Hydrogen sulfide (H2S) is an endogenous signaling molecule which like nitric oxide plays important roles in physiological functions related to cardiovascular health, but it’s mechanism of action remains poorly understood. We have developed methodology to characterize and quantify endogenous hydrogen sulfide (H2S) and small oxoacids of sulfur (SOS = HOSH, HOSOH) in numerous cell lines and mammalian tissue samples via translation of well-studied nucleophilic and electrophilic traps for cysteine and oxidized cysteines into a mass spectrometry methodology to trap small molecular weight species. This method allows quantification of these species giving both intra- and extra-cellular concentrations based on an H2S calibration curve. We propose the term sulfomic index, a ratio of S-O content, as a profile of non-protein sulfide redox status. The production of H2S and SOS is orders of magnitude higher in human melanoma and smooth muscle cells (nanomolar) as compared to other human cells (picomolar) and yeast and bacteria (femtomolar). In many cells the analytes concentrations following pattern of H2S &gt; HOSOH &gt; HOSH, but in primary vascular lines had distributions where HOSOH &gt; H2S &gt; HSOH, with much higher sulfomic indexes than seen in non-vascular derived cell lines. An important observation is that H2S and SOS are effluxed into surrounding media in much higher concentrations than found intracellularly. The efflux was continuous over time and against a concentration gradient, implying an active transport. Human primary aortic smooth muscle cells were used to examine changes in intracellular and efflux concentrations of H2S and SOS under hypoxic growth conditions. Under hypoxia, the smooth muscle cells demonstrated reduced mitochondrial respiration and increased ROS production, with a corresponding reduction in levels of H2S and HOSOH, but large increases in levels of HSOH, which we attribute to retrograde electron transfer to Complex 1 during hypoxia. We believe that measurements and characterizations these analytes under different disease states may help uncover the mechanisms of action attributed to H2S in cardiovascular health and disease.

Expanding the sulfome: Characterization of hydrogen sulfide (H2S) and small oxoacids of sulfur (SOS) using derivatization and mass spectrometry

ACS Spring 2022

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