Extended hypoxia‐mediated H₂S production provides for long‐term oxygen sensing

Abstract

Aim Numerous studies have shown that H2S serves as an acute oxygen sensor in a variety of cells. We hypothesize that H2S also serves in extended oxygen sensing. Methods Here, we compare the effects of extended exposure (24‐48 hours) to varying O2 tensions on H2S and polysulphide metabolism in human embryonic kidney (HEK 293), human adenocarcinomic alveolar basal epithelial (A549), human colon cancer (HTC116), bovine pulmonary artery smooth muscle, human umbilical‐derived mesenchymal stromal (stem) cells and porcine tracheal epithelium (PTE) using sulphur‐specific fluorophores and fluorometry or confocal microscopy. Results All cells continuously produced H2S in 21% O2 and H2S production was increased at lower O2 tensions. Decreasing O2 from 21% to 10%, 5% and 1% O2 progressively increased H2S production in HEK293 cells and this was partially inhibited by a combination of inhibitors of H2S biosynthesis, aminooxyacetate, propargyl glycine and compound 3. Mitochondria appeared to be the source of much of this increase in HEK 293 cells. H2S production in all other cells and PTE increased when O2 was lowered from 21% to 5% except for HTC116 cells where 1% O2 was necessary to increase H2S, presumably reflecting the hypoxic environment in vivo. Polysulphides (H2Sn, where n = 2‐7), the key signalling metabolite of H2S also appeared to increase in many cells although this was often masked by high endogenous polysulphide concentrations. Conclusion These results show that cellular H2S is increased during extended hypoxia and they suggest this is a continuously active O2‐sensing mechanism in a variety of cells.

Effects of Manganese Porphyrins on Cellular Sulfur Metabolism

“…Reducing ambient oxygen from 21% to 5% increased intracellular H2S (AzMC fluorescence), consistent with previous studies showing that oxygen-dependent metabolism of H2S is a mechanism for short-and long-term cellular oxygen sensing [7,11]. H2S is constitutively produced from cysteine catabolism via the transsulfuration pathway and metabolized in the mitochondrion, initially by the mitochondrial enzyme sulfur:quinone oxidoreductase [12].…”

“…If each cell produces 3 × 10 −16 mol of H 2 S per cell [23], intracellular H 2 S could theoretically increase to 200 µM in one hour if H 2 S was not metabolized. Mitochondrial oxidation of H 2 S to sulfate normally keeps pace with H 2 S production but this can quickly change as evidenced by the rapid rise in H 2 S associated with hypoxia [11] and by the longer term responses observed in the present and other studies [7]. While it is tempting to propose that MnP-oxidation diverts H 2 S generated from cysteine catabolism away from mitochondrial oxidation, this does not appear to be the case.…”

“…shows the effects of 24 h exposure to 1 µM MnPs on intracellular sulfur distribution in HEK293 cells. MnTE and MnHex significantly reduced cellular H 2 S and all three MnPs reduced H 2 S in organelles that have been identified in other studies as mitochondria[7].…”

“…Cells cultured under the above conditions are 'hyperoxic' relative to their 'physioxic' environment in vivo [46]. Due to the fact that exposure to more physioxic conditions also affects sulfur metabolism [7], a number of additional experiments were performed at more appropriate oxygen levels (5% O 2 /5% CO 2 /balance N 2 ) in a model 856-HYPO hypoxia chamber (Plas Labs, Inc. Lansing, MI, USA) at 37 • C. They were covered with a standard 96-well plate cover before they were removed from the hypoxia chamber for plate reader measurements.…”

Environment and exposure

“…24 Lastly, Olson et al found that cellular H 2 S is increased during extended hypoxia, probably functioning as a continuously active O 2 -sensing mechanism in several cells. 25 Protection from detrimental environmental influences has, without doubt, contributed to modern man living longer and healthier lives than ever before in human history. However, exposure to the natural environment has, in an increasingly urbanized world, often become a planned project to be undertaken at leisure.…”