Mitochondrial Ca2+ Uniporter Is a Mitochondrial Luminal Redox Sensor that Augments MCU Channel Activity.

Ca2+ dynamics and oxidative signaling are fundamental mechanisms for mitochondrial bioenergetics and cell function. The MCU complex is the major pathway by which these signals are integrated in mitochondria. Whether and how these coactive elements interact with MCU have not been established. As an approach toward understanding the regulation of MCU channel by oxidative milieu, we adapted inflammatory and hypoxia models. We identified the conserved cysteine 97 (Cys-97) to be the only reactive thiol in human MCU that undergoes S-glutathionylation. Furthermore, biochemical, structural, and superresolution imaging analysis revealed that MCU oxidation promotes MCU higher order oligomer formation. Both oxidation and mutation of MCU Cys-97 exhibited persistent MCU channel activity with higher [Ca2+]m uptake rate, elevated mROS, and enhanced [Ca2+]m overload-induced cell death. In contrast, these effects were largely independent of MCU interaction with its regulators. These findings reveal a distinct functional role for Cys-97 in ROS sensing and regulation of MCU activity.

Left ventricular ejection fraction digit bias and reclassification of heart failure with mildly reduced vs reduced ejection fraction based on the 2021 definition and classification of heart failure.

AIMS: Aims were to evaluate (1) reclassification of patients from heart failure with mildly reduced (HFmrEF) to reduced (HFrEF) ejection fraction when an EF = 40% was considered as HFrEF, (2) role of EF digit bias, ie, EF reporting favouring 5% increments; (3) outcomes in relation to missing and biased EF reports, in a large multinational HF registry. METHODS AND RESULTS: Of 25,154 patients in the European Society of Cardiology (ESC) HF Long-Term registry, 17% had missing EF and of those with available EF, 24% had HFpEF (EF≥50%), 21% HFmrEF (40%-49%) and 55% HFrEF (<40%) according to the 2016 ESC guidelines´ classification. EF was "exactly" 40% in 7%, leading to reclassifying 34% of the HFmrEF population defined as EF = 40% to 49% to HFrEF when applying the 2021 ESC Guidelines classification (14% had HFmrEF as EF = 41% to 49% and 62% had HFrEF as EF≤40%). EF was reported as a value ending with 0 or 5 in ∼37% of the population. Such potential digit bias was associated with more missing values for other characteristics and higher risk of all-cause death and HF hospitalization. Patients with missing EF had higher risk of all-cause and CV mortality, and HF hospitalization compared to those with recorded EF. CONCLUSIONS: Many patients had reported EF = 40%. This led to substantial reclassification of EF from old HFmrEF (40%-49%) to new HFrEF (≤40%). There was considerable digit bias in EF reporting and missing EF reporting, which appeared to occur not at random and may reflect less rigorous overall care and worse outcomes.

Self-propelling thrombin powder enables hemostasis with no observable recurrent bleeding or thrombosis over 3 days in a porcine model of upper GI bleeding.

BACKGROUND AND AIMS: Hemostatic powders used to manage upper GI bleeding continue to exhibit high recurrent bleeding rates. Previously, self-propelling thrombin powder (SPTP) sprayed endoscopically managed severe Forrest class 1A bleeding. Here, we evaluate SPTP in a 3-day recovery model of diffuse ulcerated bleeding. METHODS: Five anesthetized pigs underwent an endoscopic mucosal snare resection to trigger diffuse ulcer bleeding and were treated with SPTP. The time to hemostasis and the amount of powder delivered were measured. Pigs were recovered and monitored. RESULTS: Five pigs achieved hemostasis in 4.5 ± 1.2 minutes At 3 days after the procedure, the pigs were rescoped and showed no recurrent bleeding. Measured blood parameters were not significantly different from baseline. There were no signs of foreign bodies or thromboembolism during gross necropsy and histopathology of key organs. CONCLUSIONS: SPTP is a promising novel material that stopped diffuse ulcer bleeding in 5 pigs without recurrent bleeding or adverse local or systemic events.

Emerging gene therapies for enhancing the hemostatic potential of platelets.

Platelet transfusions are an integral component of balanced hemostatic resuscitation protocols used to manage severe hemorrhage following trauma. Enhancing the hemostatic potential of platelets could lead to further increases in the efficacy of transfusions, particularly for non-compressible torso hemorrhage or severe hemorrhage with coagulopathy, by decreasing blood loss and improving overall patient outcomes. Advances in gene therapies, including RNA therapies, are leading to new strategies to enhance platelets for better control of hemorrhage. This review will highlight three approaches for creating modified platelets using gene therapies: (i) direct transfection of transfusable platelets ex vivo, (ii) in vitro production of engineered platelets from platelet-precursor cells, and (iii) modifying the bone marrow for in vivo production of modified platelets. In summary, modifying platelets to enhance their hemostatic potential is an exciting new frontier in transfusion medicine, but more preclinical development as well as studies testing the safety and efficacy of these agents are needed.

Comprehensive multicomponent cardiac rehabilitation in cardiac implantable electronic devices recipients: a consensus document from the European Association of Preventive Cardiology (EAPC; Secondary prevention and rehabilitation section) and European Heart Rhythm Association (EHRA).

Cardiac rehabilitation (CR) is a multidisciplinary intervention including patient assessment and medical actions to promote stabilization, management of cardiovascular risk factors, vocational support, psychosocial management, physical activity counselling, and prescription of exercise training. Millions of people with cardiac implantable electronic devices live in Europe and their numbers are progressively increasing, therefore, large subsets of patients admitted in CR facilities have a cardiac implantable electronic device. Patients who are cardiac implantable electronic devices recipients are considered eligible for a CR programme. This is not only related to the underlying heart disease but also to specific issues, such as psychological adaptation to living with an implanted device and, in implantable cardioverter-defibrillator patients, the risk of arrhythmia, syncope, and sudden cardiac death. Therefore, these patients should receive special attention, as their needs may differ from other patients participating in CR. As evidence from studies of CR in patients with cardiac implantable electronic devices is sparse, detailed clinical practice guidelines are lacking. Here, we aim to provide practical recommendations for CR in cardiac implantable electronic devices recipients in order to increase CR implementation, efficacy, and safety in this subset of patients.

Biological nanopores elucidate the differences between isomers of mercaptobenzoic-capped gold clusters.

Identification of isomers using traditional mass spectroscopy methods has proven an interesting challenge due to their identical mass to charge ratios. This proves particularly consequential for gold clusters, as subtle variations in the ligand and cluster structure can have drastic effects on the cluster functionalization, solubility, and chemical properties. Biological nanopores have proven an effective tool in identifying subtle variations at the single molecule limit. This paper reports on the ability of an α-hemolysin (αHL) pore to differentiate between para-, meta-, and ortho- (p-, m-, and o-, respectively) mercaptobenzoic acid ligands attached to gold clusters at the single cluster limit. Detecting differences between p-MBA and m-MBA requires pH-dependent studies that illustrate the role inter-ligand binding plays in stabilizing m-MBA-capped clusters. Additionally, this paper investigates the difference in behavior for these clusters when isolated, and when surrounded by small ligand-Au complexes (AunLm, n = 0, 1, 2… and m = 1, 2,…) that are present following cluster synthesis. It is found that continuous exposure of clusters to freely diffusing ligand complexes stabilizes the clusters, while isolated clusters either disintegrate or exit the nanopore in seconds. This has implications for long term cluster stability.

Passive bilateral leg cycling with concomitant regional circulatory occlusion for testing mechanoreflex-metaboreflex interactions in humans.

PURPOSE: The exercise pressor reflex (EPR) plays a fundamental role in physiological reactions to exercise in humans and in the pathophysiology of cardiovascular disorders. There is no "gold standard" method for EPR assessment; therefore, we propose a new protocol for testing interactions between the muscle mechanoreflex and metaboreflex (major components of EPR). METHODS: Thirty-four healthy subjects (mean age [± standard deviation] 24 ± 4 years, 22 men) were enrolled in the study. During the study, the hemodynamic and ventilatory parameters of these subjects were continuously monitored using our proposed assessment method. This assessment method consists of an initial 5-min rest period (baseline) followed by 5 min of passive cycling (PC) on an automated cycle ergometer (mechanoreceptor stimulation), after which tourniquet cuffs located bilaterally on the upper thighs are inflated for 3 min to evoke venous and arterial regional circulatory occlusion (CO) during PC (metaboreceptor stimulation). Deflation of the tourniquet cuffs is followed by a second 5 min of PC and finally by a 5-min recovery time. The control test comprises a 5-min rest period, followed by 3 min of CO only and a final 5-min recovery. RESULTS: Mean arterial pressure (MAP) and minute ventilation (MV) increased significantly during PC (MAP: from 90 ± 9.3 to 95 ± 9.7 mmHg; MV: from 11.5 ± 2.5 to 13.5 ± 2.9 L/min; both p < 0.05) and again when CO was applied (MAP: from 95 ± 9.7 to 101 ± 11.0 mmHg; MV: from 13.5 ± 2.9 to 14.8 ± 3.8 L/min; both p < 0.05). In the control test there was a slight increase in MAP during CO (from 92 ± 10.5 to 94 ± 10.0 mmHg; p < 0.05) and no changes in the ventilatory parameters. CONCLUSION: Bilateral leg passive cycling with concomitant circulatory occlusion is a new, simple and effective method for testing interactions between the mechanoreflex and metaboreflex in humans.

Ligand-Induced Structural Changes of Thiolate-Capped Gold Nanoclusters Observed with Resistive-Pulse Nanopore Sensing.

Nanopore-based resistive pulse sensing with biological nanopores has traditionally been applied to biopolymer analysis, but more recently, interest has grown in applying the technique to characterizing water-soluble metallic clusters. This paper reports on the use of α-hemolysin (αHL) for detecting a variety of thiolate-capped gold nanoclusters. The ligands studied here are p-mercaptobenzoic acid ( p-MBA), tiopronin (TP), and thiolated PEG7 (S-PEG7). Individual clusters trapped in the cis-side of an αHL pore for extended periods (>10 s) exhibit fluctuations between numerous substates. We compare these current steps between the three different ligands and find that they scale with the mass of the corresponding ligand, which suggests that nanopore sensing could be used to characterize intraparticle surface modifications.

Monitoring functional capacity in heart failure.

This document reflects the key points of a consensus meeting of the Heart Failure Association of European Society of Cardiology (ESC) held to provide an overview the role of physiological monitoring in the complex multimorbid heart failure (HF) patient. This article reviews assessments of the functional ability of patients with HF. The gold standard measurement of cardiovascular functional capacity is peak oxygen consumption obtained from a cardiopulmonary exercise test. The 6-min walk test provides an indirect measure of cardiovascular functional capacity. Muscular functional capacity is assessed using either a 1-repetition maximum test of the upper and lower body or other methods, such as handgrip measurement. The short physical performance battery may provide a helpful, indirect indication of muscular functional capacity.

Heart rate and blood pressure monitoring in heart failure.

It has been long known that incessant tachycardia and severe hypertension can cause heart failure (HF). In recent years, it has also been recognized that more modest elevations in either heart rate (HR) or blood pressure (BP), if sustained, can be a risk factor both for the development of HF and for mortality in patients with established HF. Heart rate and BP are thus both modifiable risk factors in the setting of HF. What is less clear is the question whether routine systematic monitoring of these simple physiological parameters to a target value can offer clinical benefits. Measuring these parameters clinically during patient review is recommended in HF management in most HF guidelines, both in the acute and chronic phases of the disease. More sophisticated systems now allow long-term automatic or remote monitoring of HR and BP and whether this more detailed patient information can improve clinical outcomes will require prospective RCTs to evaluate. In addition, analysis of patterns of both HR and BP variability can give insights into autonomic function, which is also frequently abnormal in HF. This window into autonomic dysfunction in our HF patients can also provide further independent prognostic information and may in itself be target for future interventional therapies. This article, developed during a consensus meeting of the Heart Failure Association of the ESC concerning the role of physiological monitoring in the complex multi-morbid HF patient, highlights the importance of repeated assessment of HR and BP in HF, and reviews gaps in our knowledge and potential future directions.

2016 focused update: clinical recommendations for cardiopulmonary exercise testing data assessment in specific patient populations.

In the past several decades, cardiopulmonary exercise testing (CPX) has seen an exponential increase in its evidence base. The growing volume of evidence in support of CPX has precipitated the release of numerous scientific statements by societies and associations. In 2012, the European Association for Cardiovascular Prevention & Rehabilitation and the American Heart Association developed a joint document with the primary intent of redefining CPX analysis and reporting in a way that would streamline test interpretation and increase clinical application. Specifically, the 2012 joint scientific statement on CPX conceptualized an easy-to-use, clinically meaningful analysis based on evidence-vetted variables in color-coded algorithms; single-page algorithms were successfully developed for each proposed test indication. Because of an abundance of new CPX research in recent years and a reassessment of the current algorithms in light of the body of evidence, a focused update to the 2012 scientific statement is now warranted. The purposes of this update are to confirm algorithms included in the initial scientific statement not requiring revision, to propose revisions to algorithms included in the initial scientific statement, to propose new algorithms based on emerging scientific evidence, to further clarify the application of oxygen consumption at ventilatory threshold, to describe CPX variables with an emerging scientific evidence base, to describe the synergistic value of combining CPX with other assessments, to discuss personnel considerations for CPX laboratories, and to provide recommendations for future CPX research.

Energy-Chirp Compensation in a Laser Wakefield Accelerator.

The energy spread in laser wakefield accelerators is primarily limited by the energy chirp introduced during the injection and acceleration processes. Here, we propose the use of longitudinal density tailoring to reduce the beam chirp at the end of the accelerator. Experimental data sustained by quasi-3D particle-in-cell simulations show that broadband electron beams can be converted to quasimonoenergetic beams of ≤10% energy spread while maintaining a high charge of more than 120 pC. In the linear and quasilinear regimes of wakefield acceleration, the method could provide even lower, subpercent level, energy spread.

2016 Focused Update: Clinical Recommendations for Cardiopulmonary Exercise Testing Data Assessment in Specific Patient Populations.

In the past several decades, cardiopulmonary exercise testing (CPX) has seen an exponential increase in its evidence base. The growing volume of evidence in support of CPX has precipitated the release of numerous scientific statements by societies and associations. In 2012, the European Association for Cardiovascular Prevention & Rehabilitation and the American Heart Association developed a joint document with the primary intent of redefining CPX analysis and reporting in a way that would streamline test interpretation and increase clinical application. Specifically, the 2012 joint scientific statement on CPX conceptualized an easy-to-use, clinically meaningful analysis based on evidence-vetted variables in color-coded algorithms; single-page algorithms were successfully developed for each proposed test indication. Because of an abundance of new CPX research in recent years and a reassessment of the current algorithms in light of the body of evidence, a focused update to the 2012 scientific statement is now warranted. The purposes of this update are to confirm algorithms included in the initial scientific statement not requiring revision, to propose revisions to algorithms included in the initial scientific statement, to propose new algorithms based on emerging scientific evidence, to further clarify the application of oxygen consumption at ventilatory threshold, to describe CPX variables with an emerging scientific evidence base, to describe the synergistic value of combining CPX with other assessments, to discuss personnel considerations for CPX laboratories, and to provide recommendations for future CPX research.

Clinical-Grade-Expanded Regulatory T Cells Prevent Graft-versus-Host Disease While Allowing a Powerful T Cell-Dependent Graft-versus-Leukemia Effect in Murine Models.

We developed a good manufacturing practices-compatible expansion protocol to improve number and purity of regulatory T cells (Tregs) available for clinical trials. Six clinical-grade separation procedures were performed, followed by expansion with high-dose interleukin (IL)-2, anti-CD3/anti-CD28 TCR stimulation, and rapamycin for 19 days achieving a median of 8.5-fold (range, 6.25 to 13.7) expansion. FOXP3 expression was stably maintained over the culture period, while the percentage of CD127 was significantly reduced. The in vitro suppression assay showed a strong Mixed Lymphocytes Reaction inhibition. In vitro amplification did not induce any karyotypic modification. To evaluate the graft-versus-host disease (GVHD)/graft-versus-leukemia (GVL) bifunctional axis, expanded Tregs and conventional T cells (Tcons) were tested in NOD/SCID/IL2Rgnull mice injected with primary acute myeloid leukemia (AML) cells, AML cell line, acute lymphoid leukemia Philadelphia cell line, or Burkitt-like lymphoma cell line. All mice that received leukemia cells together with expanded Tregs and Tcons were rescued from leukemia and survived without GVHD, showing that Treg expansion procedure did not compromise GVHD control and the strong Tcon-mediated GVL activity. This report might represent the basis for treating high-risk leukemia and/or relapsed/refractory leukemia patients with high-dose Treg/Tcons.

Haploidentical hematopoietic transplantation from KIR ligand-mismatched donors with activating KIRs reduces nonrelapse mortality.

Because activating killer cell immunoglobulinlike receptors (KIRs) are heterogeneously expressed in the population, we investigated the role of donor activating KIRs in haploidentical hematopoietic transplants for acute leukemia. Transplants were grouped according to presence vs absence of KIR-ligand mismatches in the graft-vs-host direction (ie, of donor-vs-recipient natural killer [NK]-cell alloreactivity). In the absence of donor-vs-recipient NK-cell alloreactivity, donor activating KIRs had no effects on outcomes. In the 69 transplant pairs with donor-vs-recipient NK-cell alloreactivity, transplantation from donors with KIR2DS1 and/or KIR3DS1 was associated with reduced risk of nonrelapse mortality, largely infection related (KIR2DS1 present vs absent: hazard ratio [HR], 0.25; P = .01; KIR3DS1 present vs absent: HR, 0.18; P = .006), and better event-free survival (KIR2DS1 present vs absent: HR, 0.31; P = .011; KIR3DS1 present vs absent: HR, 0.30; P = .008). Transplantation from donors with KIR2DS1 and/or KIR3DS1 was also associated with a 50% reduction in infection rate (P = .003). In vitro analyses showed that KIR2DS1 binding to its HLA-C2 ligand upregulated inflammatory cytokine production by alloreactive NK cells in response to infectious challenges. Because ∼40% of donors able to exert donor-vs-recipient NK-cell alloreactivity carry KIR2DS1 and/or KIR3DS1, searching for them may become a feasible, additional criterion in donor selection.

A novel role for microphthalmia-associated transcription factor-regulated pigment epithelium-derived factor during melanoma progression.

Microphthalmia-associated transcription factor (MITF) acts via pigment epithelium-derived factor (PEDF), an antiangiogenic protein, to regulate retinal pigment epithelium migration. PEDF expression and/or regulation during melanoma development have not been investigated previously. Using immunohistochemistry, we determined expression of PEDF in common and dysplastic melanocytic nevi, melanoma in situ, invasive melanoma, and metastatic melanoma (n = 102). PEDF expression was consistently decreased in invasive and metastatic melanoma, compared with nevi and melanoma in situ (P < 0.0001). PEDF was lost in thicker melanomas (P = 0.003), and correlated with depth of invasion (P = 0.003) and distant metastasis (P = 0.0331), but only marginally with mitotic index, AJCC stage, nodal metastasis, or blood vascular density (0.05 < P < 0.10). Quantitative real-time PCR and microarray analyses confirmed PEDF down-regulation at the mRNA level in several melanoma lines, compared with melanocytes. MITF positively correlated with PEDF expression in invasive melanomas (P = 0.0003). Searching for PEDF regulatory mechanisms revealed two occupied conserved E-boxes (DNA recognition elements) in the first intron of the human and mouse PEDF promoter regions, confirmed by binding assays. Dominant-negative and siRNA approaches in vivo demonstrated direct transcriptional influence of MITF on PEDF, establishing the PEDF gene (SERPINF1) as a MITF target in melanocytes and melanoma cells. These findings suggest that loss of PEDF expression promotes early invasive melanoma growth.

"Designed" grafts for HLA-haploidentical stem cell transplantation.

Today human leukocyte antigen-haploidentical transplantation is a feasible option for patients with high-risk acute leukemia who do not have matched donors. Whether it is T-cell replete or T-cell depleted, it is still, however, associated with issues of transplant-related mortality and posttransplant leukemia relapse. After reports that adoptive immunotherapy with T-regulatory cells controls the alloreactivity of conventional T lymphocytes in animal models, tomorrow's world of haploidentical transplantation will focus on new "designed" grafts. They will contain an appropriate ratio of conventional T lymphocytes and T-regulatory cells, natural killer cells, γ δ T cells, and other accessory cells. Preliminary results of ongoing clinical trials show the approach is feasible. It is associated with better immune reconstitution and a quite powerful graft-versus-leukemia effect with a low incidence of graft-versus-host disease and no need for posttransplant pharmacological prophylaxis. Future strategies will focus on enhancing the clinical benefit of T-regulatory cells by increasing their number and strengthening their function.

HLA-haploidentical transplantation with regulatory and conventional T-cell adoptive immunotherapy prevents acute leukemia relapse.

Posttransplant relapse is still the major cause of treatment failure in high-risk acute leukemia. Attempts to manipulate alloreactive T cells to spare normal cells while killing leukemic cells have been unsuccessful. In HLA-haploidentical transplantation, we reported that donor-derived T regulatory cells (Tregs), coinfused with conventional T cells (Tcons), protected recipients against graft-versus-host disease (GVHD). The present phase 2 study investigated whether Treg-Tcon adoptive immunotherapy prevents posttransplant leukemia relapse. Forty-three adults with high-risk acute leukemia (acute myeloid leukemia 33; acute lymphoblastic leukemia 10) were conditioned with a total body irradiation-based regimen. Grafts included CD34(+) cells (mean 9.7 × 10(6)/kg), Tregs (mean 2.5 × 10(6)/kg), and Tcons (mean 1.1 × 10(6)/kg). No posttransplant immunosuppression was given. Ninety-five percent of patients achieved full-donor type engraftment and 15% developed ≥grade 2 acute GVHD. The probability of disease-free survival was 0.56 at a median follow-up of 46 months. The very low cumulative incidence of relapse (0.05) was significantly better than in historical controls. These results demonstrate the immunosuppressive potential of Tregs can be used to suppress GVHD without loss of the benefits of graft-versus-leukemia (GVL) activity. Humanized murine models provided insights into the mechanisms underlying separation of GVL from GVHD, suggesting the GVL effect is due to largely unopposed Tcon alloantigen recognition in bone marrow.

Healthy lifestyle interventions to combat noncommunicable disease-a novel nonhierarchical connectivity model for key stakeholders: a policy statement from the American Heart Association, European Society of Cardiology, European Association for Cardiovascular Prevention and Rehabilitation, and American College of Preventive Medicine.

Noncommunicable diseases (NCDs) have become the primary health concern for most countries around the world. Currently, more than 36 million people worldwide die from NCDs each year, accounting for 63% of annual global deaths; most are preventable. The global financial burden of NCDs is staggering, with an estimated 2010 global cost of $6.3 trillion (US dollars) that is projected to increase to $13 trillion by 2030. A number of NCDs share one or more common predisposing risk factors, all related to lifestyle to some degree: (1) cigarette smoking, (2) hypertension, (3) hyperglycemia, (4) dyslipidemia, (5) obesity, (6) physical inactivity, and (7) poor nutrition. In large part, prevention, control, or even reversal of the aforementioned modifiable risk factors are realized through leading a healthy lifestyle (HL). The challenge is how to initiate the global change, not toward increasing documentation of the scope of the problem but toward true action-creating, implementing, and sustaining HL initiatives that will result in positive, measurable changes in the previously defined poor health metrics. To achieve this task, a paradigm shift in how we approach NCD prevention and treatment is required. The goal of this American Heart Association/European Society of Cardiology/European Association for Cardiovascular Prevention and Rehabilitation/American College of Preventive Medicine policy statement is to define key stakeholders and highlight their connectivity with respect to HL initiatives. This policy encourages integrated action by all stakeholders to create the needed paradigm shift and achieve broad adoption of HL behaviors on a global scale.