likelihood for inducible atrial tachyarrhythmias has been shown for different animal models of HTN . The rolIn patients with documented AF, hypertension increases the risk of thromboembolism and bleeding, which was confirmed in studies in anticoagulated patients with AF, and this facilitates the progression from paroxysmal to persistent or permanent AF ,154. AtrPrevious studies demonstrated that both SV and ventricular premature beats occur frequently in hypertensive patients with LVH . In a reA subanalysis of the LIFE study found that the presence of a left bundle branch block (LBBB) significantly increased CV mortality (1.6-fold more), risk of sudden cardiac death (3.5-fold more), and symptomatic HF (1.7-fold more) in patients with hypertension and LVH compared with those without LBBB .The intensive studying of the relationship between AF and HTN in recent decades has provided valuable information to many questions, but still aspects from a clinical point of view remain not fully elucidated. Therefore, it is not clear whether the risk of AF increases linearly with BP level or whether there is a certain BP threshold above which the risk of this arrhythmia increases. Additionally, the role of out-of-office BP and the cumulative BP load as risk predictors in the setting of HHD and AF, as well as arrhythmia risk, have not been entirely studied. Further, it is not clear if the use of specific antihypertensive and especially antifibrotic drugs reduces AF burden or the risk of new-onset AF in patients with sinus rhythm. Moreover, the ideal BP level for hypertensive patients with anticoagulated AF to minimize the risk for haemorrhagic and thrombolytic complications is uncertain. Also, the role of novel biomarkers of structural or functional cardiac changes as risk predictors of arrhythmias, including AF, remains to be clarified. As the risk of arrhythmia is closely related to the structure and remodelling of the heart, the role of new imaging techniques needs also to be determined, as well as when they should be used, to properly identify patients with high arrhythmogenic risk in HHD patients.This review attempts to analyse the constellation of the currently available experimental and clinical data on macroscopic structural changes, including left and right heart alterations, heart failure, and rhythm and conduction disturbances as clinical manifestations of developing and progressing HHD.Thus, over the last decade, a number of extensive research works addressed the complexity of the causal relationships between structural and as well as functional alterations in HHD. The application of modern technology and methodology has resulted in major advances in our understanding of the pathophysiological processes occurring in HHD and its consequences. There is strong evidence for the influence of chronic high blood pressure on changes that occur in the left heart, such LVH and fibrosis, as well as LA remodelling, with their consequences of HF and arrhythmias. However, significant gaps remain in the literature regarding the impact of HTN on the right heart chambers and their involvement and role in HHD progression. It is worth noting that the mechanisms linking LVH to the risk of major CV events as well as the mechanisms by which regression of LVH reduces the risk of CV disease in hypertensive subjects are also still incompletely understood. The correct evaluation of the heterogenous cardiac alterations occurring in HHD is important because they have been established as strong predictors of fatal and non-fatal adverse clinical outcomes in both asymptomatic hypertensive individuals and those with prevalent disease.With the predicted increase in the burden of hypertension and life expectancy over the next few decades, HHD can be expected to have an even greater impact in the future of health care. Further studies are needed to elucidate the relative prognostic contributions of distinct singular or combined phenotypes that constitute HHD and create the optimal approach to stratify and reduce CV in different patterns of HHD.


Our findings promote the exploitation of the multiparametric potential of CMR, including LGE, for better risk stratification for TM patients.We assessed the prognostic value of multiparametric cardiovascular magnetic resonance (CMR) in predicting death from heart failure (HF) in thalassemia major (TM). We considered 1398 white TM patients  without a history of HF at baseline CMR, which was performed within the Myocardial Iron Overload in Thalassemia (MIOT) network. Iron overload was quantified by using the T2* technique, and biventricular function was determined with cine images. Late gadolinium enhancement (LGE) images were acquired to detect replacement myocardial fibrosis. During a mean follow-up of 4.83 ± 2.05 years, 49.1% of the patients changed the chelation regimen at least once; these patients were more likely to have significant myocardial iron overload (MIO) than patients who maintained the same regimen. Twelve (1.0%) patients died from HF. Significant MIO, ventricular dysfunction, ventricular dilation, and replacement myocardial fibrosis were identified as significant univariate prognosticators. Based on the presence of the four CMR predictors of HF death, patients were divided into three subgroups. Patients having all four markers had a significantly higher risk of dying for HF than patients without markers (hazard ratio (HR) = 89.93; 95%CI = 5.62–1439.46;  Beta thalassemia major (β-TM) is a genetic blood disease with a high incidence in the Mediterranean basin, the Middle East, the Indian subcontinent, Central Asia, and the Far East . HoweverThanks to its multiparametric potential, CMR represents a unique tool for the characterization and quantification of myocardial involvement and damage. CMR is the gold standard for the quantification of biventricular size and function by cine images. Because it does not incorporate ionizing radiation, does not exhibit window or geometric limitations, and provides precise ventricular endocardial definition, it allows for highly reproducible and accurate measurements of ventricular volumes. This is of particular value in TM, where the “normal” heart pumps at larger volumes and against lower peripheral resistances than the normal heart in non-thalassemic individuals, and where the heart’s biventricular size can be influenced by the iron previously accumulated. Moreover, late gadolinium enhancement (LGE) CMR is the only non-invasive imaging method that can detect replacement myocardial fibrosis, a common finding among TM patients ,16,17.A study of 481 Italian TM patients showed that heart iron, ventricular dysfunction, and replacement myocardial fibrosis could predict the future development of heart failure. Moreover, all three of these CMR markers remained independent prognosticators in a multivariate model that included a previous history of heart failure . To the The aim of this multicenter study was to evaluate the prognostic value of multiparametric CMR  in predicting death from heart failure in a large cohort of well-treated TM patients.We considered 1485 TM patients  consecutively enrolled in the Myocardial Iron Overload in Thalassemia (MIOT) network, comprising 70 thalassemia centers and 10 magnetic resonance imaging (MRI) centers, where MRI exams were performed using homogeneous, standardized, and validated procedures . The incAll patients were from an Italian background and were uniformly treated. They had been regularly transfused since early childhood and started undergoing chelation therapy from the mid-to-late 1970s, whereas patients born after the 1970s received chelation therapy from early childhood.All patients performed their first MRI scan between April 2006 and November 2015. All scans were performed in the week immediately prior to the scheduled blood transfusion. The clinical-anamnestic history of the patients, from birth to the date of the first MRI scan, was recorded in the MIOT web-based database. At every MRI follow-up, which was performed by protocol every 18 ± 3 months, the clinical, instrumental, and laboratory data were updated.All patients gave informed consent in compliance with the Declaration of Helsinki, and the study was approved by the institutional ethics committees of all MRI sites.All patients underwent MRI using the clinical 1.5 T scanners of three main vendors  equipped with phased-array coils. Breath-holding in end-expiration and ECG-gating were used.®) [For iron overload assessment, a validated T2* gradient–echo multiecho sequence was used. The intersite, interstudy, intraobserver, and interobserver variability of the proposed methodology had been previously assessed ,20. For ®) . The sof®) . The glo®) , and wer®) .Steady-state free precession (SSFP) cines were acquired in sequential 8-mm short axis slices (gap 0 mm) from the atrio-ventricular ring to the apex to assess biventricular function parameters quantitatively in a standard way . Thirty ®; Bayer Schering Pharma; Berlin, Germany) intravenous administration at the standard dose of 0.2 mmol/kg using a fast gradient-echo inversion recovery sequence. In addition, vertical, horizontal, and oblique long-axis views were acquired. Inversion times were adjusted to null the normal myocardium (from 210 ms to 300 ms). LGE was evaluated visually by two independent observers using a two-point scale (enhancement absent or present) and was considered present when visualized in two different views [2 and in patients who refused the contrast medium administration.Late gadolinium enhancement short-axis images were acquired 10–18 min after Gadobutrol , signs, biomarkers, and instrumental parameters, according to the current guidelines .The follow-up date coincided with the date of the last available MRI. For patients who did not perform a follow-up MRI, a case report form detailing patient outcomes between the baseline MRI and September 2018 was completed by the caring hematologist.All data were analyzed using SPSS version 27.0  statistical package.Continuous variables were described as mean ± SD. Categorical variables were expressed as frequencies and percentages.The normality of the distribution of the continuous variables was assessed by using the Kolmogorov–Smirnov test.2 testing was performed for non-continuous variables. The Bonferroni post hoc test was used for multiple comparisons between pairs of groups.For continuous values with normal distributions, comparisons between groups were made by performing the independent-samples t-test (2 groups) or a one-way analysis of variance (ANOVA) (>2 groups). Wilcoxon’s signed rank test or the Kruskal–Wallis test were applied for continuous values with non-normal distribution. χCorrelation analysis was performed using Pearson’s test or Spearman’s test where appropriate.The Cox proportional hazard model was used to test the association between the considered prognostic variables and the outcome (HF death). The results are presented as hazard ratios (HR) with 95% confidence intervals (CI). Kaplan–Meier curves were generated by relating the development of an outcome over time to each significant prognosticator. The log rank test was used to compare different strata in Kaplan–Meier analyses.p < 0.05 was considered statistically significant.In all tests, a 2-tailed At the baseline MRI, eighty-seven (5.9%) patients had a history of heart failure and were excluded from this study.p = 0.001), significantly lower global heart T2* values , a significantly higher number of segments with a T2* < 20 ms , significantly higher LV EDVI  and RV EDVI , and significantly lower LV EF  and RV EF .Compared to HF-free patients, patients with a history of HF were characterized at the baseline MRI by a significantly higher age , but they showed comparable global heart T2* values .The contrast medium was not administrated in 286 (20.5%) patients. Among the 187 (16.8%) patients with replacement myocardial fibrosis, none had an ischemic pattern, and two or more foci were detected in 59.9% of cases. The septum was involved in 80.6% of the cases. Patients with replacement myocardial fibrosis were significantly older than patients without replacement myocardial fibrosis  and a significant inverse correlation with global heart T2* values . A significant inverse correlation was detected between global heart T2* and MRI LIC values . Global heart T2* values were not correlated with biventricular volume indexes or LV cardiac indexes but showed a weak positive association with both LV EF  and RV EF .At baseline, serum ferritin levels showed a significant positive correlation with MRI LIC values .At the baseline MRI, patients received the following chelation regimens: deferoxamine (33.3%), deferiprone (19.0%), deferasirox (25.0%), combined deferoxamine + deferiprone (17.2%), sequential deferoxamine/deferiprone (5.0%), and others (0.5%).p < 0.0001) and to have a baseline LIC ≥ 3 mg/g/dw .During the follow-up, 49.1% of the patients changed their chelation regimen at least once, i.e., they switched to a different type of chelator or underwent modification of dose and/or frequency. Compared to patients who maintained the same regimen, those who changed the chelation regimen were more likely to have a baseline global heart T2* value < 20 ms .The percentage of patients with good compliance (correspondence between the time history of drug administration and the prescribed regimen > 60%) was significantly higher at the end of the study than at the baseline MRI  patients died from heart failure. Ten patients had HF with reduced EF at echocardiography. The majority of them presented to the healthcare provider with a reduction in their effort tolerance due to dyspnea and/or fatigue. One patient presented not only with fatigue but also with chest pain and tachycardia and had elevated troponin levels. Two patients presented with palpitations. Two patients had chronic heart failures diagnosed >1 year after the CMR scan, that rapidly worsened. One patient had an HF with mildly reduced EF. One patient had HF with preserved EF and had evidence of structural heart disease. The mean age at death was 35.06 ± 8.68 years (range: 17–47 years).Mean time from the baseline MRI to the HF-related death was 1.68 ± 1.78 years and, six (50.0%) deaths occurred within the first year of follow-up.When compared to the other patients, patients who died by HF showed at the baseline MRI significantly higher serum ferritin levels and MRI LIC values, significantly lower global heart T2* values, a significantly higher numbers of segments with T2* < 20 ms, significantly lower biventricular EFs, and a significantly higher incidence of replacement myocardial fibrosis .One patient who died from HF had a previous history of myocarditis.p = 0.010, ventricular dysfunction: p = 0.030, ventricular dilation: p < 0.0001, and replacement myocardial fibrosis: p = 0.010).(1)N = 488);patients with none of the four CMR markers N = 617);patients with one to three CMR markers N = 7).patients with four CMR markers . Global heart T2* values were significantly lower in group 2 than in groups 0 and 1  and in group 1 than in group 0 (p < 0.0001), whereas the number of segments with a T2* < 20 ms was significantly higher in group 2 than in groups 0 and 1  and in group 1 than in group 0 (p < 0.0001). Significantly lower LV EF and RV EF values were found in group 2 compared to both group 1 (p < 0.0001 for both ventricles) and group 0 (p < 0.0001 for both ventricles) and in group 1 compared to group 0 (p < 0.0001 for both ventricles). LV EDVI and RVEDVI were significantly increased in group 2 compared to group 1 (p < 0.0001 for both ventricles) and to group 0 (p < 0.0001 for both ventricles) and in group 1 compared to group 0 . The frequency of replacement myocardial fibrosis was significantly higher in group 2 than in groups 1 and 0 and in group 1 than in group 0 .p < 0.0001) and group 1   .p = 0.001) or with one to three CMR markers . p < 0.0001).Patients having all four markers had a significantly higher risk of dying by HF than patients without markers  during the follow-up were more likely to have significant MIO at baseline. Moreover, all MRI scans were performed after 2006, the year when a new era of chelation treatment had started thanks to the availability in the clinical arena of three different iron chelators and the evidence that they could be used in association to intensify chelation or make it more tolerable .No prospective association was detected between hepatic iron or serum ferritin levels and HF mortality. These parameters cannot be used to infer cardiac iron status, as demonstrated by weak cross-sectional correlation with the cardiac T2* found in the present study and in other published studies ,34,35,36As expected, MIO was a significant prognosticator of HF death. Excess iron can be detrimental to human cells through the production of hydroxyl radicals via Haber–Weiss–Fenton reactions, which cause oxidative damage to cellular components like lipids, proteins, and DNA ,42. FreeImportantly, when the four CMR indices  were evaluated in combination, they fine-tuned the prognostic stratification of TM patients. Thus, the results of our study strengthen the usefulness of a multiparametric CMR approach which integrates biventricular ejection fractions and volumes and LGE with cardiac T2* to further ameliorate the prognosis of TM patients via the early identification of high-risk patients. Conversely, relying only on cardiac T2* as a unique marker of cardiac death may lead to suboptimal prognostic stratification.It deserves mention that in our study, both ventricular dysfunction and dilation were diagnosed using previously derived “normal for TM” reference ranges in order to avoid a misdiagnosis of cardiomyopathy (underdiagnosis of dysfunction and overdiagnosis of dilatation) . Indeed,This study suffers from several limitations.The small number of HF deaths that occurred during the follow-up did not allow us to perform a multivariate analysis that included all variables identified in the univariate analysis. For this reason, we performed a model with the four CMR univariate prognosticators.The prognostic value of the CMR mapping techniques  was not evaluated because they were not available at the time of patient enrolment.We did not measure myocardial deformation (strain), which could be a more sensitive marker of myocardial dysfunction than EF . AlthougMore studies are needed to evaluate the transferability of our results to other TM populations with a lower prevalence of HCV infection, in which a lower frequency of myocardial fibrosis may be expected.In TM patients, significant MIO, ventricular dysfunction, ventricular dilation, and replacement myocardial fibrosis were associated with a significantly higher risk of heart failure death, and the combined use of all four CMR indexes provided incremental prognostic information. Hence, the present study’s findings promote exploiting the multiparametric potential of CMR, including LGE, for better risk stratification for TM patients. Further studies are needed to verify if, in addition to the adjustment of iron chelation therapy, the adoption of treatment directed to myocardial performance may further open the prognosis of TM patients.