Cysteine Trisulfide Protects E. coli from Electrophile-Induced Death through the Generation of Cysteine Hydropersulfide.

Hydropersulfide and polysulfide species have recently been shown to elicit a wide variety of biological and physiological responses. In this study, we examine the effects of cysteine trisulfide (Cys-SSS-Cys; also known as thiocystine) treatment on E. coli. Previous studies in mammalian cells have shown that Cys-SSS-Cys treatment results in protection from the electrophiles. Here, we show that the protective effect of Cys-SSS-Cys treatment against electrophile-induced cell death is conserved in E. coli. This protection correlates with the rapid generation of cysteine hydropersulfide (Cys-SSH) in the culture media. We go on to demonstrate that an exogenous phosphatase expressed in E. coli, containing only a single catalytic cysteine, is protected from electrophile-induced inactivation in the presence of hydropersulfides. These data together demonstrate that E. coli can utilize Cys-SSS-Cys to generate Cys-SSH and that the Cys-SSH can protect cellular thiols from reactivity with the electrophiles.

The Uptake and Release of Polysulfur Cysteine Species by Cells: Physiological and Toxicological Implications.

Hydropersulfides and related polysulfides have recently become topics of significant interest due to their physiological prevalence and proposed biological functions. Currently, examination of the effects of hydropersulfide treatment on cells is difficult due to their lack of inherent stability with respect to disproportionation. Herein, it is reported that the treatment of a variety of cell types with cysteine trisulfide (also known as thiocystine; Cys-SSS-Cys), results in an increase in intracellular hydropersulfide levels (e.g., cysteine hydropersulfide; Cys-SSH, and glutathione hydropersulfide; GSSH). Thus, Cys-SSS-Cys represents a possible pharmacological agent for examining the effects of hydropersulfides on cell function/viability. It has also been found that cells with increased intracellular hydropersulfide levels can export Cys-SSH into the extracellular media. Interestingly, the Cys-SSH is the major hydropersulfide exported by cells, although GSSH is the predominant intracellular species. The possible implications of cellular export are discussed.

The reaction of hydrogen sulfide with disulfides: formation of a stable trisulfide and implications for biological systems.

BACKGROUND AND PURPOSE: The signalling associated with hydrogen sulfide (H2 S) remains to be established, and recent studies have alluded to the possibility that H2 S-derived species play important roles. Of particular interest are hydropersulfides (RSSH) and related polysulfides (RSSn R, n > 1). This work elucidates the fundamental chemical relationship between these sulfur species as well as examines their biological effects. EXPERIMENTAL APPROACH: Using standard analytical techniques (1 H-NMR and MS), the equilibrium reactions between H2 S, disulfides (RSSR), RSSH, dialkyltrisulfides (RSSSR) and thiols (RSH) were examined. Their ability to protect cells from electrophilic and/or oxidative stress was also examined using cell culture. KEY RESULTS: H2 S, RSSR, RSSH, RSSSR and RSH are all in a dynamic equilibrium. In a biological system, these species can exist simultaneously, and thus, it is difficult to discern which species is (are) the biological effector(s). Treatment of cells with the dialkyl trisulfide cysteine trisulfide (Cys-SSS-Cys) resulted in high intracellular levels of hydropersulfides and protection from electrophilic stress. CONCLUSIONS AND IMPLICATIONS: In aqueous systems, the reaction between H2 S and RSSR results in the formation of equilibria whereby H2 S, RSH, RSSR, RSSH and RSSSR are present. In a biological system, any of these species can be responsible for the observed biological activity. These equilibrium species can also be generated via the reaction of RSH with RSSSR. Due to these equilibria, Cys-SSS-Cys can be a method for generating any of the other species. Importantly, HEK293T cells treated with Cys-SSS-Cys results in increased levels of hydropersulfides, allowing examination of the biological effects of RSSH. LINKED ARTICLES: This article is part of a themed section on Chemical Biology of Reactive Sulfur Species. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v176.4/issuetoc.

Chronic exposure of the RAW246.7 macrophage cell line to H2O2 leads to increased catalase expression.

Altered cellular redox states have been associated with a variety of chronic diseases, especially those correlated with inflammation. One of the primary oxidants generated during the inflammatory response is hydrogen peroxide (H2O2). Macrophages in particular are thought to produce large amounts of H2O2, however they must somehow protect themselves from the potentially lethal concentrations they produce. To investigate how immune cells protect themselves from H2O2 observed in chronic inflammatory diseases, we established an adapted population of macrophages in culture by gradually increasing sub-lethal concentrations of H2O2 in the media to typically lethal concentrations over the course of more than a month. The resulting cells were tolerant to very high concentrations of H2O2. Further investigation revealed that the cells were able to rapidly neutralize the H2O2 added to their culture media due to a dramatic upregulation of catalase. Interestingly, T cells, which are also implicated in chronic inflammation, were unable to adapt to H2O2 under the same procedure, however when T cells were cultured in media from adapted macrophages, they were able to survive typically lethal concentrations of H2O2. These data support the hypothesis that macrophages are able to protect themselves and neighboring cells during states of chronic inflammation from the oxidizing environment they create.