Adult Multipotent Cardiac Progenitor-Derived Spheroids: A Reproducible Model of In Vitro Cardiomyocyte Commitment and Specification.

BACKGROUND: Three-dimensional cell culture systems hold great promise for bridging the gap between in vitro cell-based model systems and small animal models to study tissue biology and disease. Among 3D cell culture systems, stem-cell-derived spheroids have attracted significant interest as a strategy to better mimic in vivo conditions. Cardiac stem cell/progenitor (CSC)-derived spheroids (CSs) provide a relevant platform for cardiac regeneration. METHODS: We compared three different cell culture scaffold-free systems, (i) ultra-low attachment plates, (ii) hanging drops (both requiring a 2D/3D switch), and (iii) agarose micro-molds (entirely 3D), for CSC-derived CS formation and their cardiomyocyte commitment in vitro. RESULTS: The switch from a 2D to a 3D culture microenvironment per se guides cell plasticity and myogenic differentiation within CS and is necessary for robust cardiomyocyte differentiation. On the contrary, 2D monolayer CSC cultures show a significant reduced cardiomyocyte differentiation potential compared to 3D CS culture. Forced aggregation into spheroids using hanging drop improves CS myogenic differentiation when compared to ultra-low attachment plates. Performing CS formation and myogenic differentiation exclusively in 3D culture using agarose micro-molds maximizes the cardiomyocyte yield. CONCLUSIONS: A 3D culture system instructs CS myogenic differentiation, thus representing a valid model that can be used to study adult cardiac regenerative biology.

A Mouse Model of Dilated Cardiomyopathy Produced by Isoproterenol Acute Exposure Followed by 5-Fluorouracil Administration.

Appropriate dilated cardiomyopathy (DCM) animal models are highly desirable considering the pathophysiological and clinical heterogeneity of DCM. Genetically modified mice are the most widely and intensively utilized research animals for DCM. However, to translate discoveries from basic science into new and personalized medical applications, research in non-genetically based DCM models remains a key issue. Here, we characterized a mouse model of non-ischemic DCM induced by a stepwise pharmacologic regime of Isoproterenol (ISO) high dose bolus followed by a low dose systemic injection of the chemotherapy agent, 5-Fluorouracil (5-FU). C57BL/6J mice were injected with ISO and, 3 days after, were randomly assigned to saline or 5-FU. Echocardiography and a strain analysis show that ISO + 5FU in mice induces progressive left ventricular (LV) dilation and reduced systolic function, along with diastolic dysfunction and a persistent global cardiac contractility depression through 56 days. While mice treated with ISO alone recover anatomically and functionally, ISO + 5-FU causes persistent cardiomyocyte death, ensuing in cardiomyocyte hypertrophy through 56 days. ISO + 5-FU-dependent damage was accompanied by significant myocardial disarray and fibrosis along with exaggerated oxidative stress, tissue inflammation and premature cell senescence accumulation. In conclusions, a combination of ISO + 5FU produces anatomical, histological and functional cardiac alterations typical of DCM, representing a widely available, affordable, and reproducible mouse model of this cardiomyopathy.

COVID-19 Vaccines and Atrial Fibrillation: Analysis of the Post-Marketing Pharmacovigilance European Database.

Atrial fibrillation (AF) has been described in COVID-19 patients. Recently, some case reports and US pharmacovigilance analyses described AF onset as a rare adverse event following COVID-19 vaccination. The possible correlation is unclear. We systematically analyzed the reports of AF related to COVID-19 vaccines collected in the European pharmacovigilance database, EudraVigilance (EV), from 2020 to November 2022. We carried out descriptive and disproportionality analyses. Moreover, we performed a sensitivity analysis, excluding the reports describing other possible alternative AF causes (pericarditis, myocarditis, COVID-19, or other drugs that may cause/exacerbate AF). Overall, we retrieved 6226 reports, which represented only 0.3% of all those related to COVID-19 vaccines collected in EV during our study period. AF reports mainly referred to adults (in particular, >65 years old), with an equal distribution in sex. Reports were mainly related to tozinameran (54.04%), elasomeran (28.3%), and ChAdOx1-S (14.32%). The reported AF required patient hospitalization in 35% of cases and resulted in a life-threatening condition in 10% of cases. The AF duration (when reported) was highly variable, but the majority of the events had a short duration (moda = 24 h). Although an increased frequency of AF reporting with mRNA vaccines emerges from our study, other investigations are required to investigate the possible correlation between COVID-19 vaccination and the rare AF occurrence.

Cytochalasin D restores nuclear size acting on F-actin and IZUMO1 localization in low-quality spermatozoa.

In spermatozoa, the nuclear F-actin supports the acroplaxome, a subacrosomal structure involved in the correct exposure of several acrosomal membrane proteins; among them, the glycoprotein IZUMO1 is the major protein involved in sperm-oocyte fusion. Nuclear F-actin is also involved in sperm head shaping and chromosome compartmentalization. To date, few notions regarding the bivalent role of F-actin on sperm chromatin organization and IZUMO1 positioning have been reported. In our work, we characterized subcellular organization of F-actin in human high- and low-quality spermatozoa (A- and B-SPZ), respectively, showing that F-actin over-expression in sperm head of B-SPZ affected IZUMO1 localization. A correct IZUMO1 repositioning following in vitro induction of F-actin depolymerization, by cytochalasin D treatment, occurred. Interestingly, F-actin depolymerization was also associated with a correct acrosome repositioning, thus to favor a proper acrosome reaction onset, with changes in sperm nuclear size parameters and histone acetylation rate reaching high-quality conditions. In conclusion, the current work shows a key role of F-actin in the control of IZUMO1 localization as well as chromatin remodeling and acetylation events.

COVID-19 and atrial fibrillation: Intercepting lines.

Almost 20% of COVID-19 patients have a history of atrial fibrillation (AF), but also a new-onset AF represents a frequent complication in COVID-19. Clinical evidence demonstrates that COVID-19, by promoting the evolution of a prothrombotic state, increases the susceptibility to arrhythmic events during the infective stages and presumably during post-recovery. AF itself is the most frequent form of arrhythmia and is associated with substantial morbidity and mortality. One of the molecular factors involved in COVID-19-related AF episodes is the angiotensin-converting enzyme (ACE) 2 availability. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) uses ACE2 to enter and infect multiple cells. Atrial ACE2 internalization after binding to SARS-CoV-2 results in a raise of angiotensin (Ang) II, and in a suppression of cardioprotective Ang(1-7) formation, and thereby promoting cardiac hypertrophy, fibrosis and oxidative stress. Furthermore, several pharmacological agents used in COVID-19 patients may have a higher risk of inducing electrophysiological changes and cardiac dysfunction. Azithromycin, lopinavir/ritonavir, ibrutinib, and remdesivir, used in the treatment of COVID-19, may predispose to an increased risk of cardiac arrhythmia. In this review, putative mechanisms involved in COVID-19-related AF episodes and the cardiovascular safety profile of drugs used for the treatment of COVID-19 are summarized.

Dapagliflozin protects the kidney in a non-diabetic model of cardiorenal syndrome.

Cardiorenal syndrome encompasses a spectrum of disorders involving heart and kidney dysfunction, and sharing common risk factors, such as hypertension and diabetes. Clinical studies have shown that patients with and without diabetes may benefit from using sodium-glucose cotransporter 2 inhibitors to reduce the risk of heart failure and ameliorate renal endpoints. Because the underlying mechanisms remain elusive, we investigated the effects of dapagliflozin on the progression of renal damage, using a model of non-diabetic cardiorenal disease. Dahl salt-sensitive rats were fed a high-salt diet for five weeks and then randomized to dapagliflozin or vehicle for the following six weeks. After treatment with dapagliflozin, renal function resulted ameliorated as shown by decrease of albuminuria and urine albumin-to-creatinine ratio. Functional benefit was accompanied by a decreased accumulation of extracellular matrix and a reduced number of sclerotic glomeruli. Dapagliflozin significantly reduced expression of inflammatory and endothelial activation markers such as NF-κB and e-selectin. Upregulation of pro-oxidant-releasing NADPH oxidases 2 and 4 as well as downregulation of antioxidant enzymes were also counteracted by drug treatment. Our findings also evidenced the modulation of both classic and non-classic renin-angiotensin-aldosterone system (RAAS), and effects of dapagliflozin on gene expression of ion channels/transporters involved in renal homeostasis. Thus, in a non-diabetic model of cardiorenal syndrome, dapagliflozin provides renal protection by modulating inflammatory response, endothelial activation, fibrosis, oxidative stress, local RAAS and ion channels.

Streptozotocin-Induced Type 1 and 2 Diabetes Mellitus Mouse Models Show Different Functional, Cellular and Molecular Patterns of Diabetic Cardiomyopathy.

The main cause of morbidity and mortality in diabetes mellitus (DM) is cardiovascular complications. Diabetic cardiomyopathy (DCM) remains incompletely understood. Animal models have been crucial in exploring DCM pathophysiology while identifying potential therapeutic targets. Streptozotocin (STZ) has been widely used to produce experimental models of both type 1 and type 2 DM (T1DM and T2DM). Here, we compared these two models for their effects on cardiac structure, function and transcriptome. Different doses of STZ and diet chows were used to generate T1DM and T2DM in C57BL/6J mice. Normal euglycemic and nonobese sex- and age-matched mice served as controls (CTRL). Immunohistochemistry, RT-PCR and RNA-seq were employed to compare hearts from the three animal groups. STZ-induced T1DM and T2DM affected left ventricular function and myocardial performance differently. T1DM displayed exaggerated apoptotic cardiomyocyte (CM) death and reactive hypertrophy and fibrosis, along with increased cardiac oxidative stress, CM DNA damage and senescence, when compared to T2DM in mice. T1DM and T2DM affected the whole cardiac transcriptome differently. In conclusion, the STZ-induced T1DM and T2DM mouse models showed significant differences in cardiac remodeling, function and the whole transcriptome. These differences could be of key relevance when choosing an animal model to study specific features of DCM.

Initial Phase of Anthracycline Cardiotoxicity Involves Cardiac Fibroblasts Activation and Metabolic Switch.

The application of doxorubicin (DOX) is hampered by cardiotoxicity, with diastolic dysfunction as the earliest manifestation. Fibrosis leads to impaired relaxation, but the mechanisms that operate shortly after DOX exposure are not clear. We asked whether the activation of cardiac fibroblasts (CFs) anticipates myocardial dysfunction and evaluated the effects of DOX on CF metabolism. CFs were isolated from the hearts of rats after the first injection of DOX. In another experiment, CFs were exposed to DOX in vitro. Cell phenotype and metabolism were determined. Early effects of DOX consisted of diastolic dysfunction and unchanged ejection fraction. Markers of pro-fibrotic remodeling and evidence of CF transformation were present immediately after treatment completion. Oxygen consumption rate and extracellular acidification revealed an increased metabolic activity of CFs and a switch to glycolytic energy production. These effects were consistent in CFs isolated from the hearts of DOX-treated animals and in naïve CFs exposed to DOX in vitro. The metabolic switch was paralleled with the phenotype change of CFs that upregulated markers of myofibroblast differentiation and the activation of pro-fibrotic signaling. In conclusion, the metabolic switch and activation of CFs anticipate DOX-induced damage and represent a novel target in the early phase of anthracycline cardiomyopathy.

Safety profile of sodium glucose co-transporter 2 (SGLT2) inhibitors: A brief summary.

A new therapeutic class of oral agents firstly used for the treatment of type 2 diabetes mellitus is represented by gliflozines or sodium-glucose co-transporter 2 (SGLT2) inhibitors. SGLT2 inhibitors might be effective alone or in combination with any other drugs. This therapeutic class currently includes five agents: canagliflozin, dapagliflozin, empagliflozin, ertugliflozin, and sotagliflozin. SGLT2 inhibitors prevent the renal reabsorption of filtered glucose and sodium by blocking the SGLT2 co-transporters in the proximal convoluted renal tubule, facilitating glucose excretion in the urine (glycosuria) and lowering blood glucose levels. SGLT2 inhibitors have also shown to have pleiotropic effects and determine cardiovascular and renal prevention, thus leading to an extension of their therapeutic indication to include the heart failure. Despite their clinical benefits, warnings about adverse events have been implemented by Regulatory Agencies in the product's information since their introduction to the market. In particular, SGLT2 inhibitors have shown a strong impact on a high number of risk factors. They can cause hypoglycaemia, hypotension, lower limb amputation, fractures, genito-urinary infections, and diabetic ketoacidosis with different frequencies of onset. Despite some of these events are rare, they can lead to serious and dangerous complications, highlighting the importance of a strict monitoring of patients. Overall, SLGT-2 inhibitors are effective antidiabetic drugs with favorable advantages in renal and cardiovascular protection, and with a generally well-tolerated safety profile. This review aims to summarize the safety profile of SGLT2 inhibitors available in the market.

Pharmacological clearance of senescent cells improves cardiac remodeling and function after myocardial infarction in female aged mice.

Cardiovascular diseases (CVD) are predominantly an aging disease. Important sex-specific differences exist and the mechanism(s) by which this sex-by-age interaction influences CVD development and progression remains elusive. Accordingly, it is still unknown whether cell senescence, a main feature of cardiac male aging, is a significant feature also of the female aged mouse heart and whether senolytics, senescence-clearing compounds, promote myocardial repair and regeneration after myocardial infarction (MI) in aged female mice. To this aim, the combination of two senolytics, dasatinib and quercetin (D+Q) or just their vehicle was administered to 22-24 months old C57BL/6 female mice after MI. D+Q improved global left ventricle function and myocardial performance after MI whereby female cardiac aging is characterized by accumulation of cardiac senescent cells that are further increased by MI. Despite their terminal differentiation nature, also cardiomyocytes acquire a senescent phenotype with age in females. D+Q removed senescent cardiac non-myocyte and myocyte cells ameliorating cardiac remodeling and regeneration. Senolytics removed aged dysfunctional cardiac stem/progenitor cells (CSCs), relieving healthy CSCs with normal proliferative and cardiomyogenic differentiation potential. In conclusions, cardiac senescent cells accumulate in the aged female hearts. Removing senescent cells is a key therapeutic target for efficient repair of the aged female heart.

Anti-inflammatory role of SGLT2 inhibitors as part of their anti-atherosclerotic activity: Data from basic science and clinical trials.

Atherosclerosis is a progressive inflammatory disease leading to mortality and morbidity in the civilized world. Atherosclerosis manifests as an accumulation of plaques in the intimal layer of the arterial wall that, by its subsequent erosion or rupture, triggers cardiovascular diseases. Diabetes mellitus is a well-known risk factor for atherosclerosis. Indeed, Type 2 diabetes mellitus patients have an increased risk of atherosclerosis and its associated-cardiovascular complications than non-diabetic patients. Sodium-glucose co-transport 2 inhibitors (SGLT2i), a novel anti-diabetic drugs, have a surprising advantage in cardiovascular effects, such as reducing cardiovascular death in a patient with or without diabetes. Numerous studies have shown that atherosclerosis is due to a significant inflammatory burden and that SGLT2i may play a role in inflammation. In fact, several experiment results have demonstrated that SGLT2i, with suppression of inflammatory mechanism, slows the progression of atherosclerosis. Therefore, SGLT2i may have a double benefit in terms of glycemic control and control of the atherosclerotic process at a myocardial and vascular level. This review elaborates on the anti-inflammatory effects of sodium-glucose co-transporter 2 inhibitors on atherosclerosis.

Ranolazine Counteracts Strength Impairment and Oxidative Stress in Aged Sarcopenic Mice.

Sarcopenia is defined as the loss of muscle mass associated with reduced strength leading to poor quality of life in elderly people. The decline of skeletal muscle performance is characterized by bioenergetic impairment and severe oxidative stress, and does not always strictly correlate with muscle mass loss. We chose to investigate the ability of the metabolic modulator Ranolazine to counteract skeletal muscle dysfunctions that occur with aging. For this purpose, we treated aged C57BL/6 mice with Ranolazine/vehicle for 14 days and collected the tibialis anterior and gastrocnemius muscles for histological and gene expression analyses, respectively. We found that Ranolazine treatment significantly increased the muscle strength of aged mice. At the histological level, we found an increase in centrally nucleated fibers associated with an up-regulation of genes encoding MyoD, Periostin and Osteopontin, thus suggesting a remodeling of the muscle even in the absence of physical exercise. Notably, these beneficial effects of Ranolazine were also accompanied by an up-regulation of antioxidant and mitochondrial genes as well as of NADH-dehydrogenase activity, together with a more efficient protection from oxidative damage in the skeletal muscle. These data indicate that the protection of muscle from oxidative stress by Ranolazine might represent a valuable approach to increase skeletal muscle strength in elderly populations.

Myocardial regeneration protocols towards the routine clinical scenario: An unseemly path from bench to bedside.

Heart failure secondary to cardiomyocyte loss and/or dysfunction is the number one killer worldwide. The field of myocardial regeneration with its far-reaching primary goal of cardiac remuscularization and its hard-to-accomplish translation from bench to bedside, has been filled with ups and downs, steps forward and steps backward, controversies galore and, unfortunately, scientific scandals. Despite the present morass in which cardiac remuscularization is stuck in, the search for clinically effective regenerative approaches remains keenly active. Starting with a concise overview of the still highly debated regenerative capacity of the adult mammalian heart, we focus on the main interventions, that have reached or are close to clinical use, critically discussing key findings, successes, and failures. Finally, some promising and innovative approaches for myocardial repair/regeneration still at the pre-clinical stage are discussed to offer a holistic view on the future of myocardial repair/regeneration for the prevention/management of heart failure in the clinical scenario. Funding: This research was funded by Grants from the Ministry of University and Research PRIN2015 2015ZTT5KB_004; PRIN2017NKB2N4_005; PON-AIM - 1829805-2.

Disentangling a Thorny Issue: Myocarditis and Pericarditis Post COVID-19 and Following mRNA COVID-19 Vaccines.

Considering the clinical significance for myocarditis and pericarditis after immunization with mRNA COVID-19 vaccines, the present pharmacovigilance study aimed to describe these events reported with mRNA COVID-19 vaccines in the Vaccine Adverse Events Reporting System (VAERS). From 1990 to July 2021, the mRNA vaccines were the most common suspected vaccines related to suspected cases of myocarditis and/or pericarditis (myocarditis: N = 1,165; 64.0%; pericarditis: N = 743; 55.1%), followed by smallpox vaccines (myocarditis: N = 222; 12.2%; pericarditis: N = 200; 14.8%). We assessed all suspected cases through the case definition and classification of the Brighton Collaboration Group, and only definitive, probable, and possible cases were included in the analysis. Our findings suggested that myocarditis and pericarditis mostly involve young male, especially after the second dose with a brief time to onset. Nevertheless, this risk is lower (0.38/100,000 vaccinated people; 95% CI 0.36-0.40) than the risk of developing myocarditis after SARS-CoV-2 infection (1000-4000 per 100,000 people) and the risk of developing "common" viral myocarditis (1-10 per 100,000 people/year). Comparing with the smallpox vaccine, for which is already well known the association with myocarditis and pericarditis, our analysis showed a lower probability of reporting myocarditis (ROR 0.12, 95% CI 0.10-0.14) and pericarditis (ROR 0.06, 95% CI 0.05-0.08) following immunization with mRNA COVID-19 vaccines.

The Reporting Frequency of Ketoacidosis Events with Dapagliflozin from the European Spontaneous Reporting System: The DAPA-KETO Study.

Dapagliflozin was associated with an increased risk of diabetic ketoacidosis that has led to the European withdrawal of the authorization for the type 1 diabetes. However, it is still used for the treatment of type 2 diabetes. Therefore, we aim to evaluate the occurrence of dapagliflozin-induced ketoacidosis events by using the European spontaneous reporting system. The reporting odds ratios (ROR) were computed to assess the reporting frequency of ketoacidosis events for dapagliflozin compared to Dipeptidyl peptidase-4 (DPP-4) inhibitors, insulins, or all other Sodium-glucose cotransporter-2 (SGLT-2) inhibitors. A total of 2406 cases with dapagliflozin reported at least one event of ketoacidosis. The three most reported events were: diabetic ketoacidosis (1412; 55.39%), ketoacidosis (476; 18.67%), and euglycaemic diabetic ketoacidosis (296; 11.61%). Dapagliflozin was associated with the higher reporting frequency of ketoacidosis events compared to DPP-4 inhibitors (ROR 12.07, 95%CI 11.67-13.81) or insulins (ROR 7.59, 95%CI 7.13-7.89). A lower reporting frequency was instead observed compared to other SGLT2 inhibitors (ROR 0.91, 95%CI 0.87-0.96). Considering the higher reporting frequency of ketoacidosis observed with dapagliflozin then DPP-4 inhibitors or insulins, attention should be given to patients treated with this drug.

Unraveling and Targeting Myocardial Regeneration Deficit in Diabetes.

Cardiomyopathy is a common complication in diabetic patients. Ventricular dysfunction without coronary atherosclerosis and hypertension is driven by hyperglycemia, hyperinsulinemia and impaired insulin signaling. Cardiomyocyte death, hypertrophy, fibrosis, and cell signaling defects underlie cardiomyopathy. Notably, detrimental effects of the diabetic milieu are not limited to cardiomyocytes and vascular cells. The diabetic heart acquires a senescent phenotype and also suffers from altered cellular homeostasis and the insufficient replacement of dying cells. Chronic inflammation, oxidative stress, and metabolic dysregulation damage the population of endogenous cardiac stem cells, which contribute to myocardial cell turnover and repair after injury. Therefore, deficient myocardial repair and the progressive senescence and dysfunction of stem cells in the diabetic heart can represent potential therapeutic targets. While our knowledge of the effects of diabetes on stem cells is growing, several strategies to preserve, activate or restore cardiac stem cell compartments await to be tested in diabetic cardiomyopathy.

Diabetes-Induced Cellular Senescence and Senescence-Associated Secretory Phenotype Impair Cardiac Regeneration and Function Independently of Age.

Diabetes mellitus (DM) affects the biology of multipotent cardiac stem/progenitor cells (CSCs) and adult myocardial regeneration. We assessed the hypothesis that senescence and senescence-associated secretory phenotype (SASP) are main mechanisms of cardiac degenerative defect in DM. Accordingly, we tested whether ablation of senescent CSCs would rescue the cardiac regenerative/reparative defect imposed by DM. We obtained cardiac tissue from nonaged (50- to 64-year-old) patients with type 2 diabetes mellitus (T2DM) and without DM (NDM) and postinfarct cardiomyopathy undergoing cardiac surgery. A higher reactive oxygen species production in T2DM was associated with an increased number of senescent/dysfunctional T2DM-human CSCs (hCSCs) with reduced proliferation, clonogenesis/spherogenesis, and myogenic differentiation versus NDM-hCSCs in vitro. T2DM-hCSCs showed a defined pathologic SASP. A combination of two senolytics, dasatinib (D) and quercetin (Q), cleared senescent T2DM-hCSCs in vitro, restoring their expansion and myogenic differentiation capacities. In a T2DM model in young mice, diabetic status per se (independently of ischemia and age) caused CSC senescence coupled with myocardial pathologic remodeling and cardiac dysfunction. D + Q treatment efficiently eliminated senescent cells, rescuing CSC function, which resulted in functional myocardial repair/regeneration, improving cardiac function in murine DM. In conclusion, DM hampers CSC biology, inhibiting CSCs' regenerative potential through the induction of cellular senescence and SASP independently from aging. Senolytics clear senescence, abrogating the SASP and restoring a fully proliferative/differentiation-competent hCSC pool in T2DM with normalization of cardiac function.

Current and future therapeutic perspective in chronic heart failure.

The incidence of heart failure is primarily flat or declining for a presumably reflecting better management of cardiovascular diseases, but that of heart failure with preserved ejection fraction (HFpEF) is probably increasing for the lack of an established effective treatment. Moreover, there is no specific pharmacological treatment for patients with heart failure with mildly reduced ejection fraction (HFmrEF) since no substantial prospective randomized clinical trial has been performed exclusively in such population. According to the recent 2021 European Society of Cardiology (ESC) guidelines, the triad composed of an Angiotensin Converting Enzyme inhibitor or Angiotensin Receptor-Neprilysin Inhibitor (ARNI), a beta-blocker, and a Mineralcorticoid Receptor Antagonist is the cornerstone therapy for all patients with heart failure with reduced ejection fraction (HFrEF) but a substantial gap exists for patients with HFpEF/HFmrEF. Despite the important role of the Renin-Angiotensin-Aldosterone System (RAAS) in heart failure pathophysiology, RAAS blockers were found ineffective for HFpEF patients. Indeed, even the new drug class of ARNI was found effective only in HFrEF patients. In this regard, a therapeutic alternative may be represented by drug stimulating the non-classic RAAS (ACE2 and A1-7) as well as other emerging drug classes (such as SGLT2 inhibitors). Reflecting on this global health burden and the gap in treatments among heart failure phenotypes, we summarize the leading players of heart failure pathophysiology, the available pharmacological treatments for each heart failure phenotype, and that in future development.

FUS driven circCNOT6L biogenesis in mouse and human spermatozoa supports zygote development.

Circular RNA (circRNA) biogenesis requires a backsplicing reaction, promoted by inverted repeats in cis-flanking sequences and trans factors, such as RNA-binding proteins (RBPs). Among these, FUS plays a key role. During spermatogenesis and sperm maturation along the epididymis such a molecular mechanism has been poorly explored. With this in mind, we chose circCNOT6L as a study case and wild-type (WT) as well as cannabinoid receptor type-1 knock-out (Cb1-/-) male mice as animal models to analyze backsplicing mechanisms. Our results suggest that spermatozoa (SPZ) have an endogenous skill to circularize mRNAs, choosing FUS as modulator of backsplicing and under CB1 stimulation. A physical interaction between FUS and CNOT6L as well as a cooperation among FUS, RNA Polymerase II (RNApol2) and Quaking (QKI) take place in SPZ. Finally, to gain insight into FUS involvement in circCNOT6L biogenesis, FUS expression was reduced through RNA interference approach. Paternal transmission of FUS and CNOT6L to oocytes during fertilization was then assessed by using murine unfertilized oocytes (NF), one-cell zygotes (F) and murine oocytes undergoing parthenogenetic activation (PA) to exclude a maternal contribution. The role of circCNOT6L as an active regulator of zygote transition toward the 2-cell-like state was suggested using the Embryonic Stem Cell (ESC) system. Intriguingly, human SPZ exactly mirror murine SPZ.