HLA-DR15 Molecules Jointly Shape an Autoreactive T Cell Repertoire in Multiple Sclerosis.

The HLA-DR15 haplotype is the strongest genetic risk factor for multiple sclerosis (MS), but our understanding of how it contributes to MS is limited. Because autoreactive CD4+ T cells and B cells as antigen-presenting cells are involved in MS pathogenesis, we characterized the immunopeptidomes of the two HLA-DR15 allomorphs DR2a and DR2b of human primary B cells and monocytes, thymus, and MS brain tissue. Self-peptides from HLA-DR molecules, particularly from DR2a and DR2b themselves, are abundant on B cells and thymic antigen-presenting cells. Furthermore, we identified autoreactive CD4+ T cell clones that can cross-react with HLA-DR-derived self-peptides (HLA-DR-SPs), peptides from MS-associated foreign agents (Epstein-Barr virus and Akkermansia muciniphila), and autoantigens presented by DR2a and DR2b. Thus, both HLA-DR15 allomorphs jointly shape an autoreactive T cell repertoire by serving as antigen-presenting structures and epitope sources and by presenting the same foreign peptides and autoantigens to autoreactive CD4+ T cells in MS.

A regulatory role for TGF-ß signaling in the establishment and function of the thymic medulla.

Medullary thymic epithelial cells (mTECs) are critical in establishing and maintaining the appropriate microenvironment for negative selection and maturation of immunocompetent T cells with a self-tolerant T cell antigen receptor repertoire. Cues that direct proliferation and maturation of mTECs are provided by members of the tumor necrosis factor (TNF) superfamily expressed on developing thymocytes. Here we demonstrate a negative role of the morphogen TGF-ß in tempering these signals under physiological conditions, limiting both growth and function of the thymic medulla. Eliminating TGF-ß signaling specifically in TECs or by pharmacological means increased the size of the mTEC compartment, enhanced negative selection and functional maturation of medullary thymocytes as well as the production of regulatory T cells, thus reducing the autoreactive potential of peripheral T cells.

Reducing Mortality and Morbidity in Children with Severe Combined Immunodeficiency in Switzerland: the Role of Newborn Screening.

Newborn screening (NBS) for severe combined immunodeficiency (SCID) has been introduced in various countries with the aim of reducing morbidity and mortality. However, studies analyzing outcomes before and after the implementation of NBS programs remain limited. This study sought to compare the outcomes of SCID patients identified through Switzerland's national SCID NBS program, introduced in January 2019, with those of a historical cohort diagnosed between 2007 and 2019. The study included seven patients (32%) identified through NBS, and 15 (68%) born before NBS implementation and diagnosed based on clinical signs. Children in the NBS group were younger at diagnosis (median age 9 days vs 9 months, P = .002) and at hematopoietic stem cell transplantation (HSCT, median age 5 months vs 11 months, P = .003) compared to the clinical group. The NBS group had a lower incidence of infections before HSCT (29% vs 93%, P = .004). Although not statistically significant, the overall survival rate on last follow-up was higher in the NBS group (86% vs 67%, P = .62). Importantly, patients with active infections undergoing HSCT had a significantly lower overall survival probability compared to those without (P = .01). In conclusion, the introduction of NBS in Switzerland has led to earlier and often asymptomatic diagnosis of affected children, enabling timely intervention, infection prevention, and prompt treatment. These factors have contributed to higher survival rates in the NBS group. These findings underscore the critical importance of NBS for SCID, offering potential life-saving benefits through early detection and intervention.

Renal Denervation Prevents Atrial Arrhythmogenic Substrate Development in CKD.

BACKGROUND: In patients with chronic kidney disease (CKD), atrial fibrillation (AF) is highly prevalent and represents a major risk factor for stroke and death. CKD is associated with atrial proarrhythmic remodeling and activation of the sympathetic nervous system. Whether reduction of the sympathetic nerve activity by renal denervation (RDN) inhibits AF vulnerability in CKD is unknown. METHODS: Left atrial (LA) fibrosis was analyzed in samples from patients with AF and concomitant CKD (estimated glomerular filtration rate [eGFR], <60 mL/min per 1.73 m2) using picrosirius red and compared with AF patients without CKD and patients with sinus rhythm with and without CKD. In a translational approach, male Sprague Dawley rats were fed with 0.25% adenine (AD)-containing chow for 16 weeks to induce CKD. At week 5, AD-fed rats underwent RDN or sham operation (AD). Rats on normal chow served as control. After 16 weeks, cardiac function and AF susceptibility were assessed by echocardiography, radiotelemetry, electrophysiological mapping, and burst stimulation, respectively. LA tissue was histologically analyzed for sympathetic innervation using tyrosine hydroxylase staining, and LA fibrosis was determined using picrosirius red. RESULTS: Sirius red staining demonstrated significantly increased LA fibrosis in patients with AF+CKD compared with AF without CKD or sinus rhythm. In rats, AD demonstrated LA structural changes with enhanced sympathetic innervation compared with control. In AD, LA enlargement was associated with prolonged duration of induced AF episodes, impaired LA conduction latency, and increased absolute conduction inhomogeneity. RDN treatment improved LA remodeling and reduced LA diameter compared with sham-operated AD. Furthermore, RDN decreased AF susceptibility and ameliorated LA conduction latency and absolute conduction inhomogeneity, independent of blood pressure reduction and renal function. CONCLUSIONS: In an experimental rat model of CKD, RDN inhibited progression of atrial structural and electrophysiological remodeling. Therefore, RDN represents a potential therapeutic tool to reduce the risk of AF in CKD, independent of changes in renal function and blood pressure.

Good engraftment after reduced intensity targeted busulfan-based conditioning and matched related donor hematopoietic cell transplantation in hemoglobinopathies.

BACKGROUND: Hematopoietic cell transplantation (HCT) is an established curative therapy for transfusion-dependent thalassemia (TDT) and sickle cell disease (SCD). The latest American Society of Hematology guidelines recommend myeloablative preparative regimen in patients under 18 years of age. PROCEDURE: The objective was to demonstrate safety and efficacy of a reduced intensity conditioning (RIC) regimen including high-dose fludarabine, anti-thymocyte globulin, and targeted busulfan as a single alkylator to sub-myeloablative exposures. RESULTS: Between 2012 and 2021, 11 patients with SCD and five patients with TDT and matched related donor (MRD) HCT were included. The median age at transplantation was 8.3 years (range: 3.7-18.8 years). The median administered busulfan AUC was 67.4 mg/L×h (range: 60.7-80 mg/L×h). Overall survival was 93.8% and event-free survival 87.5% with one engrafted SCD patient with pre-existing moyamoya disease succumbing after drainage of a subdural hematoma. One SCD patient developed a secondary graft failure and was treated with a second HCT. Myeloid chimerism was full in all other patients with a median follow-up time of 4.1 years (range: 2.0-11.1 years), whereas T-cell donor chimerism was frequently mixed. CONCLUSION: This RIC conditioning followed by MRD HCT is sufficiently myeloablative to cure pediatric patients with hemoglobinopathies without the need for additional total body irradiation or thiotepa.

Semiautomated pipeline for quantitative analysis of heart histopathology.

BACKGROUND: Heart diseases are among the leading causes of death worldwide, many of which lead to pathological cardiomyocyte hypertrophy and capillary rarefaction in both patients and animal models, the quantification of which is both technically challenging and highly time-consuming. Here we developed a semiautomated pipeline for quantification of the size of cardiomyocytes and capillary density in cardiac histology, termed HeartJ, by generating macros in ImageJ, a broadly used, open-source, Java-based software. METHODS: We have used modified Gomori silver staining, which is easy to perform and digitize in high throughput, or Fluorescein-labeled lectin staining. The latter can be easily combined with other stainings, allowing additional quantitative analysis on the same section, e.g., the size of cardiomyocyte nuclei, capillary density, or single-cardiomyocyte protein expression. We validated the pipeline in a mouse model of cardiac hypertrophy induced by transverse aortic constriction, and in autopsy samples of patients with and without aortic stenosis. RESULTS: In both animals and humans, HeartJ-based histology quantification revealed significant hypertrophy of cardiomyocytes reflecting other parameters of hypertrophy and rarefaction of microvasculature and enabling the analysis of protein expression in individual cardiomyocytes. The analysis also revealed that murine and human cardiomyocytes had similar diameters in health and extent of hypertrophy in disease confirming the translatability of our murine cardiac hypertrophy model. HeartJ enables a rapid analysis that would not be feasible by manual methods. The pipeline has little hardware requirements and is freely available. CONCLUSIONS: In summary, our analysis pipeline can facilitate effective and objective quantitative histological analyses in preclinical and clinical heart samples.

Loss of Mitochondrial Ca2+ Uniporter Limits Inotropic Reserve and Provides Trigger and Substrate for Arrhythmias in Barth Syndrome Cardiomyopathy.

BACKGROUND: Barth syndrome (BTHS) is caused by mutations of the gene encoding tafazzin, which catalyzes maturation of mitochondrial cardiolipin and often manifests with systolic dysfunction during early infancy. Beyond the first months of life, BTHS cardiomyopathy typically transitions to a phenotype of diastolic dysfunction with preserved ejection fraction, blunted contractile reserve during exercise, and arrhythmic vulnerability. Previous studies traced BTHS cardiomyopathy to mitochondrial formation of reactive oxygen species (ROS). Because mitochondrial function and ROS formation are regulated by excitation-contraction coupling, integrated analysis of mechano-energetic coupling is required to delineate the pathomechanisms of BTHS cardiomyopathy. METHODS: We analyzed cardiac function and structure in a mouse model with global knockdown of tafazzin (Taz-KD) compared with wild-type littermates. Respiratory chain assembly and function, ROS emission, and Ca2+ uptake were determined in isolated mitochondria. Excitation-contraction coupling was integrated with mitochondrial redox state, ROS, and Ca2+ uptake in isolated, unloaded or preloaded cardiac myocytes, and cardiac hemodynamics analyzed in vivo. RESULTS: Taz-KD mice develop heart failure with preserved ejection fraction (>50%) and age-dependent progression of diastolic dysfunction in the absence of fibrosis. Increased myofilament Ca2+ affinity and slowed cross-bridge cycling caused diastolic dysfunction, in part, compensated by accelerated diastolic Ca2+ decay through preactivated sarcoplasmic reticulum Ca2+-ATPase. Taz deficiency provoked heart-specific loss of mitochondrial Ca2+ uniporter protein that prevented Ca2+-induced activation of the Krebs cycle during ß-adrenergic stimulation, oxidizing pyridine nucleotides and triggering arrhythmias in cardiac myocytes. In vivo, Taz-KD mice displayed prolonged QRS duration as a substrate for arrhythmias, and a lack of inotropic response to ß-adrenergic stimulation. Cellular arrhythmias and QRS prolongation, but not the defective inotropic reserve, were restored by inhibiting Ca2+ export through the mitochondrial Na+/Ca2+ exchanger. All alterations occurred in the absence of excess mitochondrial ROS in vitro or in vivo. CONCLUSIONS: Downregulation of mitochondrial Ca2+ uniporter, increased myofilament Ca2+ affinity, and preactivated sarcoplasmic reticulum Ca2+-ATPase provoke mechano-energetic uncoupling that explains diastolic dysfunction and the lack of inotropic reserve in BTHS cardiomyopathy. Furthermore, defective mitochondrial Ca2+ uptake provides a trigger and a substrate for ventricular arrhythmias. These insights can guide the ongoing search for a cure of this orphaned disease.

Interleukin-1α Is a Central Regulator of Leukocyte-Endothelial Adhesion in Myocardial Infarction and in Chronic Kidney Disease.

BACKGROUND: Cardiovascular diseases and chronic kidney disease (CKD) are highly prevalent, aggravate each other, and account for substantial mortality. Both conditions are characterized by activation of the innate immune system. The alarmin interleukin-1α (IL-1α) is expressed in a variety of cell types promoting (sterile) systemic inflammation. The aim of the present study was to examine the role of IL-1α in mediating inflammation in the setting of acute myocardial infarction (AMI) and CKD. METHODS: We assessed the expression of IL-1α on the surface of monocytes from patients with AMI and patients with CKD and determined its association with atherosclerotic cardiovascular disease events during follow-up in an explorative clinical study. Furthermore, we assessed the inflammatory effects of IL-1α in several organ injury models in Il1a-/- and Il1b-/- mice and investigated the underlying mechanisms in vitro in monocytes and endothelial cells. RESULTS: IL-1α is strongly expressed on the surface of monocytes from patients with AMI and CKD compared with healthy controls. Higher IL-1α surface expression on monocytes from patients with AMI and CKD was associated with a higher risk for atherosclerotic cardiovascular disease events, which underlines the clinical relevance of IL-1α. In mice, IL-1α, but not IL-1ß, mediates leukocyte-endothelial adhesion as determined by intravital microscopy. IL-1α promotes accumulation of macrophages and neutrophils in inflamed tissue in vivo. Furthermore, IL-1α on monocytes stimulates their homing at sites of vascular injury. A variety of stimuli such as free fatty acids or oxalate crystals induce IL-1α surface expression and release by monocytes, which then mediates their adhesion to the endothelium via IL-1 receptor-1. IL-1α also promotes expression of the VCAM-1 (vascular cell adhesion molecule-1) on endothelial cells, thereby fostering the adhesion of circulating leukocytes. IL-1α induces inflammatory injury after experimental AMI, and abrogation of IL-1α prevents the development of CKD in oxalate or adenine-fed mice. CONCLUSIONS: IL-1α represents a key mediator of leukocyte-endothelial adhesion and inflammation in AMI and CKD. Inhibition of IL-1α may serve as a novel anti-inflammatory treatment strategy.

A systematic review and meta-analysis of murine models of uremic cardiomyopathy.

Chronic kidney disease (CKD) triggers the risk of developing uremic cardiomyopathy as characterized by cardiac hypertrophy, fibrosis and functional impairment. Traditionally, animal studies are used to reveal the underlying pathological mechanism, although variable CKD models, mouse strains and readouts may reveal diverse results. Here, we systematically reviewed 88 studies and performed meta-analyses of 52 to support finding suitable animal models for future experimental studies on pathological kidney-heart crosstalk during uremic cardiomyopathy. We compared different mouse strains and the direct effect of CKD on cardiac hypertrophy, fibrosis and cardiac function in "single hit" strategies as well as cardiac effects of kidney injury combined with additional cardiovascular risk factors in "multifactorial hit" strategies. In C57BL/6 mice, CKD was associated with a mild increase in cardiac hypertrophy and fibrosis and marginal systolic dysfunction. Studies revealed high variability in results, especially regarding hypertrophy and systolic function. Cardiac hypertrophy in CKD was more consistently observed in 129/Sv mice, which express two instead of one renin gene and more consistently develop increased blood pressure upon CKD induction. Overall, "multifactorial hit" models more consistently induced cardiac hypertrophy and fibrosis compared to "single hit" kidney injury models. Thus, genetic factors and additional cardiovascular risk factors can "prime" for susceptibility to organ damage, with increased blood pressure, cardiac hypertrophy and early cardiac fibrosis more consistently observed in 129/Sv compared to C57BL/6 strains.

Renal denervation reduces atrial remodeling in hypertensive rats with metabolic syndrome.

Atrial fibrillation (AF) is highly prevalent in hypertensive patients with metabolic syndrome and is related to inflammation and activation of the sympathoadrenergic system. The multi-ligand Receptor-for-Advanced-Glycation-End-products (RAGE) activates inflammation-associated tissue remodeling and is regulated by the sympathetic nervous system. Its counterpart, soluble RAGE (sRAGE), serves as anti-inflammatory decoy receptor with protective properties. We investigated the effect of sympathetic modulation by renal denervation (RDN) on atrial remodeling, RAGE/sRAGE and RAGE ligands in metabolic syndrome. RDN was performed in spontaneously hypertensive obese rats (SHRob) with metabolic syndrome compared with lean spontaneously hypertensive rats (SHR) and with normotensive non-obese control rats. Blood pressure and heart rate were measured by telemetry. The animals were killed 12 weeks after RDN. Left atrial (LA) and right atrial (RA) remodeling was assessed by histological analysis and collagen types. Sympathetic innervation was measured by tyrosine hydroxylase staining of atrial nerve fibers, RAGE/sRAGE, RAGE ligands, cytokine expressions and inflammatory infiltrates were analyzed by Western blot and immunofluorescence staining. LA sympathetic nerve fiber density was higher in SHRob (+44%) versus controls and reduced after RDN (-64% versus SHRob). RAGE was increased (+718%) and sRAGE decreased (- 62%) in SHRob as compared with controls. RDN reduced RAGE expression (- 61% versus SHRob), significantly increased sRAGE levels (+162%) and induced a significant decrease in RAGE ligand levels in SHRob (- 57% CML and - 51% HMGB1) with reduced pro-inflammatory NFkB activation (- 96%), IL-6 production (- 55%) and reduced inflammatory infiltrates. This led to a reduction in atrial fibrosis (- 33%), collagen type I content (- 72%), accompanied by reduced LA myocyte hypertrophy (- 21%). Transfection experiments on H9C2 cardiomyoblasts demonstrated that RAGE is directly involved in fibrosis formation by influencing cellular production of collagen type I. In conclusion, suppression of renal sympathetic nerve activity by RDN prevents atrial remodeling in metabolic syndrome by reducing atrial sympathetic innervation and by modulating RAGE/sRAGE balance and reducing pro-inflammatory and pro-fibrotic RAGE ligands, which provides a potential therapeutic mechanism to reduce the development of AF.

Hematopoietic cell transplantation in chronic granulomatous disease: a study of 712 children and adults.

Chronic granulomatous disease (CGD) is a primary immunodeficiency resulting in life-threatening infections and inflammatory complications. Allogeneic hematopoietic cell transplantation (allo-HCT) can cure the disease, but the indication to transplant remains controversial. We performed a retrospective multicenter study of 712 patients with CGD who underwent allo-HCT transplantation from March 1993 through December 2018. We studied 635 children (aged <18 years) and 77 adults. Median follow-up was 45 months. Median age at transplantation was 7 years (range, 0.1-48.6). Kaplan-Meier estimates of overall survival (OS) and event-free survival (EFS) at 3 years were 85.7% and 75.8%, respectively. In multivariate analysis, older age was associated with reduced survival and increased chronic graft-versus-host disease. Nevertheless, OS and EFS at 3 years for patients ≥18 years were 76% and 69%, respectively. Use of 1-antigen-mismatched donors was associated with reduced OS and EFS . No significant difference was found in OS, but a significantly reduced EFS was noted in the small group of patients who received a transplant from a donor with a >1 antigen mismatch. Choice of conditioning regimen did not influence OS or EFS. In summary, we report an excellent outcome after allo-HCT in CGD, with low incidence of graft failure and mortality in all ages. Older patients and recipients of 1-antigen-mismatched grafts had a less favorable outcome. Transplantation should be strongly considered at a younger age and particularly in the presence of a well-matched donor.

Cathepsin A contributes to left ventricular remodeling by degrading extracellular superoxide dismutase in mice.

In the heart, the serine carboxypeptidase cathepsin A (CatA) is distributed between lysosomes and the extracellular matrix (ECM). CatA-mediated degradation of extracellular peptides may contribute to ECM remodeling and left ventricular (LV) dysfunction. Here, we aimed to evaluate the effects of CatA overexpression on LV remodeling. A proteomic analysis of the secretome of adult mouse cardiac fibroblasts upon digestion by CatA identified the extracellular antioxidant enzyme superoxide dismutase (EC-SOD) as a novel substrate of CatA, which decreased EC-SOD abundance 5-fold. In vitro, both cardiomyocytes and cardiac fibroblasts expressed and secreted CatA protein, and only cardiac fibroblasts expressed and secreted EC-SOD protein. Cardiomyocyte-specific CatA overexpression and increased CatA activity in the LV of transgenic mice (CatA-TG) reduced EC-SOD protein levels by 43%. Loss of EC-SOD-mediated antioxidative activity resulted in significant accumulation of superoxide radicals (WT, 4.54 µmol/mg tissue/min; CatA-TG, 8.62 µmol/mg tissue/min), increased inflammation, myocyte hypertrophy (WT, 19.8 µm; CatA-TG, 21.9 µm), cellular apoptosis, and elevated mRNA expression of hypertrophy-related and profibrotic marker genes, without affecting intracellular detoxifying proteins. In CatA-TG mice, LV interstitial fibrosis formation was enhanced by 19%, and the type I/type III collagen ratio was shifted toward higher abundance of collagen I fibers. Cardiac remodeling in CatA-TG was accompanied by an increased LV weight/body weight ratio and LV end diastolic volume (WT, 50.8 µl; CatA-TG, 61.9 µl). In conclusion, CatA-mediated EC-SOD reduction in the heart contributes to increased oxidative stress, myocyte hypertrophy, ECM remodeling, and inflammation, implicating CatA as a potential therapeutic target to prevent ventricular remodeling.

Fibroblast Growth Factor 21 Response in a Preclinical Alcohol Model of Acute-on-Chronic Liver Injury.

BACKGROUND AND AIMS: Fibroblast growth factor (FGF) 21 has recently been shown to play a potential role in bile acid metabolism. We aimed to investigate the FGF21 response in an ethanol-induced acute-on-chronic liver injury (ACLI) model in Abcb4-/- mice with deficiency of the hepatobiliary phospholipid transporter. METHODS: Total RNA was extracted from wild-type (WT, C57BL/6J) and Abcb4-/- (KO) mice, which were either fed a control diet (WT-Cont and KO-Cont groups; n = 28/group) or ethanol diet, followed by an acute ethanol binge (WT-EtOH and KO-EtOH groups; n = 28/group). A total of 58 human subjects were recruited into the study, including patients with alcohol-associated liver disease (AALD; n = 31) and healthy controls (n = 27). The hepatic and ileal expressions of genes involved in bile acid metabolism, plasma FGF levels, and bile acid and its precursors 7α- and 27-hydroxycholesterol (7α- and 27-OHC) concentrations were determined. Primary mouse hepatocytes were isolated for cell culture experiments. RESULTS: Alcohol feeding significantly induced plasma FGF21 and decreased hepatic Cyp7a1 levels. Hepatic expression levels of Fibroblast growth factor receptor 1 (Fgfr1), Fgfr4, Farnesoid X-activated receptor (Fxr), and Small heterodimer partner (Shp) and plasma FGF15/FGF19 levels did not differ with alcohol challenge. Exogenous FGF21 treatment suppressed Cyp7a1 in a dose-dependent manner in vitro. AALD patients showed markedly higher FGF21 and lower 7α-OHC plasma levels while FGF19 did not differ. CONCLUSIONS: The simultaneous upregulation of FGF21 and downregulation of Cyp7a1 expressions upon chronic plus binge alcohol feeding together with the invariant plasma FGF15 and hepatic Shp and Fxr levels suggest the presence of a direct regulatory mechanism of FGF21 on bile acid homeostasis through inhibition of CYP7A1 by an FGF15-independent pathway in this ACLI model. Lay Summary: Alcohol challenge results in the upregulation of FGF21 and repression of Cyp7a1 expressions while circulating FGF15 and hepatic Shp and Fxr levels remain constant both in healthy and pre-injured livers, suggesting the presence of an alternative FGF15-independent regulatory mechanism of FGF21 on bile acid homeostasis through the inhibition of Cyp7a1.

CaMKII activity contributes to homeometric autoregulation of the heart: A novel mechanism for the Anrep effect.

KEY POINTS: The Anrep effect represents the alteration of left ventricular (LV) contractility to acutely enhanced afterload in a few seconds, thereby preserving stroke volume (SV) at constant preload. As a result of the missing preload stretch in our model, the Anrep effect differs from the slow force response and has a different mechanism. The Anrep effect demonstrated two different phases. First, the sudden increased afterload was momentary equilibrated by the enhanced LV contractility as a result of higher power strokes of strongly-bound myosin cross-bridges. Second, the slightly delayed recovery of SV is perhaps dependent on Ca2+ /calmodulin-dependent protein kinase II activation caused by oxidation and myofilament phosphorylation (cardiac myosin-binding protein-C, myosin light chain 2), maximizing the recruitment of available strongly-bound myosin cross-bridges. Short-lived oxidative stress might present a new facet of subcellular signalling with respect to cardiovascular regulation. Relevance for human physiology was demonstrated by echocardiography disclosing the Anrep effect in humans during handgrip exercise. ABSTRACT: The present study investigated whether oxidative stress and Ca2+ /calmodulin-dependent protein kinase II (CaMKII) activity are involved in triggering the Anrep effect. LV pressure-volume (PV) analyses of isolated, preload controlled working hearts were performed at two afterload levels (60 and 100 mmHg) in C57BL/6N wild-type (WT) and CaMKII-double knockout mice (DKOCaMKII ). In snap-frozen WT hearts, force-pCa relationship, H2 O2 generation, CaMKII oxidation and phosphorylation of myofilament and Ca2+ handling proteins were assessed. Acutely raised afterload showed significantly increased wall stress, H2 O2 generation and LV contractility in the PV diagram with an initial decrease and recovery of stroke volume, whereas end-diastolic pressure and volume, as well as heart rate, remained constant. Afterload induced increase in LV contractility was blunted in DKOCaMKII -hearts. Force development of single WT cardiomyocytes was greater with elevated afterload at submaximal Ca2+ concentration and associated with increases in CaMKII oxidation and phosphorylation of cardiac-myosin binding protein-C, myosin light chain and Ca2+ handling proteins. CaMKII activity is involved in the regulation of the Anrep effect and associates with stimulation of oxidative stress, presumably starting a cascade of CaMKII oxidation with downstream phosphorylation of myofilament and Ca2+ handling proteins. These mechanisms improve LV inotropy and preserve stroke volume within few seconds.

Differential effects of endurance, interval, and resistance training on telomerase activity and telomere length in a randomized, controlled study.

Aims: It is unknown whether different training modalities exert differential cellular effects. Telomeres and telomere-associated proteins play a major role in cellular aging with implications for global health. This prospective training study examines the effects of endurance training, interval training (IT), and resistance training (RT) on telomerase activity and telomere length (TL). Methods and results: One hundred and twenty-four healthy previously inactive individuals completed the 6 months study. Participants were randomized to three different interventions or the control condition (no change in lifestyle): aerobic endurance training (AET, continuous running), high-intensive IT (4 × 4 method), or RT (circle training on 8 devices), each intervention consisting of three 45 min training sessions per week. Maximum oxygen uptake (VO2max) was increased by all three training modalities. Telomerase activity in blood mononuclear cells was up-regulated by two- to three-fold in both endurance exercise groups (AET, IT), but not with RT. In parallel, lymphocyte, granulocyte, and leucocyte TL increased in the endurance-trained groups but not in the RT group. Magnet-activated cell sorting with telomerase repeat-ampliflication protocol (MACS-TRAP) assays revealed that a single bout of endurance training-but not RT-acutely increased telomerase activity in CD14+ and in CD34+ leucocytes. Conclusion: This randomized controlled trial shows that endurance training, IT, and RT protocols induce specific cellular pathways in circulating leucocytes. Endurance training and IT, but not RT, increased telomerase activity and TL which are important for cellular senescence, regenerative capacity, and thus, healthy aging.

Renal Denervation: Is It Ready for Prime Time?

PURPOSE OF REVIEW: Interventional cardiology and in particular the field of renal denervation is subject to constant change. This review provides an up to date overview of renal denervation trials and an outlook on what to expect in the future. RECENT FINDINGS: After the sham-controlled SYMPLICITY HTN-3 trial dampened the euphoria following early renal denervation trials, the recently published results of the sham-controlled SPYRAL HTN and RADIANCE HTN trials provided proof-of-principle for the blood pressure-lowering efficacy of renal denervation. However, these studies underline the major issue of patients' non-adherence to antihypertensive medication as well as the need for reliable patient- and procedure-related predictors of response. The second generation of sham-controlled renal denervation trials provided proof of principle for the blood pressure-lowering efficacy of RDN. However, larger trials have to assess long-term safety and efficacy.

Sympathoadrenergic suppression improves heart function by upregulating the ratio of sRAGE/RAGE in hypertension with metabolic syndrome.

Receptors-for-Advanced-Glycation-End-products (RAGE) activate pro-inflammatory programs mediated by carboxymethyllysine (CML) and high-mobility-group-box1 protein (HMGB1). The soluble isoform sRAGE neutralizes RAGE-ligands preventing cardiovascular complications in conditions associated with increased sympathetic activation like hypertension and diabetes. The effects of sympathetic modulation on RAGE/sRAGE-balance and end-organ damage in metabolic syndrome on top of hypertension remains unknown. We hypothesized that increased sympathoadrenergic activity might lead to an unfavourable RAGE/sRAGE regulation. Renal denervation (RDN) was used to modulate sympathetic activation in obese spontaneously hypertensive rats (SHRobRDN) versus sham-operated obese spontaneously hypertensive rats (SHRob), their hypertensive lean controls (SHR) and non-hypertensive controls. Cardiac fibrosis was assessed by histological analysis and sRAGE/RAGE and ligand levels by Western blotting. Levels of CML and HMGB1 were highest in SHRob and were significantly lowered by RDN in serum (-44% and -45%) and myocardium (-25% and -52%). Myocardial RAGE was increased in SHR (+72% versus controls) and in SHRob (+68% versus SHR) while sRAGE decreased (-50% in SHR versus controls and -51% in SHRob versus SHR). RDN reduced myocardial RAGE expression. (-20%) and increased sRAGE levels in heart (+80%) and serum (+180%) versus sham-operated SHRob. Myocardial fibrosis correlated inversely with myocardial sRAGE content (r = -0.79; p = .004; n = 10). Myocardial sRAGE shedding active A-Disintegrin-And-Metalloprotease-10 (ADAM-10) was decreased in SHR (-33% versus controls) and in SHRob (-54% versus SHR), and was restored after RDN (+129% versus SHRob). Serum ADAM-10 activity was also decreased in SHRob (-66% versus SHR) and restored after RDN (+150% versus SHRob). In vitro, isoproterenol induced a ß1-adrenergic receptor mediated increase of RAGE expression in splenocytes (+200%) and decreased sRAGE secretion of splenocytes and cardiac fibroblasts (-50% and -49%) by ß2-adrenergic receptor stimulation mediated suppression of ADAM-10 activity. In conclusion, sympathetic activity affects sRAGE/RAGE-balance, which can be suppressed through sympathetic modulation by RDN, preventing RAGE-induced cardiac damage in hypertension with metabolic syndrome.

A novel subtilase inhibitor in plants shows structural and functional similarities to protease propeptides.

The propeptides of subtilisin-like serine proteinases (subtilases, SBTs) serve dual functions as intramolecular chaperones that are required for enzyme folding and as inhibitors of the mature proteases. SBT propeptides are homologous to the I9 family of protease inhibitors that have only been described in fungi. Here we report the identification and characterization of subtilisin propeptide-like inhibitor 1 (SPI-1) from Arabidopsis thaliana Sequence similarity and the shared ß-α-ß-ß-α-ß core structure identified SPI-1 as a member of the I9 inhibitor family and as the first independent I9 inhibitor in higher eukaryotes. SPI-1 was characterized as a high-affinity, tight-binding inhibitor of Arabidopsis subtilase SBT4.13 with Kd and Ki values in the picomolar range. SPI-1 acted as a stable inhibitor of SBT4.13 over the physiologically relevant range of pH, and its inhibitory profile included many other SBTs from plants but not bovine chymotrypsin or bacterial subtilisin A. Upon binding to SBT4.13, the C-terminal extension of SPI-1 was proteolytically cleaved. The last four amino acids at the newly formed C terminus of SPI-1 matched both the cleavage specificity of SBT4.13 and the consensus sequence of Arabidopsis SBTs at the junction of the propeptide with the catalytic domain. The data suggest that the C terminus of SPI-1 acts as a competitive inhibitor of target proteases as it remains bound to the active site in a product-like manner. SPI-1 thus resembles SBT propeptides with respect to its mode of protease inhibition. However, in contrast to SBT propeptides, SPI-1 could not substitute as a folding assistant for SBT4.13.

Modulation of renal sympathetic innervation: recent insights beyond blood pressure control.

Renal afferent and efferent sympathetic nerves are involved in the regulation of blood pressure and have a pathophysiological role in hypertension. Additionally, several conditions that frequently coexist with hypertension, such as heart failure, obstructive sleep apnea, atrial fibrillation, renal dysfunction, and metabolic syndrome, demonstrate enhanced sympathetic activity. Renal denervation (RDN) is an approach to reduce renal and whole body sympathetic activation. Experimental models indicate that RDN has the potential to lower blood pressure and prevent cardio-renal remodeling in chronic diseases associated with enhanced sympathetic activation. Studies have shown that RDN can reduce blood pressure in drug-naïve hypertensive patients and in hypertensive patients under drug treatment. Beyond its effects on blood pressure, sympathetic modulation by RDN has been shown to have profound effects on cardiac electrophysiology and cardiac arrhythmogenesis. RDN can display anti-arrhythmic effects in a variety of animal models for atrial fibrillation and ventricular arrhythmias. The first non-randomized studies demonstrate that RDN may promote the maintenance of sinus rhythm following catheter ablation in patients with atrial fibrillation. Registry data point towards a beneficial effect of RDN to prevent ventricular arrhythmias in patients with heart failure and electrical storm. Further large randomized placebo-controlled trials are needed to confirm the antihypertensive and anti-arrhythmic effects of RDN. Here, we will review the current literature on anti-arrhythmic effects of RDN with the focus on atrial fibrillation and ventricular arrhythmias. We will discuss new insights from preclinical and clinical mechanistic studies and possible clinical implications of RDN.

Nocturnal hypoxemic burden is associated with epicardial fat volume in patients with acute myocardial infarction.

BACKGROUND: Increased epicardial fat volume (EFV) is a common feature of patients with sleep-disordered breathing (SDB), is considered as an established marker of cardiovascular risk, and is associated with adverse cardiovascular events after myocardial infarction (MI). METHODS: To investigate the association between different measures of SDB severity and EFV after acute MI, we enrolled 105 patients with acute MI in this study. Unattended in-hospital polysomnography was performed to determine the number of apneas and hypopneas per hour during sleep (apnea-hypopnea index, AHI). To determine nocturnal hypoxemic burden, we used pulse oximetry and applied a novel parameter, the hypoxia load representing the integrated area of desaturation divided by total sleep time (HLTST). Of 105 patients, 56 underwent cardiovascular magnetic resonance to define EFV. RESULTS: HLTST was significantly associated with EFV (r2 = 0.316, p = 0.025). Multivariate linear regression analysis accounting for age, sex, body mass index, smoking, and left ventricular mass demonstrated that the HLTST was an independent modulator of EFV (B-coefficient 0.435 (95% CI 0.021-0.591); p = 0.015). In contrast, AHI or established measures of hypoxemia did not correlate with EFV. CONCLUSIONS: HLTST, a novel parameter to determine nocturnal hypoxemic burden, and not AHI as an event-based measure of SDB, was associated with EFV in patients with acute MI. Further studies are warranted to confirm the link between nocturnal hypoxemia and EFV and to determine the prognostic value of a more detailed characterization of nocturnal hypoxemic burden in patients with high cardiovascular risk.