Hydrogen sulfide (H 2 S), as a reducing agent and an antioxidant molecule, exerts protective effects against hyperglycemic stress in the vascular endothelium. The mitochondrial enzyme 3-mercaptopyruvate sulfurtransferase (3-MST) is an important biological source of H 2 S. We have recently demonstrated that 3-MST activity is inhibited by oxidative stress in vitro and speculated that this may have an adverse effect on cellular homeostasis. In the current study, given the importance of H 2 S as a vasorelaxant, angiogenesis stimulator and cellular bioenergetic mediator, we first determined whether the 3-MST/H 2 S system plays a physiological regulatory role in endothelial cells. Next, we tested whether a dysfunction of this pathway develops during the development of hyperglycemia and diabetes-associated vascular complications. Intraperitoneal (IP) 3-MP (1 mg/kg) raised plasma H 2 S levels in rats. 3-MP (10 μmol/L to 1 mmol/L) promoted angiogenesis in vitro in bEnd3 microvascular endothelial cells and in vivo in a Matrigel assay in mice (0.3-1 mg/kg). In vitro studies with bEnd3 cell homogenates demonstrated that the 3-MP-induced increases in H 2 S production depended on enzymatic activity, although at higher concentrations (1-3 mmol/L) there was also evidence for an additional nonenzymatic H 2 S production by 3-MP. In vivo, 3-MP facilitated wound healing in rats, induced the relaxation of dermal microvessels and increased mitochondrial bioenergetic function. In vitro hyperglycemia or in vivo streptozotocin diabetes impaired angiogenesis, attenuated mitochondrial function and delayed wound healing; all of these responses were associated with an impairment of the proangiogenic and bioenergetic effects of 3-MP. The antioxidants DL-α-lipoic acid (LA) in vivo, or dihydrolipoic acid (DHLA) in vitro restored the ability of 3-MP to stimulate angiogenesis, cellular bioenergetics and wound healing in hyperglycemia and diabetes. We conclude that diabetes leads to an impairment of the 3-MST/H 2 S pathway, and speculate that this may contribute to the pathogenesis of hyperglycemic endothelial cell dysfunction. We also suggest that therapy with H 2 S donors, or treatment with the combination of 3-MP and lipoic acid may be beneficial in improving angiogenesis and bioenergetics in hyperglycemia.

Hydrogen sulfide (H2S) is an endogenous gaseous biological mediator, which regulates, among others, the oxidative balance of cells under normal physiological conditions, as well as in various diseases. Several previous studies have reported that H2S attenuates inflammatory mediator production. In this study, we investigated the role of H2S in chromatin modulation in an in vitro model of lipopolysaccharide (LPS)-induced inflammation and evaluated its effects on inflammatory cytokine production. Tamm-Horsfall protein 1 (THP-1) differentiated macrophages were pre-treated with sodium hydrosulfide (NaHS) (an H2S donor) at 0.01, 0.1, 0.5 or 1 mM for 30 min. To stimulate cytokine production, the cells were challenged with bacterial LPS (1 μg/ml) for 1, 4, 8 or 24 h. Histone H3 acetylation was analyzed by chromatin immunoprecipitation (ChIP), cytokine production was measured by ELISA and histone deacetylase (HDAC) activity was analyzed using a standard biochemical assay. H2S inhibited the production of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) in a concentration-dependent manner; it was most effective at the two highest concentrations used. This effect was associated with a decrease in histone H3 acetylation at the IL-6 and TNF-α promoters in the cells exposed to H2S or H2S + LPS. The findings of the present study suggest that H2S suppresses histone acetylation, which, in turn, inhibits chromatin openness, leading to a decrease in the gene transcription of various pro-inflammatory cytokines. Therefore, this mechanism may contribute to the previously demonstrated anti-inflammatory effects of H2S and various H2S donors.

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Sepsis survivors suffer from additional morbidities, including higher disk of readmissions, nervous system disturbances and cognitive dysfunction, and increased mortality, even several years after the initial episode of sepsis. In many ways, the phenotype of sepsis survivors resembles the phenotype associated with accelerated aging. Since telomere shortening is a hallmark of aging, we investigated whether sepsis also leads to telomere shortening. Male balb/c mice were divided into two groups: the control group received 100 μl of normal saline intraperitoneally; the sepsis group received 15 mg/kg of bacterial lipopolysaccharide i.p. After 48 hours, animals were sacrificed to collect blood, spleen and kidney. The human component of our study utilized blood samples obtained from patients in the Trauma Department and samples collected 7 days later in those patients who developed sepsis. Telomere length was measured by quantitative PCR. Since oxidative stress is a known inducer of telomere shortening, thiobarbituric acid reactive substances and superoxide dismutase (SOD) activity were analyzed in order to evaluate oxidative stress burden. Induction of endotoxemia in mice resulted in significant telomere shortening in spleen and kidney.Blood cells from patients that progressed to sepsis also exhibited a statistically significant reduction of telomere length. Endotoxemia in mice also induced an earlyonset increase in oxidative stress markers, but was not associated with a downregulation of telomerase protein expression. We conclude that endotoxemia and sepsis induce telomere shortening in various tissues and hypothesize that this may contribute to the pathogenesis of the delayed pathophysiological events in sepsis survivors.