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Comparing the effects of chemical Ca2+ dyes and R-GECO on contractility and Ca2+ transients in adult and human iPSC cardiomyocytes.
We compared commonly used BAPTA-derived chemical Ca2+ dyes (fura2, Fluo-4, and Rhod-2) with a newer genetically encoded indicator (R-GECO) in single cell models of the heart. We assessed their performance and effects on cardiomyocyte contractility, determining fluorescent signal-to-noise ratios and sarcomere shortening in primary ventricular myocytes from adult mouse and guinea pig, and in human iPSC-derived cardiomyocytes. Chemical Ca2+ dyes displayed dose-dependent contractile impairment in all cell types, and we observed a negative correlation between contraction and fluorescence signal-to-noise ratio, particularly for fura2 and Fluo-4. R-GECO had no effect on sarcomere shortening. BAPTA-based dyes, but not R-GECO, inhibited in vitro acto-myosin ATPase activity. The presence of fura2 accentuated or diminished changes in contractility and Ca2+ handling caused by small molecule modulators of contractility and intracellular ionic homeostasis (mavacamten, levosimendan, and flecainide), but this was not observed when using R-GECO in adult guinea pig left ventricular cardiomyocytes. Ca2+ handling studies are necessary for cardiotoxicity assessments of small molecules intended for clinical use. Caution should be exercised when interpreting small molecule studies assessing contractile effects and Ca2+ transients derived from BAPTA-like chemical Ca2+ dyes in cellular assays, a common platform for cardiac toxicology testing and mechanistic investigation of cardiac disease physiology and treatment.
Cohort profile: characterisation, determinants, mechanisms and consequences of the long-term effects of COVID-19 - providing the evidence base for health care services (CONVALESCENCE) in the UK.
PURPOSE: The pathogenesis of the long-lasting symptoms which can follow an infection with the SARS-CoV-2 virus ('long covid') is not fully understood. The 'COroNaVirus post-Acute Long-term EffectS: Constructing an evidENCE base' (CONVALESCENCE) study was established as part of the Longitudinal Health and Wellbeing COVID-19 UK National Core Study. We performed a deep phenotyping case-control study nested within two cohorts (the Avon Longitudinal Study of Parents and Children and TwinsUK) as part of CONVALESCENCE. PARTICIPANTS: From September 2021 to May 2023, 349 participants attended the CONVALESCENCE deep phenotyping clinic at University College London. Four categories of participants were recruited: cases of long covid (long covid(+)/SARS-CoV-2(+)), alongside three control groups: those with neither long covid symptoms nor evidence of prior COVID-19 (long covid(-)/SARS-CoV-2(-); control group 1), those who self-reported COVID-19 and had evidence of SARS-CoV-2 infection, but did not report long covid (long covid(-)/SARS-CoV-2(+); control group 2) and those who self-reported persistent symptoms attributable to COVID-19 but no evidence of SARS-CoV-2 infection (long covid(+)/SARS-CoV-2(-); control group 3). Remote wearable measurements were performed up until February 2024. FINDINGS TO DATE: This cohort profile describes the baseline characteristics of the CONVALESCENCE cohort. Of the 349 participants, 141 (53±15 years old; 21 (15%) men) were cases, 89 (55±16 years old; 11 (12%) men) were in control group 1, 75 (49±15 years old; 25 (33%) men) were in control group 2 and 44 (55±16 years old; 9 (21%) men) were in control group 3. FUTURE PLANS: The study aims to use a multiorgan score calculated as the cumulative total for each of nine domains (ie, lung, vascular, heart, kidney, brain, autonomic function, muscle strength, exercise capacity and physical performance). The availability of data preceding acute COVID-19 infection in cohorts may help identify the consequences of infection independent of pre-existing subclinical disease and also provide evidence of determinants that influence the development of long covid.
Hepcidin and Tissue-Specific Iron Regulatory Networks.
Hepcidin is primarily secreted by the liver and functions as an endocrine hormone. However, a growing number of studies show that hepcidin can also be produced locally by other cells and organs, where it acts in an autocrine/paracrine manner to mediate important iron-dependent pathways. These pathways can operate under normal homeostatic conditions or become relevant in pathophysiological conditions (inflammation, infection, cancer, liver disease, myocardial infarction, etc.). This chapter will delve into the local roles of hepcidin, highlighting its unconventional functions in barrier maintenance, host defense, growth, tissue housekeeping, and injury repair.
Consensus Recommendations for Hyperpolarized [1-13C]pyruvate MRI Multi-center Human Studies.
Magnetic resonance imaging of hyperpolarized (HP) [1-13C]pyruvate allows in-vivo assessment of metabolism and has translated into human studies across diseases at 15 centers worldwide. Consensus on best practice for multi-center studies is required to develop clinical applications. This paper presents the results of a 2-round formal consensus building exercise carried out by experts with HP [1-13C]pyruvate human study experience. Twenty-nine participants from 13 sites brought together expertise in pharmacy methods, MR physics, translational imaging, and data-analysis; with the goal of providing recommendations and best practice statements on conduct of multi-center human studies of HP [1-13C]pyruvate MRI. Overall, the group reached consensus on approximately two-thirds of 246 statements in the questionnaire, covering 'HP 13C-Pyruvate Preparation', 'MRI System Setup, Calibration, and Phantoms', 'Acquisition and Reconstruction', and 'Data Analysis and Quantification'. Consensus was present across categories, examples include that: (i) different HP pyruvate preparation methods could be used in human studies, but that the same release criteria have to be followed; (ii) site qualification and quality assurance must be performed with phantoms and that the same field strength must be used, but that the rest of the system setup and calibration methods could be determined by individual sites;(iii) the same pulse sequence and reconstruction methods were preferable, but the exact choice should be governed by the anatomical target; (iv) normalized metabolite area-under-curve (AUC) values and metabolite AUC were the preferred metabolism metrics. The work confirmed areas of consensus for multi-center study conduct and identified where further research is required to ascertain best practice.
Risk thresholds for soft versus hard cardiovascular disease outcome models for initiating statin therapy among Chinese adults: a cost-utility analysis.
BACKGROUND: Current guidelines for atherosclerotic cardiovascular disease (ASCVD) primary prevention mostly recommend risk scores that predict risk of non-fatal myocardial infarction, fatal ischemic heart disease (IHD), and fatal or non-fatal ischemic stroke (hard outcomes), ignoring the burden from other non-fatal IHD outcomes. We explored the optimal risk thresholds for statin initiation using non-laboratory-based soft and hard ASCVD outcome models and compared the cost-utility of such models in the Chinese population. METHODS: We constructed Markov cohort models to estimate the incidence of ASCVD events, costs, and quality-adjusted life years (QALYs) over a lifetime from a social perspective. The simulation cohort was constructed using data from the China Kadoorie Biobank (CKB). Input data included cost, utility, statin efficacy, and other parameters were derived from published literature. We used CKB-ASCVD models to predict 10-year risk and different risk thresholds to guide statin initiation. The incremental cost-effectiveness ratio (ICER) was estimated as cost per QALY gained. Sensitivity analyses were performed to explore the uncertainty in the models. RESULTS: The optimal risk threshold was 18% for the soft ASCVD model and 10% for the hard ASCVD model, with ICERs of $7013.48/QALY and $6540.71/QALY, respectively. The optimal thresholds were robust in stratified analyses by region and sex, and one-way sensitivity analyses over a wide range of input parameters. Probabilistic sensitivity analyses showed that these optimal thresholds had around 70% chance of being cost-effective. When analyzed by age group, above optimal thresholds were cost-effective in adults aged 30-59 years but not in those aged 60-75 years. The threshold strategies based on soft ASCVD model were mostly cost-saving compared with those based on hard models to treat the same proportions of the population. CONCLUSIONS: The risk threshold of 18% for soft ASCVD model and 10% for hard ASCVD model have acceptable cost-utility profiles in the Chinese population. The soft ASCVD model is more cost-effective than the hard model and should be used as a screening tool for ASCVD primary prevention.
Ketosis regulates K+ ion channels, strengthening brain-wide signaling disrupted by age.
Aging is associated with impaired signaling between brain regions when measured using resting-state fMRI. This age-related destabilization and desynchronization of brain networks reverses itself when the brain switches from metabolizing glucose to ketones. Here, we probe the mechanistic basis for these effects. First, we confirmed their robustness across measurement modalities using two datasets acquired from resting-state EEG (Lifespan: standard diet, 20-80 years, N = 201; Metabolic: individually weight-dosed and calorically-matched glucose and ketone ester challenge, μ a g e = 26.9 ± 11.2 years , N = 36). Then, using a multiscale conductance-based neural mass model, we identified the unique set of mechanistic parameters consistent with our clinical data. Together, our results implicate potassium (K+) gradient dysregulation as a mechanism for age-related neural desynchronization and its reversal with ketosis, the latter finding of which is consistent with direct measurement of ion channels. As such, the approach facilitates the connection between macroscopic brain activity and cellular-level mechanisms.
Verification and Adjudication of Health Outcomes in Prospective Cohort Studies
The value of prospective biobank studies critically depends on their ability to collect large number of well-characterised disease outcomes on study participants over a prolonged period of time. In contrast with case-control studies which typically collect data on disease cases directly from hospitals, prospective studies collect incident disease outcomes during follow-up over several years or decades by linkage to death or disease registries. Verification of reported disease outcomes in prospective studies is the process of independent validation of the reporting sources for disease outcomes. Adjudication is the process of independent review of all the available evidence on clinical symptoms, signs, imaging, biochemical or histological investigations to classify reported disease outcomes into major disease types and their pathological and/or aetiological sub-types. Hence, disease verification and disease adjudication are complimentary processes to verify the accuracy of reported diagnoses and to classify major diseases into their pathological sub-types. Since most major diseases present as clinical syndromes, reliable classification of disease outcomes is required for studies of genetic and other determinants of such diseases. Verification and adjudication systems require collection of additional information from external sources, including disease registers or medical records from hospitals or primary health care systems for independent review by clinical specialists. The design and implementation of practical procedures needed for disease verification and adjudication in large prospective studies requires feasible and cost-effective systems. Both verification and adjudication systems require regulatory approval to safeguard the confidentiality of personally identifiable data. This chapter discusses the principles and practical procedures required to establish systems for verification and adjudication of disease outcomes in large prospective studies, which will also be of general relevance for other studies.
Metabolic flexibility and reverse remodelling of the failing human heart.
BACKGROUND AND AIMS: Cardiac resynchronization therapy (CRT) produces long-term reverse remodelling which requires greater adenosine triphosphate delivery to the contractile machinery. Whilst the heart retains some metabolic flexibility in non-ischaemic cardiomyopathy, whether this correlates with reverse remodelling is unknown. This study investigated whether CRT acutely changes cardiac substrate uptake, and whether this translates to favourable reverse remodelling. METHODS: The effect of CRT on cardiac substrate uptake was assessed via direct coronary flow and arteriovenous measurements, with metabolomic/lipidomic analysis on infusions of insulin/glucose and intralipid. Cardiac function was assessed with left ventricular pressure-volume loops during implantation, and cardiac magnetic resonance before and 6 months following CRT, with and without biventricular pacing. RESULTS: Regardless of substrate infusion, CRT acutely improved stroke work without increasing O2 uptake on both insulin/glucose (by 34%, P = .05) and intralipid (by 36%, P = .03). This was followed by increased fatty acid (FA) uptake on insulin/glucose (R = 0.89, P = .03) and increased β-hydroxybutyrate uptake (R = 0.81, P = .05) during intralipid infusion. After 6 months, there was a 48% (P < .001) reduction in left ventricular end diastolic volume, beyond that achievable by acutely shortening or lengthening QRS duration. Reverse remodelling significantly correlated with increased FA uptake with CRT on insulin/glucose (R = 0.71, P = .05) driven by long and medium chain uptake, and increased ketone uptake with CRT on intralipid (R = 0.79, P = .05). CONCLUSIONS: CRT acutely alters the metabolic phenotype of non-ischaemic cardiomyopathy towards a more physiological picture of FA uptake which correlates with reverse remodelling. Retained metabolic flexibility may therefore be critical for subsequent reverse remodelling.
Complex relationship between Amerindian ancestry and obesity in the Mexican population.
The high prevalence of obesity in Mexican-American populations in the United States has suggested that the different genetic composition of the Mexican population may be related to the high prevalence of obesity in Mexico. Recently, the genome of 140,000 individuals in the Mexico City Prospective Study (MCPS) cohort was explored, and it was found that the average Amerindian ancestry (AMR) was 66.2%, followed by European (29.2%), African (3.7%), and Asian (0.8%) ancestries. However, the proportions of ancestry vary by geographic region of the country, with an increasing gradient of AMR from north to south. Despite the importance of this relationship, there are few studies that have analyzed the relationship between obesity and AMR, and the results are controversial. The relationship between AMR and central obesity has been more consistent, especially in women. Few genetic variants associated with obesity have been found in Mexico, due to the small number of individuals analyzed. Future analysis of the MCPS cohort will likely clarify the relationship between AMR and obesity, and identify genetic variations and genes associated with obesity and other metabolic diseases, specific to the Amerindian genome.
Genetic Risk and Prognosis of the First Incident Stroke Survivors: Findings from China Kadoorie Biobank and UK Biobank.
BACKGROUND AND OBJECTIVES: Stroke is known for its poor prognosis. Although genetic instruments have shown promise in stratifying first stroke risk in the general population, it is unknown whether they are associated with stroke prognosis. Our study aims to explore the role of genetic risk of stroke in the progression from stroke-free to first stroke and then to recurrent stroke, subsequent coronary heart disease (CHD), and death in China and the United Kingdom. METHODS: We used data from 2 prospective population-based cohorts, China Kadoorie Biobank (CKB) and UK Biobank (UKB). Participants who were unrelated and free of stroke and CHD at baseline were included. Genetic risks of stroke were quantified using integrative polygenic risk scores (iPRSs), which incorporated summary statistics from multiple genome-wide association studies for stroke outcomes and its subtypes, and vascular-risk traits. We used a multistate model to analyze the roles of genetic risk in the transitions from baseline to first incident stroke and then to recurrent stroke, subsequent CHD, and death. RESULTS: Our study included 80,908 CKB participants and 380,348 UKB participants, with mean ages (% female) of 54.0 years (58.6%) and 56.1 years (55.4%). During median follow-ups of 11.9 years and 13.4 years in the CKB and UKB, respectively, 13,481 and 5,772 participants had their first stroke, neither experienced a CHD, or died within 28 days. These survivors had 5,707 and 943 recurrent strokes, as well as 1,196 and 418 CHD events, respectively. iPRSs were associated with recurrent stroke and CHD among stroke survivors in both populations. The corresponding hazard ratios (HRs) and 95% CIs per SD of iPRSs were 1.08 (1.05-1.11) and 1.08 (1.02-1.15) in CKB and 1.11 (1.03-1.19) and 1.23 (1.10-1.37) in UKB, respectively. There was no association between iPRSs and mortality risk. When we further divided the first stroke into 4 pathologic subtypes, both populations revealed statistically significant associations between iPRSs and the transitions from first ischemic stroke to recurrent stroke and CHD. DISCUSSION: Our study shows that the genetic risk of first stroke also influences the prognosis of stroke survivors, indicating that PRS has the potential to improve stroke prognosis.
Baseline serum ferritin predicts myocardial iron uptake following intravenous iron therapy - a hypothesis-generating study.
AIMS: Many patients with heart failure (HF) are iron-deficient. Intravenous (IV) iron therapy improves symptoms and reduces hospitalizations for HF. Several mechanisms have been proposed, including myocardial iron repletion. However, it is unknown if serum iron markers predict the extent of this repletion. To address this question, data from two clinical studies that evaluated changes in myocardial iron using cardiac magnetic resonance (CMR) were harnessed. METHODS AND RESULTS: The Myocardial-IRON trial measured change in myocardial iron, denoted by a decrease in CMR T1 and T2*, at 7 and 30 days after IV ferric carboxymaltose (FCM) in patients with iron deficiency (ID) and HF (n = 53). The STUDY trial measured myocardial and spleen iron at multiple timepoints after FCM in patients with ID without HF (n = 12). In this post-hoc analysis, we examined the association between baseline serum iron markers (transferrin saturation and ferritin) and change in myocardial iron in the weeks after FCM therapy. Changes in spleen iron were also examined, due its role as an intermediary in the redistribution of iron from iron-carbohydrate complexes such as FCM. In patients with or without HF, higher serum ferritin at baseline predicted lower rise in myocardial iron in the weeks after therapy with FCM. In contrast, higher serum ferritin at baseline predicted a greater rise in spleen iron. CONCLUSIONS: These data point towards the hypothesis that functional ID, which is characterized by elevated ferritin, could limit myocardial iron repletion after IV iron therapy, by favouring iron trapping in the spleen.