Combinatorial immune checkpoint blockade increases myocardial expression of NLRP-3 and secretion of H-FABP, NT-Pro-BNP, interleukin-1ß and interleukin-6: biochemical implications in cardio-immuno-oncology.

Background: Immune checkpoint blockade in monotherapy or combinatorial regimens with chemotherapy or radiotherapy have become an integral part of oncology in recent years. Monoclonal antibodies against CTLA-4 or PD-1 or PDL-1 are the most studied ICIs in randomized clinical trials, however, more recently, an anti-LAG3 (Lymphocyte activation gene-3) antibody, Relatlimab, has been approved by FDA in combination with Nivolumab for metastatic melanoma therapy. Moreover, Atezolizumab is actually under study in association with Ipilimumab for therapy of metastatic lung cancer. Myocarditis, vasculitis and endothelitis are rarely observed in these patients on monotherapy, however new combination therapies could expose patients to more adverse cardiovascular events. Methods: Human cardiomyocytes co-cultured with human peripheral blood lymphocytes (hPBMCs) were exposed to monotherapy and combinatorial ICIs (PD-L1 and CTLA-4 or PD-1 and LAG-3 blocking agents, at 100 nM) for 48 h. After treatments, cardiac cell lysis and secretion of biomarkers of cardiotoxicity (H-FABP, troponin-T, BNP, NT-Pro-BNP), NLRP3-inflammasome and Interleukin 1 and 6 were determined through colorimetric and enzymatic assays. Mitochondrial functions were studied in cardiomyocyte cell lysates through quantification of intracellular Ca++, ATP content and NADH:ubiquinone oxidoreductase core subunit S1 (Ndufs1) levels. Histone deacetylases type 4 (HDAC-4) protein levels were also determined in cardiomyocyte cell lysates to study potential epigenetic changes induced by immunotherapy regimens. Results: Both combinations of immune checkpoint inhibitors exert more potent cardiotoxic side effects compared to monotherapies against human cardiac cells co-cultured with human lymphocytes. LDH release from cardiac cells was 43% higher in PD-L1/CTLA-4 blocking agents, and 35.7% higher in PD-1/LAG-3 blocking agents compared to monotherapies. HDAC4 and intracellular Ca++ levels were increased, instead ATP content and Ndufs1 were reduced in myocardial cell lysates (p < 0.001 vs. untreated cells). Troponin-T, BNP, NT-Pro-BNP and H-FABP, were also strongly increased in combination therapy compared to monotherapy regimen. NLRP3 expression, IL-6 and IL-1ß levels were also increased by PDL-1/CTLA-4 and PD-1/LAG-3 combined blocking agents compared to untreated cells and monotherapies. Conclusions: Data of the present study, although in vitro, indicate that combinatorial immune checkpoint blockade, induce a pro- inflammatory phenotype, thus indicating that these therapies should be closely monitored by the multidisciplinary team consisting of oncologists, cardiologists and immunologists.

Sodium-glucose cotransporter 2 inhibitor dapagliflozin prevents ejection fraction reduction, reduces myocardial and renal NF-κB expression and systemic pro-inflammatory biomarkers in models of short-term doxorubicin cardiotoxicity.

Background: Anthracycline-mediated adverse cardiovascular events are among the leading causes of morbidity and mortality in patients with cancer. Sodium-glucose cotransporter 2 inhibitors (SGLT2i) exert multiple cardiometabolic benefits in patients with/without type 2 diabetes, chronic kidney disease, and heart failure with reduced and preserved ejection fraction. We hypothesized that the SGLT2i dapagliflozin administered before and during doxorubicin (DOXO) therapy could prevent cardiac dysfunction and reduce pro-inflammatory pathways in preclinical models. Methods: Cardiomyocytes were exposed to DOXO alone or combined with dapagliflozin (DAPA) at 10 and 100 nM for 24 h; cell viability, iATP, and Ca++ were quantified; lipid peroxidation products (malondialdehyde and 4-hydroxy 2-hexenal), NLRP3, MyD88, and cytokines were also analyzed through selective colorimetric and enzyme-linked immunosorbent assay (ELISA) methods. Female C57Bl/6 mice were treated for 10 days with a saline solution or DOXO (2.17 mg/kg), DAPA (10 mg/kg), or DOXO combined with DAPA. Systemic levels of ferroptosis-related biomarkers, galectin-3, high-sensitivity C-reactive protein (hs-CRP), and pro-inflammatory chemokines (IL-1α, IL-1ß, IL-2, IL-4, IL-6, IL-10, IL-12, IL17-α, IL-18, IFN-γ, TNF-α, G-CSF, and GM-CSF) were quantified. After treatments, immunohistochemical staining of myocardial and renal p65/NF-kB was performed. Results: DAPA exerts cytoprotective, antioxidant, and anti-inflammatory properties in human cardiomyocytes exposed to DOXO by reducing iATP and iCa++ levels, lipid peroxidation, NLRP-3, and MyD88 expression. Pro-inflammatory intracellular cytokines were also reduced. In preclinical models, DAPA prevented the reduction of radial and longitudinal strain and ejection fraction after 10 days of treatment with DOXO. A reduced myocardial expression of NLRP-3 and MyD-88 was seen in the DOXO-DAPA group compared to DOXO mice. Systemic levels of IL-1ß, IL-6, TNF-α, G-CSF, and GM-CSF were significantly reduced after treatment with DAPA. Serum levels of galectine-3 and hs-CRP were strongly enhanced in the DOXO group; on the other hand, their expression was reduced in the DAPA-DOXO group. Troponin-T, B-type natriuretic peptide (BNP), and N-Terminal Pro-BNP (NT-pro-BNP) were strongly reduced in the DOXO-DAPA group, revealing cardioprotective properties of SGLT2i. Mice treated with DOXO and DAPA exhibited reduced myocardial and renal NF-kB expression. Conclusion: The overall picture of the study encourages the use of DAPA in the primary prevention of cardiomyopathies induced by anthracyclines in patients with cancer.

Clausius inequality and optimality of quasistatic transformations for nonequilibrium stationary states.

Nonequilibrium stationary states of thermodynamic systems dissipate a positive amount of energy per unit of time. If we consider transformations of such states that are realized by letting the driving depend on time, the amount of energy dissipated in an unbounded time window then becomes infinite. Following the general proposal by Oono and Paniconi and using results of the macroscopic fluctuation theory, we give a natural definition of a renormalized work performed along any given transformation. We then show that the renormalized work satisfies a Clausius inequality and prove that equality is achieved for very slow transformations, that is, in the quasistatic limit. We finally connect the renormalized work to the quasipotential of the macroscopic fluctuation theory, which gives the probability of fluctuations in the stationary nonequilibrium ensemble.

Starting dose and dose adjustment of non-vitamin K antagonist oral anticoagulation agents in a nationwide cohort of patients with atrial fibrillation.

This study aims to provide real-world data about starting-dose of NOACs and dose-adjustment in patients with atrial fibrillation (AF). In fact, even if new oral anticoagulation agents (NOACs) have a predictable effect without need for regular monitoring, dose-adjustments should be performed according to the summary of product information and international guidelines. We employed the Italian Medicines Agency monitoring registries comprising data on a nationwide cohort of patients with AF treated with NOACs from 2013 to 2018. Logistic regression analysis was used to evaluate the determinants of dosage choice. During the reference period, treatment was commenced for 866,539 patients. Forty-five percent of the first prescriptions were dispensed at a reduced dose (dabigatran 60.3%, edoxaban 45.2%, apixaban 40.9%, rivaroxaban 37.4%). The prescription of reduced dose was associated with older age, renal disease, bleeding risk and the concomitant use of drugs predisposing to bleeding, but not with CHA2DS2-VASc and HAS-BLED. A relative reduction of the proportion of patients treated with low dosages was evident overtime for dabigatran and rivaroxaban; whereas prescription of low dose apixaban and edoxaban increased progressively among elderly patients. Evidence based on real-world data shows a high frequency of low dose prescriptions of NOACs in AF patients. Except for older age, renal disease, bleeding risk and the concomitant use of drugs predisposing to bleeding, other factors that may determine the choice of reduced dose could not be ascertained. There may be potential under-treatment of AF patients, but further evaluation is warranted.

Non-vitamin K antagonist oral anticoagulation agents in patients with atrial fibrillation: Insights from Italian monitoring registries.

BACKGROUND: Atrial fibrillation (AF) is the most common cardiac arrhythmia associated with an increased risk of stroke and thromboembolism. Anticoagulation with Vitamin K antagonists (VKAs) or with novel oral anti-coagulants (NOACs) represents the cornerstone of the pharmacological treatment to reduce the risk of thromboembolism. This study aims to provide real-world data from a whole large European country about NOAC use in "non-valvular atrial fibrillation" (NVAF). METHODS: We analysed the Italian Medicines Agency (AIFA) monitoring registries collecting data of a nationwide cohort of patients with "NVAF" treated with NOACs. Using logistic regression analysis, baseline characteristics and treatment discontinuation information were compared among initiators of the 4 NOACs. RESULTS: In the reference period, the NOAC database collected data for 683,172 patients. The median age was 78 years with 19.5% aged 85 or older. Overall, the treatments were in accordance with guidelines. About 1/3 of patients switched from a prior VKA treatment; in the 72.3% of cases, these patients had a labile International Normalized Ratio (INR) at first prescription. The most prescribed NOAC was rivaroxaban, followed by apixaban, dabigatran and edoxaban. CONCLUSIONS: This study is the largest European real-world study ever published on NOACs. It includes all Italian patients treated with NOACs since 2013 accounting for about 1/3 of subjects with AF. The enrolled population consisted of very elderly patients, at high risk of ischemic adverse events. The AIFA registries are consolidated tools that guarantee the appropriateness of prescription and provide important information for the governance of National Health System by collecting real-world data.

Renal transplantation in patients over 65 years of age: no more a contraindication but a growing indication.

INTRODUCTION: Worldwide organ shortage and the increasing age of end-stage renal disease patients demanding a graft have prompted extensive use of marginal donors. The "old-for-old" allocation has been proposed for the elderly. The aim of this study was to evaluate the results of a policy of free acceptance into the waiting list of recipients older than 65 years. METHODS: From 1987 to 2004 70 patients whose mean age was 67.4 +/- 2.8 years, underwent an extensive pretransplant evaluation including cardiac studies. Immunosuppression was based upon low-dose steroids, and cyclosporine (50%) or tacrolimus (44%). RESULTS: Patient and graft survival at 1, 3, 5, and 10 years were 85%, 78.5%, 75%, 50%, and 80%, 74%, 70%, 36%, respectively. Death occurred in 17/70 (24%), 14 of whom had a functioning graft. The causes of death were 30% cancer, 23% cardiovascular, 23% sepsis, 12% cerebrovascular hemorrhage, 12% meningitis. The acute rejection (AR) rate was 18.6%. The causes of graft loss were: 71% patient death, 4% irreversible AR, 4% vascular thrombosis, and 21% chronic allograft dysfunction. The main complications were: 52% prostatic hypertrophy, 40% urinary tract infections, 8.6% diabetes, 11% pneumonia, 10% cardiovascular diseases, 7% urological complications, 8% abdominal pathology, 6% acute pyelonephritis, 8% non-skin cancer. CONCLUSIONS: Despite the increased vulnerability of the elderly, they should not be excluded a priori from renal transplantation. Extensive pretransplant screening, mainly cardiovascular, and a tailored immunosuppression are two crucial issues. The moderate rate of AR suggests that these patients do not have an impaired immunocompetence as far as acute rejection is concerned.

[Ethical problems in dialysis in elderly patients]. / L'etica nel paziente anziano in dialisi. Un'interpretazione del Centro Dialisi di Aosta.

The incidence and prevalence of elderly patients are progressively increasing in most dialysis facilities with consequent medical assistance difficulties and the need to find a suitable care unit. Particularly in this age group, the clinical assessment is often difficult and the selection criteria vary widely, not only from country to country, but also from one dialysis unit to another unit in the same area. The authors discuss some of the more complex arguments for and against dialysis in elderly patients. For some of the more difficult cases, Kantian deontology and its three ethical principles of beneficence, non-maleficence and autonomy can facilitate the decision-making process regarding the acceptance or refusal of the therapy. The central role of the patients themselves, the involvement of the family, the discussion of the individual case within the dialysis team, and the good performance of the pre-dialysis program are particularly important. In certain cases it is possible to actually discontinue the treatment, or not initiate it, by using different arguments in competent or incompetent patients. In conclusion, the difficult choice of whether to treat or turn down an elderly patient must depend exclusively on the medical clinical assessment of each case, whereas economical considerations can incorrectly influence that choice.

[Nephrologists and porphyrias]. / I nefrologi e le porfirie.

As usually occurs for rare diseases, the word "PORPHYRIA" often recalls a confused topic with shaded boundaries, presenting "bullous" skin lesions, rare opportunity of diagnosis in clinical practice, unknown pathogenesis, and almost absent therapeutic options. The goal of this review is to draw attention to this topic, as new diagnostic and therapeutic tools might change the natural history of this disease. Porphyrias are disorders resulting from abnormalities of porphyrin metabolism. Porphyrins are molecules made up of four pyrrol rings, which constitute haeme-proteins, including haemoglobin. Following the "trigger" enzyme delta-aminolevulinic acid (ALA) synthase, which is capable of condensing succynil CoA and glycine, seven additional enzymes are involved in the process that eventually leads to haeme biosynthesis. Porphyrias are the result of total or partial deficiencies in these seven enzymes involved in haeme synthesis. Usually, the final haeme product exerts a negative feed-back on its synthesis. The enzyme deficiency that occurs in porphyrias is responsible for reduced haeme production, which, in turn, allows the cascade to be stimulated by increased activity of the trigger enzyme, ALA-synthase (ALA-s). However, due to the subsequent enzyme defects, notwithstanding increased ALA-s activity, haeme synthesis is blunted and intermediate metabolites accumulate. Clinical manifestations depend on which step the enzymatic defect occurs: if enzymatic defects are in the initial steps of the metabolic cascade, early metabolic intermediates will accumulate [i.e. ALA and porphobilinogen (PBG)] responsible for attacks of neurological dysfunction; if the enzymatic defects are in the final steps, sunlight-induced cutaneous lesions (phtotosensitivity) due to porphyrin accumulation in the skin will develop. The seven major human porphyrias may be classified into "hepatic or erythropoietic porphyrias" depending on the organ/tissue where metabolic alterations are more evident, or "acute or chronic porphyrias" depending on the prevalence of clinical symptoms, if neurologic (acute) or cutaneous (chronic). Only a small number of people with inherited enzyme deficiency will develop overt clinical disease, mainly because of the role of acquired aggravating and precipitating factors, such as drugs, hormonal causes, infection, caloric restriction, alcohol. The biochemical diagnosis of porphyrias relies on the detection of the consequences of increased ALA-s activity in the liver: overproduction, accumulation and increased excretion of early (ALA, PBG) or late (porphyrins) intermediate compounds in plasma, faeces and urine. A major difficulty arises from the knowledge that such abnormalities may be completely absent during the remission phases of the disease. Only in very specialised Centres it is now possible to measure specific haeme synthesis enzyme defects, and to perform molecular diagnosis by DNA analysis. The true prevalence of the diseases is unknown, ranging from 1:500 to 1:50,000. Therapeutic strategies include withdrawal of all common precipitants (drug, alcohol, fasting, infection), use of opiates and chlorpromazine, carbohydrates (300-400 g/day) and infusion of human haemine. Genetic therapies are being studied for the future.

[Dialysis in the elderly]. / La dialisi nel paziente anziano.

Assuming >/= 75 years old as the age limit to define dialysis in the elderly, the incidence in this group of patients is progressively increasing in most dialysis units, with an annual growth of 8 to 16%, and represents 20 to 33% of the overall population being affected. The prevalence of the elderly dialysis group is also high, 14 to 20%, in the main literature casistics. Vascular nephropathies, 13 to 50%, represent the major cause of end-stage renal disease, followed by diabetes, 11 to 37%. First year survival rate is an acceptable 52 to 82%, whereas the fifth year value is on average 20 %, also due to the high baseline mortality in these patients. The death causes are mainly cardiac related and represent 45% of the overall mortality. The main prognostic factors are frequency and severity of comorbid factors, in addition to nutritional indexes that are particularly important in this age group. Dialysis dose and treatment time are not related to mortality. Haemodialysis and peritoneal dialysis complement each other to allow the best results. The survival rate, however, is usually better with haemodialysis, especially in old diabetic patients and after some years of treatment. Vascular access, intradialytic hypotension, cardiopathy, intestinal bleeding and amyloidotic arthropathy represent the more critical aspects of dialysis in the elderly, while the quality of life is sometimes unexpectedly good.

Current fluctuations in stochastic lattice gases.

We study current fluctuations in lattice gases in the macroscopic limit extending the dynamic approach for density fluctuations developed in previous articles. More precisely, we establish a large deviation theory for the space-time fluctuations of the empirical current which include the previous results. We then estimate the probability of a fluctuation of the average current over a large time interval. It turns out that recent results by Bodineau and Derrida [Phys. Rev. Lett. 92, 180601 (2004)]] in certain cases underestimate this probability due to the occurrence of dynamical phase transitions.

Fluctuations in stationary nonequilibrium states of irreversible processes.

We formulate a dynamical fluctuation theory for stationary nonequilibrium states (SNS) which covers situations in a nonlinear hydrodynamic regime and is verified explicitly in stochastic models of interacting particles. In our theory a crucial role is played by the time reversed dynamics. Our results include the modification of the Onsager-Machlup theory in the SNS, a general Hamilton-Jacobi equation for the macroscopic entropy and a nonequilibrium, nonlinear fluctuation dissipation relation valid for a wide class of systems.

[Characteristics of non-sustained ventricular tachycardia found during dynamic ECG in patients with recent myocardial infarction]. / Caratteristiche delle tachicardie ventricolari non sostenute riscontrate all'ECG dinamico in pazienti con infarto miocardico recente.

The characteristics of ventricular tachycardia found during Holter ECG monitoring before discharge in patients hospitalized because of acute myocardial infarction were analyzed. One or more ventricular tachycardia episodes were found in 29 of 251 patients (11.5%). On the whole, there were 233 episodes of ventricular tachycardia: 18 patients (62%) had only one episode of ventricular tachycardia, 9 (31%) 2-5 episodes and 2, respectively, 68 and 118 episodes. Episodes of ventricular tachycardia were more numerous in patients with frequent or polymorphic premature ventricular complexes than in patients with sporadic or monomorphic premature ventricular complexes. Fifty-seven ventricular tachycardia episodes were analyzed: 30 of 3 beats, 25 of 4-9 beats and 2 of 15 beats. Forty-seven episodes were monomorphic and 10 (17.5%) were polymorphic. The ventricular tachycardia rate was 136.4 +/- 25 b/m' (range 104-200). The RR'/QT ratio (where RR' = coupling interval of the first beat of ventricular tachycardia) was 1.67 +/- 0.42 and was not correlated either with the rate or the number of beats of ventricular tachycardia. Heart rate at the moment of ventricular tachycardia was 82 +/- 15 b/m' and QT interval 0.36 +/- 0.05 sec; there was no difference when compared to their values of 1 and 5 minutes before ventricular tachycardia. Furthermore, the heart rate showed no difference when compared to the mean value of the hours in which ventricular tachycardia episodes occurred. In addition, heart rate was not correlated with ventricular tachycardia rate, whereas a good correlation was found between the last RR interval preceding ventricular tachycardia and RR' interval (r = 0.61, P less than 0.01).

Electromagnetic interference of digital and analog cellular telephones with implantable cardioverter defibrillators: in vitro and in vivo studies.

The present study examines the potential electromagnetic interference effects induced by cellular telephones on ICDs. We developed ad hoc protocols to conduct both in vitro and in vivo trials on most of the implantable cardioverter defibrillators available on the international market. Trials were conducted with three cellular telephones: two GSM (Global System for Mobile Communication) and one TACS (Total Access Communication System). A human trunk simulator was used to carry out in vitro observations on six ICDs from five manufacturers. In vivo tests were conducted on 13 informed patients with eight different ICD models. During the trials in air, GSM telephones induced interference effects on 4 out of the 6 cardioverter defibrillators tested. Specifically, pulse inhibition, reprogramming, false ventricular fibrillation, and ventricular tachycardia detections occurred, which would have entailed inappropriate therapy delivery had this been activated. Effects were circumscribed to the area closely surrounding the connectors. When the ICD was immersed in saline solution, no effects were observed. Three cases of just ventricular triggering with the interfering signal were observed in vivo.

Evaluation of mean right atrial pressure by two-dimensional and Doppler echocardiography in patients with cardiac disease.

BACKGROUND: Several approaches have been used for noninvasive estimation of right atrial pressure (RAP), but, no currently available method has gained any definite validation. The purpose of this study was to evaluate the accuracy of two-dimensional and Doppler echocardiography in estimating mean RAP in patients with cardiac disease. METHODS: We examined the relation of mean RAP to right atrial size and function, size and respiratory changes of inferior vena cava and Doppler parameters of tricuspid and hepatic vein flow in 114 consecutive patients (77 men, 37 women; mean age 57 +/- 12 years) with various cardiac diseases undergoing cardiac catheterization. Echocardiographic studies were performed within 24 hours before catheterization (mean interval 6 +/- 3 hours). Patients were assigned to 3 groups according to the values of mean RAP (group 1, < or = 8 mmHg; group 2, between 9 and 12 mmHg; group 3, > 12 mmHg). RESULTS: Mean RAP ranged from 3 to 20 mmHg (mean 9.1 +/- 4.3 mmHg). It correlated most strongly with the collapsibility index of inferior vena cava (IVCCI) (r = -0.76), minimal inspiratory diameter of inferior vena cava (r = 0.72) and deceleration time of early tricuspid flow (DT) (r = -0.61). Discriminant analysis demonstrated that IVCCI and DT were major determinants of mean RAP with 81.6% of cases correctly assigned to study groups: 96% of patients of group 1 and 87% of patients of group 3 were identified, whereas the accuracy in identifying the patients of group 2 was lower (46%). An IVCCI > 45% was the best cutoff point in predicting a mean RAP < or = 8 mmHg; an IVCCI < 35% and a DT < 150 msec were the best cutoff points in predicting a mean RAP > or = 15 mmHg. The best multivariate equation predicting mean RAP was: mean RAP = 23.3 - 0.2 IVCCI -0.026 DT (r = 0.80, R2 = 0.64). This equation was 81% sensitive and 84% specific in detecting a mean RAP < or = 8 mmHg and 74% sensitive and 97% specific in detecting a mean RAP > 12 mmHg. CONCLUSIONS: Mean RAP can be estimated noninvasively by two-dimensional and Doppler echocardiography. The combined analysis of IVCCI and DT provides an accurate prediction on mean RAP < or = 8 mmHg and > 12 mmHg, whereas the prediction of intermediate values is less accurate.