S-(3-Carboxypropyl)-L-cysteine

Hydrogen sulfide (H2S), mainly produced by cystathionine γ-lyase (CSE) in vascular system, emerges as a novel gasotransmitter exerting anti-inflammatory and anti-atherosclerotic effects. Alterations of CSE/H2S pathway may thus be involved in atherosclerosis pathogenesis. However, the underlying mechanisms are poorly understood. The present study showed that the levels of CSE mRNA and protein expression, as well as H2S production were decreased in ox-LDL-treated macrophage. CSE overexpression reduced the ox-LDL-stimulated tumor necrosis factor-α (TNF-α) generation in Raw264.7 and primary macrophage while CSE knockdown enhanced it. Exogenous supplementation of H2S with NaHS and Na2S also decreased the production of TNF-α and intercellular adhesion molecule-1 (ICAM-1) in ox-LDL-stimulated macrophage, and alleviated the adhesion of macrophage to endothelial monolayer. Cysteine, a CSE preferential substrate for H2S biosynthesis, produced similar effects on the pro-inflammatory cytokine generation, which were reversed by CSE inhibitors PAG and BCA, respectively. Moreover, NaHS and Na2S attenuated the phosphorylation and degradation of IκBα and p65 nuclear translocation, as well as JNK activation caused by ox-LDL. The JNK inhibitor suppressed the NF-κB transcription activity in ox-LDL-treated cells. Furthermore, inhibitors of NF-κB (PDTC), ERK (U0126 and PD98059) and JNK (SP600125) partially blocked the suppression by ox-LDL on the CSE mRNA levels. Taken together, the findings demonstrate that ox-LDL may down-regulate the CSE/H2S pathway, which plays an anti-inflammatory role in ox-LDL-stimulated macrophage by suppressing JNK/NF-κB signaling. The study reveals new therapeutic strategies for atherosclerosis, based on modulating CSE/H2S pathway.

Hypoxia-induced coronary artery vasodilatation protects the heart by increasing blood flow under ischemic conditions, however its mechanism is not fully elucidated. Hydrogen sulfide (H2S) is reported to be an oxygen sensor/transducer in the vasculature. The present study aimed to identify and characterise the role of H2S in the hypoxic response of the coronary artery, and to define the H2S synthetic enzymes involved. Immunoblotting and immunohistochemistry showed expression of all three H2S-producing enzymes, cystathionine-β-synthase (CBS), cystathionine-γ-lyase (CSE) and 3-mercaptopyruvate sulfurtransferase (MPST), in porcine coronary artery. Artery segments were mounted for isometric tension recording; hypoxia caused a transient endothelium-dependent contraction followed by prolonged endothelium-independent relaxation. The CBS inhibitor amino-oxyacetate (AOAA) reduced both phases of the hypoxic response. The CSE inhibitor dl-propargylglycine (PPG) and aspartate (limits MPST) had no effect alone, but when applied together with AOAA the hypoxic relaxation response was further reduced. Exogenous H2S (Na2S and NaHS) produced concentration-dependent contraction followed by prolonged relaxation. Responses to both hypoxia and exogenous H2S were dependent on the endothelium, NO, cGMP, K+ channels and Cl-/HCO3- exchange. H2S production in coronary arteries was blocked by CBS inhibition (AOAA), but not by CSE inhibition (PPG). These data show that H2S is an endogenous mediator of the hypoxic response in coronary arteries. Of the three H2S-producing enzymes, CBS, expressed in the vascular smooth muscle, appears to be the most important for H2S generated during hypoxic relaxation of the coronary artery. A contribution from other H2S-producing enzymes only becomes apparent when CBS activity is inhibited.

The onion-derived metabolite, S-1-propenylmercaptocysteine (CySSPe), protects against oxidative stress and exhibits anti-inflammatory effects by modulating cellular redox homeostasis. We sought to establish whether CySSPe activates nuclear factor erythroid 2-related factor 2 (Nrf2) and whether activation of Nrf2 by CySSPe involves modification of the Kelch-like ECH-associated protein-1 (Keap1) to manifest these effects. We found that CySSPe stabilized Nrf2 protein and facilitated nuclear translocation to induce expression of antioxidant enzymes, including NQO1, HO-1, and GCL. Moreover, CySSPe attenuated tert-butyl hydroperoxide-induced cytotoxicity and dose-dependently inhibited reactive oxygen species production. Silencing experiments using Nrf2-siRNA confirmed that CySSPe conferred protection against oxidative stress by activating Nrf2. CySSPe enhanced cellular pool of reduced glutathione (GSH) and improved GSH:GSSG ratio. Pretreatment of cells with l-buthionine-S,R-sulfoximine (BSO) confirmed that CySSPe increases de novo synthesis of GSH by upregulating expression of the GSH-synthesizing enzyme GCL. Treatment of cells with CySSPe elevated hydrogen sulfide (H2S) production. Inhibition of H2S-synthesizing enzymes, cystathionine-gamma-lyase (CSE) and cystathionine-beta-synthase (CBS), by pretreating cells with propargylglycine (PAG) and oxyaminoacetic acid (AOAA) revealed that H2S production was partially dependent on a CSE/CBS-catalyzed β-elimination reaction with CySSPe that likely produced 1-propenyl persulfide (RSSH). Depleting cells of their GSH pool by exposure to BSO and diethylmaleate attenuated H2S production, suggesting a GSH-dependent formation of H2S, likely via the reduction of RSSH by GSH. Finally, treatment of cells with CySSPe persulfidated Keap1, which may be the mechanism involved for the stabilization of Nrf2 by CySSPe. Taken together, our results showed that attenuation of oxidative stress by CySSPe is associated with its ability to produce H2S or RSSH, which persulfidates Keap1 and activates Nrf2 signaling. This study provides insights on the potential of CySSPe as an onion-derived dietary agent that modulates redox homeostasis and combats oxidative stress.