Alice Marino

Arylthioamides as H2S Donors: l-Cysteine-Activated Releasing Properties and Vascular Effects in Vitro and in Vivo

A small library of arylthioamides 1-12 was easily synthesized, and their H2S-releasing properties were evaluated both in the absence or in the presence of an organic thiol such as l-cysteine. A number of arylthioamides (1-3 and 7) showed a slow and l-cysteine-dependent H2S-releasing mechanism, similar to that exhibited by the reference slow H2S-releasing agents, such as diallyl disulfide (DADS) and the phosphinodithioate derivative GYY 4137. Compound 1 strongly abolished the noradrenaline-induced vasoconstriction in isolated rat aortic rings and hyperpolarized the membranes of human vascular smooth muscle cells in a concentration-dependent fashion. Finally, a significant reduction of the systolic blood pressure of anesthetized normotensive rats was observed after its oral administration. Altogether these results highlighted the potential of arylthioamides 1-3 and 7 as H2S-donors for basic studies, and for the rational design/development of promising pharmacotherapeutic agents to treat cardiovascular diseases.

Histamine H4-Receptors Inhibit Mast Cell Renin Release in Ischemia/Reperfusion via Protein Kinase Cε-Dependent Aldehyde Dehydrogenase Type-2 Activation

Renin released by ischemia/reperfusion (I/R) from cardiac mast cells (MCs) activates a local renin-angiotensin system (RAS) causing arrhythmic dysfunction. Ischemic preconditioning (IPC) inhibits MC renin release and consequent activation of this local RAS. We postulated that MC histamine H4-receptors (H4Rs), being Gαi/o-coupled, might activate a protein kinase C isotype–ε (PKCε)–aldehyde dehydrogenase type-2 (ALDH2) cascade, ultimately eliminating MC-degranulating and renin-releasing effects of aldehydes formed in I/R and associated arrhythmias. We tested this hypothesis in ex vivo hearts, human mastocytoma cells, and bone marrow–derived MCs from wild-type and H4R knockout mice. We found that activation of MC H4Rs mimics the cardioprotective anti-RAS effects of IPC and that protection depends on the sequential activation of PKCε and ALDH2 in MCs, reducing aldehyde-induced MC degranulation and renin release and alleviating reperfusion arrhythmias. These cardioprotective effects are mimicked by selective H4R agonists and disappear when H4Rs are pharmacologically blocked or genetically deleted. Our results uncover a novel cardioprotective pathway in I/R, whereby activation of H4Rs on the MC membrane, possibly by MC-derived histamine, leads sequentially to PKCε and ALDH2 activation, reduction of toxic aldehyde-induced MC renin release, prevention of RAS activation, reduction of norepinephrine release, and ultimately to alleviation of reperfusion arrhythmias. This newly discovered protective pathway suggests that MC H4Rs may represent a new pharmacologic and therapeutic target for the direct alleviation of RAS-induced cardiac dysfunctions, including ischemic heart disease and congestive heart failure.

E-NTPDase1/CD39 modulates renin release from heart mast cells during ischemia/reperfusion: a novel cardioprotective role

Ischemia/reperfusion (I/R) elicits renin release from cardiac mast cells (MC), thus activating a local renin‐angiotensin system (RAS), culminating in ventricular fibrillation. We hypothesized that in I/R, neurogenic ATP could degranulate juxtaposed MC and that ecto‐nucleoside triphosphate diphosphohydrolase 1/CD39 (CD39) on MC membrane could modulate ATP‐induced renin release. We report that pharmacological inhibition of CD39 in a cultured human mastocytoma cell line (HMC‐1) and murine bone marrow‐derived MC with ARL67156 (100 mM) increased ATP‐induced renin release (≥2‐fold), whereas purinergic P2X7 receptors (P2X7R) blockade with A740003 (3 mM) prevented it. Likewise, CD39 RNA silencing in HMC‐1 increased ATP‐induced renin release (≥2‐fold), whereas CD39 overexpression prevented it. Acetaldehyde, an I/R product (300 μM), elicited an 80% increase in ATP release from HMC‐1, in turn, causing an autocrine 20% increase in renin release. This effect was inhibited or potentiated when CD39 was overexpressed or silenced, respectively. Moreover, P2X7R silencing prevented ATP‐ and acetaldehyde‐induced renin release. I/R‐induced RAS activation in ex vivo murine hearts, characterized by renin and norepinephrine overflow and ventricular fibrillation, was potentiated (~2‐fold) by CD39 inhibition, an effect prevented by P2X7R blockade. Our data indicate that by regulating ATP availability at the MC surface, CD39 modulates local renin release and thus, RAS activation, ultimately exerting a cardioprotective effect.—Aldi, S., Marino, A., Tomita, K., Corti, F., Anand, R., Olson, K. E., Marcus, A. J., Levi, R., E‐NTPDase1/ CD39 modulates renin release from heart mast cells during ischemia/reperfusion: a novel cardioprotective role. FASEB J. 29, 61–69 (2015). www.fasebj.org

The novel H2S donor 4‐carboxy‐phenyl isothiocyanate inhibits mast cell degranulation and renin release by decreasing intracellular calcium

Hydrogen sulfide (H2S) modulates many pathophysiological processes, including inflammation and allergic reactions, in which mast cells act as major effector cells. IgE receptor (FcεRI) cross linking leads to an increase in intracellular calcium ([Ca+2]i), a critical step in mast cell degranulation. The aim of this study was to investigate the role of H2S in [Ca+2]i‐dependent mast cell activation.

S1P receptor 1-Mediated Anti–Renin-Angiotensin System Cardioprotection: Pivotal Role of Mast Cell Aldehyde Dehydrogenase Type 2

In the ischemic-reperfused (I/R) heart, renin-containing mast cells (MC) release enzymatically active renin, activating a local renin-angiotensin system (RAS), causing excessive norepinephrine release and arrhythmic dysfunction. Activation of Gi-receptors on MC and/or ischemic preconditioning (IPC) prevent renin release, thus providing anti-RAS cardioprotection. We questioned whether sphingosine-1-phosphate (S1P), a sphingolipid produced in the I/R heart, might afford anti-RAS cardioprotection by activating Gi-coupled S1P1 receptors (S1P1R) on MC. We report that activation of Gi-coupled S1P1R in cardiac MC confers IPC-like anti-RAS cardioprotection due to S1P1R-mediated inhibition of I/R-induced cardiac MC degranulation and renin release. This results from an initial translocation of protein kinase C subtype-ε and subsequent activation of aldehyde dehydrogenase type 2 (ALDH2), culminating in the elimination of the MC-degranulating effects of acetaldehyde and other toxic species produced during I/R. Inhibition of toxic aldehydes-induced MC-renin release prevents local RAS activation, reduces infarct size, and alleviates arrhythmias. Notably, these cardioprotective effects are lacking in hearts and MC from gene-targeted knock-in mice (ALDH2*2) in which ALDH2 enzymatic activity is maximally reduced. Thus, ALDH2 appears to play a pivotal role in this protective process. Our findings suggest that MC S1P1R may represent a new pharmacologic and therapeutic target for the direct alleviation of RAS-induced cardiac dysfunctions, including ischemic heart disease and congestive heart failure.

Selective Activation of Retinoic Acid β2 Receptors Exerts Cardioprotective Effects

We previously discovered that oral treatment with AC261066, a synthetic selective agonist for the retinoic acid β2-receptor, decreases oxidative stress in the liver, pancreas, and kidney of mice fed a high-fat diet (HFD). Since hyperlipidemic states are causally associated with myocardial ischemia and oxidative stress, we have now investigated the effects of AC261066 in an ex vivo ischemia/reperfusion (I/R) injury model in hearts of two prototypic dysmetabolic mice. We found that a 6-week oral treatment with AC261066 in both genetically hypercholesterolemic (ApoE−/−) and obese (HFD-fed) wild-type mice exerts protective effects when their hearts are subsequently subjected to I/R ex vivo in the absence of added drug. In ApoE−/− mice this cardioprotection ensued without hyperlipidemic changes. Cardioprotection consisted of attenuation of infarct size, diminution of norepinephrine (NE) spillover, and alleviation of reperfusion arrhythmias. This cardioprotection was associated with a reduction in oxidative stress and mast cell (MC) degranulation. We suggest that the reduction in myocardial injury and adrenergic activation, and the antiarrhythmic effects, result from decreased formation of oxygen radicals and toxic aldehydes known to elicit the release of MC-derived renin, promoting the activation of the local renin-angiotensin system leading to enhanced NE release and reperfusion arrhythmias. Because these beneficial effects of AC261066 occurred at the ex vivo level following oral drug treatment, our data suggest that AC261066 could be viewed as a therapeutic means to reduce I/R injury of the heart, and potentially also be considered in the treatment of other cardiovascular ailments such as chronic arrhythmias and cardiac failure.