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Although very common, the precise mechanisms that explain the symptomatology of neuroendocrine syncope (NES) remain poorly understood. This disease, which can be very incapacitating, manifests itself as a drop in blood pressure secondary to vasodilation and/or extreme slowing of heart rate. As studies continue, the involvement of the adenosinergic system is becoming increasingly evident. Adenosine, which is an ATP derivative, may be involved in a large number of cases. Adenosine acts on G protein-coupled receptors with seven transmembrane domains. A1 and A2A adenosine receptor dysfunction seem to be particularly implicated since the activation leads to severe bradycardia or vasodilation, respectively, two cardinal symptoms of NES. This mini-review aims to shed light on the links between dysfunction of the adenosinergic system and NHS. In particular, signal transduction pathways through the modulation of cAMP production and ion channels in relation to effects on the cardiovascular system are addressed. A better understanding of these mechanisms could guide the pharmacological development of new therapeutic approaches. [ABSTRACT FROM AUTHOR]

The inflammasome NLRP3 is involved in the inflammatory processes of several cardiac diseases. Recently, GBP5 and 6 transcripts have been found to be specifically deregulated in immune-related myocarditis, but the mechanism of action remains unclear. Cardiomyocyte dysfunction during the cardiotoxic reaction probably results from the combined action of cytokines secreted by T lymphocytes reactivated by ICIs and inflammatory molecules released by endocardial and vascular endothelial cells. Our hypothesis is that cardiac dysfunction in immune-mediated myocarditis is linked to activation of the NLRP3 inflammasome in cardiac cells in this inflammatory context. Our aim is to unravel the regulatory mechanisms of the NLRP3 inflammasome in response to cytokines known to be secreted during ICI-related myocarditis, and to characterise the role of paracrine effects of endothelial cells in cardiomyocyte dysfunction. hiPSC-derived cardiomyocytes (hiPSC-CM) and endothelial cells (hiPSC-EC) were stimulated by different inflammatory cytokines (IFN-γ, RANTES, Granzyme B and IL-1β). We studied several groups: healthy individual (1 clone from 1 individual), melanoma patients who reported immuno-mediated myocarditis (13 clones from 5 patients), and melanoma patients who did not report cardiotoxicity after immunotherapy (7 clones from 3 patients). IFN-γ affects the viability of cardiac cells by activating the JAK/STAT inflammatory pathway, and by overexpressing NLRP3 inflammasome transcripts such as nlrp3, nlrc5 and caspase-1. This effect is associated to overexpression of the MHC and PD-L1 proteins, as described in humans. IFN-γ alone did not induce significant differences between groups on these targets. We have developed a flow cytometry multiplexing protocol to characterise apoptosis associated with caspase activity, in particular NLRP3 inflammasome-associated caspase-1, metabolic deregulation and oxidative stress. The results are currently being analysed after stimulation by inflammatory cytokines IFN-γ, RANTES, Granzyme B and IL-1β. hiPSC-CM and hiPSC-EC respond to IFN-γ by activating inflammatory pathways, in particular the NLRP3 inflammasome, leading to cell death. Our work unravels the mechanisms associated with cardiotoxicity, triggered by inflammatory cytokines released during the cardiotoxic response to ICIs, thus clarifying the role of the two cardiac cell types in the cardiac dysfunction observed in immuno-mediated myocarditis in patients. [ABSTRACT FROM AUTHOR]

In association with immune checkpoint inhibitors (ICI) anticancer therapy, myocarditis (ICI-M), have emerged with a mortality rate reaching 50%. The IFN-γ pathway and inflammasome regulatory proteins seem to be specifically involved in ICI-M, but the mechanism of action remains unclear. In the cardiovascular system, the endocardial and vascular endothelium is involved in the regulation of the contractile function of cardiomyocytes. Our hypothesis is that the cardiotoxicity induced by the infiltration of immune cells into the myocardium is partly due to an exacerbation of inflammasome activation in endothelial cells, which subsequently affects the cardiomyocytes function. Our aim is to characterize the role of endothelial cells in ICI-related myocarditis in a unique model of induced pluripotent stem cells (hiPSC)-derived cardiac cells. The hiPSC were generated from melanoma patients who received anti-PD-1/anti-CTLA-4 immunotherapy and followed at the Cardio-Oncology Centre (North Hospital, Marseille). Twelve hiPSC clones were generated from patients who developed myocarditis (ICI-M, 6 clones from 3 patients), and patients who did not develop cardiotoxicity (non-ICI-M, 6 clones from 3 patients). hiPSC-derived endothelial cells (hiPSC-EC) were differentiated from hiPSC clones and stimulated by 10 ng/mL of IFN-γ during 48 h. IFN-γ induced a decrease in cell viability, as well as an increase in the expression of transcripts of genes associated with the JAK/STAT and NLRP3 inflammasome pathways in hiPSC-EC. However, the decrease in cell viability was significantly greater in hiPSC-EC from the ICI-M group compared to the non-ICI-M group. IFN-γ activation of inflammatory intracellular signalling is greater in the ICI-M group, in particular overexpression of the jak1, jak2, stat3 and gbp6 transcripts. At the protein level, in this ICI-M group, the IFN-γ-induced increase of MHC-I and PD-L1 is greater than in the non-ICI-M group. IFN-γ stimulation revealed specific regulations in endothelial cells derived from hiPSC clones from patients with ICI-related myocarditis. We are now analysing the apoptotic and inflammatory activity of hiPSC-EC by real-time microscopy monitoring of caspases (Incucyte S3) and by assaying pro-inflammatory molecules in the supernatants (ELISA). This study will allow to establish the role played by endothelial cells in cardiac toxicity induced by ICI immunotherapy. [ABSTRACT FROM AUTHOR]

The anthracycline doxorubicin (DOX) is a widely used anticancer drug, nonetheless, responsible for significant cardiotoxic effects. The mechanisms of this cardiotoxicity are not fully understood, although they seem to be mainly linked to mitochondrial dysfunctions, i.e. a decrease in ATP production and calcium overload. The ATP-derived adenosine and its A2A receptors (A2AR) have been shown to distinctly modulate in mice the DOX-induced cardiotoxicity, in a way dependent on the timing of A2AR activation during the cardiotoxic response (Hamad et al., 2009). On the other hand, in mouse chondrocytes, A2AR stimulation was able to improve mitochondrial metabolism (Castro et al., 2020). To better understand the impact of adenosine and its receptors in the DOX-induced cardiac toxicity, the development of human cellular models would be extremely important. Characterization of the role of A2AR in DOX-induced dysfunction in human endothelial cells and cardiomyocytes in order to better understand the pathophysiological mechanisms of this cellular cardiotoxicity. We used induced pluripotent stem cells (hiPSCs), differentiated into cardiomyocytes (CM) and endothelial cells (EC), treated with DOX (200 nM, 48 h), in the presence or absence of an A2AR agonist, CGS-21680 (70 nM, 24 h or 48 h). The differentiation of hiPSCs into CM and EC show all the expected characteristics of these cell types. We have verified the expression of A2AR, at the transcriptional and protein level, in both cell types and its quantification in the presence of DOX and/or CGS is in progress. Our preliminary results indicate a gene-expression regulation of A2AR-associated signaling pathways in these cellular models. The characterization of mitochondrial function in CM, and cytokine production in EC is currently under investigation. Our results will provide a better understanding of both the role of A2AR in DOX-induced cardiotoxicity and the implication of EC and CM in this cardiotoxicity. Moreover, this study will allow characterizing the A2AR intracellular signaling pathways functioning in hiPSCs-derived cardiac cells. [ABSTRACT FROM AUTHOR]

Immune checkpoint inhibitors (ICIs) reactivate T cells to fight tumor cells, unraveling a new approach in cancer treatment. However, cardiovascular complications induced by these ICIs, especially myocarditis, have emerged as particularly dangerous complications with a mortality rate close to 50%. A recent study on cardiac biopsies showed that the IFN-γ pathway and inflammasome regulating proteins was specifically involved in myocarditis induced by ICIs treatment; this mechanism was not found in viral myocarditis and in dilated cardiomyopathies [1]. It is therefore crucial to set up models to study these pathophysiological mechanisms so that the benefit of immunotherapies is not compromised by their adverse cardiac effects. To better understand pathophysiological mechanisms of myocarditis induced by ICIs, we characterized the response to IFN-γ of endothelial cells (hiPSC-EC) generated from cells of patients with and without ICIs-induced myocarditis. We generated human-induced pluripotent stem cell (hiPSC), differentiated in hiPSC-EC, from cells of patients followed in the MediCO Center, Marseille. We considered two groups of patients: with myocarditis (Myoc) and without myocarditis (non-Myoc) following anti-PD-1/anti-CTLA4 immunotherapy. hiPSC-EC were treated with 10 ng/mL of IFN-γ during 48 h. Endothelial cells differentiated from hiPSC clones exhibited the characteristics expected for this cell type in all clones tested, both in the Myoc and non-Myoc patient groups. Stimulation by IFN-γ activated inflammatory intracellular signaling in hiPSC-EC, in particular overexpression of MHC class I and II, and PDL-1. Moreover, changes in the expression of genes of the JAK/STAT pathway and of the inflammasome, particularly the NLRP3 inflammasome, were observed in all groups of tested cells after the IFN-γ stimulation. However, the cells from Myoc patients seemed to be more impaired than those from non-Myoc patients. In particular, hiPSC-EC proliferation was more altered in this group by IFN-γ, and the expression of genes associated with the inflammatory response was strongly increased. Further analyses are in progress to confirm these preliminary data. These preliminary results, obtained in cells reprogrammed from cells of patients with immuno-induced myocarditis, are in agreement with the data published by Finke et al. (2021) in myocardial biopsies, indicating that these hiPSCs are a good model to study the pathophysiological mechanisms associated with the cardiotoxicitiy of anticancer immunotherapy. [ABSTRACT FROM AUTHOR]