BackgroundHydrogen sulfide (H2S) exhibits protective effects in various disease models including cerebral ischemia–reperfusion (I/R) injury. Nonetheless, mechanisms and identity of molecules responsible for neuroprotective effects of H2S remain incompletely defined. In the current study, we observed that thiosulfate, an oxidation product of H2S, mediates protective effects of an H2S donor compound sodium sulfide (Na2S) against neuronal I/R injury.Methods and ResultsWe observed that thiosulfate in cell culture medium is not only required but also sufficient to mediate cytoprotective effects of Na2S against oxygen glucose deprivation and reoxygenation of human neuroblastoma cell line (SH‐SY5Y) and murine primary cortical neurons. Systemic administration of sodium thiosulfate (STS) improved survival and neurological function of mice subjected to global cerebral I/R injury. Beneficial effects of STS, as well as Na2S, were associated with marked increase of thiosulfate, but not H2S, in plasma and brain tissues. These results suggest that thiosulfate is a circulating “carrier” molecule of beneficial effects of H2S. Protective effects of thiosulfate were associated with inhibition of caspase‐3 activity by persulfidation at Cys163 in caspase‐3. We discovered that an SLC13 family protein, sodium sulfate cotransporter 2 (SLC13A4, NaS‐2), facilitates transport of thiosulfate, but not sulfide, across the cell membrane, regulating intracellular concentrations and thus mediating cytoprotective effects of Na2S and STS.ConclusionsThe protective effects of H2S are mediated by thiosulfate that is transported across cell membrane by NaS‐2 and exerts antiapoptotic effects via persulfidation of caspase‐3. Given the established safety track record, thiosulfate may be therapeutic against ischemic brain injury.

Rationale The regulation of calcium (Ca2+) homeostasis by beta-adrenergic receptor (βAR) activation provides the essential underpinnings of sympathetic regulation of myocardial function as well as a basis for understanding molecular events that result in hypertrophic signaling and heart failure. Sympathetic stimulation of the βAR not only induces protein phosphorylation but also activates nitric oxide (NO)-dependent signaling, which modulates cardiac contractility. Nonetheless, the role of NO in βAR-dependent regulation of Ca2+ handling has not yet been explicated fully. Objective To elucidate the role of protein S-nitrosylation, a major transducer of NO bioactivity, on βAR-dependent alterations in cardiomyocyte Ca2+ handling and hypertrophy. Methods and Results Using transgenic mice to titrate the levels of protein SNO, we uncovered major roles for protein S-nitrosylation generally, and for phospholamban (PLN) and cardiac troponin C (cTnC) S-nitrosylation in particular, in βAR-dependent regulation of Ca2+ homeostasis. Notably, S-nitrosylation of PLN consequent upon βAR stimulation is necessary for its inhibitory pentamerization of PLN, which activates sarcoplasmic reticulum Ca2+-ATPase (SERCA2a) and increases cytosolic Ca2+ transients. Coincident S-nitrosylation of cTnC decreases myocardial sensitivity to Ca2+. During chronic adrenergic stimulation, global reductions in cellular S-nitrosylation mitigate hypertrophic signaling resulting from Ca2+ overload. Conclusions S-nitrosylation operates in concert with phosphorylation to regulate many cardiac Ca2+-handling proteins, including PLN and cTnC, thereby playing an essential and previously unrecognized role in cardiac Ca2+ homeostasis. Manipulation of the S-nitrosylation level may prove therapeutic in heart failure.

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Aims Mitochondria-targeted hydrogen sulfide donor AP39, [(10-oxo-10-(4-(3-thioxo-3H-1,2-dithiol-5yl)phenoxy)decyl) triphenylphosphonium bromide], exhibits cytoprotective effects against oxidative stress in vitro. We examined whether or not AP39 improves neurological function and long term survival in mice subjected to cardiac arrest (CA) and cardiopulmonary resuscitation (CPR). Methods Adult C57BL/6 male mice were subjected to 8 min of CA and subsequent CPR. We examined the effects of AP39 (10, 100, 1000 nmol kg−1) or vehicle administered intravenously at 2 min before CPR (Experiment 1). Systemic oxidative stress levels, mitochondrial permeability transition, and histological brain injury were assessed. We also examined the effects of AP39 (10, 1000 nmol kg−1) or vehicle administered intravenously at 1 min after return of spontaneous circulation (ROSC) (Experiment 2). ROSC was defined as the return of sinus rhythm with a mean arterial pressure > 40 mm Hg lasting at least 10 seconds. Results Vehicle treated mice subjected to CA/CPR had poor neurological function and 10-day survival rate (Experiment 1; 15%, Experiment 2; 23%). Administration of AP39 (100 and 1000 nmol kg−1) 2 min before CPR significantly improved neurological function and 10-day survival rate (54% and 62%, respectively) after CA/CPR. Administration of AP39 before CPR attenuated mitochondrial permeability transition pore opening, reactive oxygen species generation, and neuronal degeneration after CA/CPR. Administration of AP39 1 min after ROSC at 10 nmol kg−1, but not at 1000 nmol kg−1, significantly improved neurological function and 10 day-survival rate (69%) after CA/CPR. Conclusion The current results suggest that administration of mitochondria-targeted sulfide donor AP39 at the time of CPR or after ROSC improves neurological function and long term survival rates after CA/CPR by maintaining mitochondrial integrity and reducing oxidative stress.