Pathophysiological roles of H2S in inflammation: inhibition of mast cell degranulation and potential pharmacotherapeutic uses of H2S-releasing drugs

Hydrogen sulfide (H2S) is an endogenous gasotransmitter involved in the regulation of many physiological mechanisms in the cardiovascular, respiratory, gastroenteric, endocrine and central nervous systems. Accordingly, a pharmacological modulation of this gasotransmitter is becoming a challenging research field in drug discovery. In particular, H2S-donors, molecules able to release H2S and restore its impaired production, appear to be a promising class of drugs. It is now accepted that H2S is involved in inflammation, although there are many controversial hypothesis on its roles. Mast cells are important cells related to inflammatory events and a rational strategy to reduce the inflammation process could be to reduce mast cell activation and the consequent release of pro-inflammatory mediators from their internal granules. Yet, the influence of endogenous H2S and of H2S-releasing drugs on these cells has been poorly investigated to date. This PhD thesis presents new results in the discovery of novel H2S-donors, both of natural and synthetic origin, belonging to different chemotypes, and the evaluation of their effects on mast cell degranulation and in mast cell-mediated cardiac injury. In a first phase, a wide array of H2S-releasing compounds was probed; compounds exhibiting the most interesting profile were selected for subsequent studies. After that, possible anti-degranulating effects of H2S were tested on three different cell lines, RBL-2H3 (rat basophilic leukemia cells), BMMC (bone marrow-derived mast cells) and HMC-1 (human mastocytoma cells) degranulated by different stimuli (antigenic and non-antigenic). This part of the study revealed significant inhibitory effect of H2S-releasing drugs on the mast cell degranulation of all the mast cell lines, indicating that: a) endogenous H2S may play significant roles in modulating mast cell degranulation, and thus in influencing mast cell-mediated inflammatory processes; b) H2S-releasing drugs may be useful in treating pathological situations associated with mast cell degranulation. Recently, it has been demonstrated that cardiac mast cells, closely located to the nerve endings, are a previously unrecognized source of renin, which is stored and released from intracellular granules. Released renin activates a local renin-angiotensin system (RAS) and the promotes norepinephrine (NE) release from sympathetic nerve endings, culminating in severe cardiac dysfunction. In particular, mast cell degranulation is elicited by ischemia/reperfusion (I/R) injury and is correlated to the severity of the I/R damage. Since the cardioprotective activity of H2S in the cardiovascular system is widely reported, but its mechanisms are still largely unexplored, the last phase of the experimental work investigated the possible relation between H2S-induced cardioprotection and inhibition of heart mast cell degranulation in rat and mouse models of myocardial I/R. The preliminary results of this phase show: a) that the selected H2S-releasing compound failed to inhibit mast cell degranulation and renin release when induced by I/R; b) the H2S-releasing compound significantly reduced NE release. In addition, other cardiac parameters indicating the functionality of the heart were measured, such as rate pressure product (RPP), expressed as product of heart rate (HR) and left ventricular developed pressure (LVDP), arrhythmias duration (VT/VF) and coronary flow (CF). Although those factors were restored to basal levels by the H2S-donor both in rat and murine models of I/R, these results suggest that cardioprotection might not be linked to inhibition of mast cell degranulation. Indeed, when 5-hydroxydecanoic acid (5-HD), a selective inhibitor of mitochondrial ATP potassium channels (mitoKATP), was pre-injected in the presence of the H2S-donor, a clear antagonism was observed on the anti-ischemic effect of the H2S-donor. This preliminary result indicates that the activity of H2S, released by the donor, has an important role in cardioprotection and that mitoKATP could be considered a promising target for its cardioprotective role.