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Recent studies have demonstrated that ferroptosis, a novel form of nonapoptotic regulated cell death plays an important role in doxorubicin (DOX)-induced cardiotoxicity (DoIC). Hydrogen sulfide (H 2 S) is emerging as the third important gaseous mediator in cardiovascular system. However, whether H 2 S has an effect on DOX-induced ferroptosis remains unknown. Here, we found that DOX not only triggered cardiomyocyte ferroptosis but also significantly inhibited the synthesis of endogenous H 2 S in the murine model of chronic DoIC. Application of NaHS, an H 2 S donor obviously activated the SLC7A11/GSH/GPx4 antioxidant pathway and thus alleviated DOX-induced ferroptosis and cardiac injury in mice. In contrast, cardiac-specific knockout of cystathionine γ-lyase gene (Cse) in mice (Cse f/f /Cre + ) to abolish the cardiac synthesis of endogenous H 2 S evidently exacerbated DOX-induced ferroptosis and cardiac dysfunction. A further suppression of SLC7A11/GSH/GPx4 pathway was obtained in Cse f/f /Cre + mice with DoIC, as compared to Cse f/f /Cre - mice with DoIC. The aggravation caused by cardiac-specific Cse deficiency was remarkably rescued by exogenous supplementation of NaHS. Moreover, in DOX-stimulated H9c2 cardiomyocytes, pretreatment with NaHS dose-dependently enhanced the activity of SLC7A11/GSH/GPx4 pathway and subsequently mitigated ferroptosis and mitochondrial impairment. On the contrary, transfection with Cse siRNA in DOX-stimulated H9c2 cardiomyocytes markedly inhibited SLC7A11/GSH/GPx4 pathway, thus leading to aggravated ferroptosis and more damage to mitochondrial structure and function. In addition, the protective effect of NaHS on DOX-induced ferroptosis was closely related to the S-sulfhydrated Keap1, which in turn promoted nuclear translocation of Nrf2 and the transcription of SLC7A11 and GPx4. In conclusion, our findings suggest that H 2 S may exert protective effect on DoIC by inhibiting DOX-induced ferroptosis via Keap1/Nrf2-dependent SLC7A11/GSH/GPx4 antioxidant pathway., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Endothelial‑to‑mesenchymal transition (EndMT) serves an important role in the vascular remodeling of pulmonary arterial hypertension (PAH). However, little is known about the correlation between hydrogen sulfide (H2S), a protective gaseous mediator in PAH and the process of EndMT. Male Sprague‑Dawley rats (10 weeks old) received a single dose of monocrotaline (MCT; i.p., 60 mg/kg) and were randomly treated with NaHS [an H2S donor; intraperitoneal (i.p.) 1 mg/kg/day], DL‑propagylglycine (an inhibitor of H2S synthesis; PAG; i.p., 10 mg/kg/day) or saline, 7 days after MCT injection. Rats were sacrificed 21 days after MCT injection. A selection of human pulmonary artery endothelial cells (HPAECs) were pretreated with NaHS or saline and stimulated with transforming growth factor (TGF)‑β1 (10 ng/ml), and the other HPAECs were transfected with a cystathionine γ‑lyase (CSE, an H2S synthesizing enzyme) plasmid and subsequently stimulated with TGF‑β1. NaHS was indicated to inhibit EndMT and PAH progression by inhibiting the induction of the nuclear factor (NF)‑κB‑Snail pathway. In contrast, the depletion of H2S formation by PAG exacerbated EndMT and PAH by activating NF‑κB‑Snail molecules. In HPAECs, NaHS dose‑dependently inhibited TGF‑β1‑induced EndMT and the activation of the NF‑κB‑Snail pathway. Transfection with a CSE plasmid significantly repressed TGF‑β1‑induced expression of the mesenchymal marker and upregulated the expression of the endothelial marker, which was accompanied by the suppression of the NF‑κB‑Snail pathway. The inhibitory effect of CSE overexpression on TGF‑β1‑induced EndMT was significantly reversed by pretreatment with PAG. In conclusion, the current study provides novel information elucidating the beneficial effect of H2S on PAH through inhibiting the induction of the NF‑κB‑Snail pathway and the subsequent process of EndMT in pulmonary arteries.

Hydrogen sulfide (H2S) functions as a protective gas transmitter in various physiological and pathological processes, but the lack of ideal donors severely hampers the clinical application of H2S. This study aims to construct a controlled release H2S donor and evaluate its protective effect on graft endothelium. Mesoporous silica nanoparticles (MSNs) were synthesized using the sol-gel method and loaded with diallyl trisulfide (DATS), an H2S-releasing agent named DATS-MSN. In vitro experiments showed that DATS-MSN could alleviate endothelial cell inflammation and enhance endothelial cell proliferation and migration. In vivo experiments demonstrated that the apoptosis of graft endothelium was mitigated in the presence of DATS-MSN. Our results indicated that DATS-MSN, releasing H2S in a controlled release fashion, could serve as an ideal H2S donor.

Hydrogen sulfide, as a novel gaseous mediator, has been suggested to play a key role in atherogenesis. However, the precise mechanisms by which H(2)S affects atherosclerosis remain unclear. Therefore, the present study aimed to investigate the potential role of H(2)S in atherosclerosis and the underlying mechanism with respect to chemokines (CCL2, CCL5 and CX3CL1) and chemokine receptors (CCR2, CCR5, and CX3CR1) in macrophages. Mouse macrophage cell line RAW 264.7 or mouse peritoneal macrophages were pre-incubated with saline or NaHS (50 µM, 100 µM, 200 µM), an H(2)S donor, and then stimulated with interferon-γ (IFN-γ) or lipopolysaccharide (LPS). It was found that NaHS dose-dependently inhibited IFN-γ or LPS-induced CX3CR1 and CX3CL1 expression, as well as CX3CR1-mediated chemotaxis in macrophages. Overexpression of cystathionine γ-lyase (CSE), an enzyme that catalyzes H(2)S biosynthesis resulted in a significant reduction in CX3CR1 and CX3CL1 expression as well as CX3CR1-mediated chemotaxis in stimulated macrophages. The inhibitory effect of H(2)S on CX3CR1 and CX3CL1 expression was mediated by modulation of proliferators-activated receptor-γ (PPAR-γ) and NF-κB pathway. Furthermore, male apoE(-/-) mice were fed a high-fat diet and then randomly given NaHS (1 mg/kg, i.p., daily) or DL-propargylglycine (PAG, 10 mg/kg, i.p., daily). NaHS significantly inhibited aortic CX3CR1 and CX3CL1 expression and impeded aortic plaque development. NaHS had a better anti-atherogenic benefit when it was applied at the early stage of atherosclerosis. However, inhibition of H(2)S formation by PAG increased aortic CX3CR1 and CX3CL1 expression and exacerbated the extent of atherosclerosis. In addition, H(2)S had minimal effect on the expression of CCL2, CCL5, CCR2 and CCR5 in vitro and in vivo. In conclusion, these data indicate that H(2)S hampers the progression of atherosclerosis in fat-fed apoE(-/-) mice and downregulates CX3CR1 and CX3CL1 expression on macrophages and in lesion plaques.

Hydrogen sulfide (H(2)S) is recognized increasingly as a proinflammatory mediator in various inflammatory conditions. Here, we have investigated the role of H(2)S in regulating expression of some endothelial adhesion molecules and recruitment of leukocytes to inflamed sites in sepsis. Male Swiss mice were subjected to cecal ligation and puncture (CLP)-induced sepsis and treated with saline (i.p.), DL-propargylglycine (PAG; 50 mg/kg, i.p.), an inhibitor of H(2)S formation or NaHS (10 mg/kg, i.p.), an H(2)S donor. PAG was administered 1 h before or after the induction of sepsis, and NaHS was given at the same time of CLP. Using intravital microcopy, we found that in sepsis, prophylactic and therapeutic administration of PAG reduced leukocyte rolling and adherence significantly in mesenteric venules coupled with decreased mRNA and protein levels of adhesion molecules (ICAM-1, P-selectin, and E-selectin) in lung and liver. In contrast, injection of NaHS up-regulated leukocyte rolling and attachment significantly, as well as tissue levels of adhesion molecules in sepsis. Conversely, normal mice were given NaHS (10 mg/kg, i.p.) to induce lung inflammation, with or without NF-kappaB inhibitor BAY 11-7082 pretreatment. NaHS treatment enhanced the level of adhesion molecules and neutrophil infiltration in lung. These alterations were reversed by pretreatment with BAY 11-7082. Moreover, expression of CXCR2 in neutrophils obtained from H(2)S-treated mice was up-regulated significantly, leading to an obvious elevation in MIP-2-directed migration of neutrophils. Therefore, H(2)S acts as an important endogenous regulator of leukocyte activation and trafficking during an inflammatory response.

Recent studies have implied that hydrogen sulfide (H2S) plays a crucial role in several inflammatory conditions. However, so far little is known about the mechanism by which H2S provokes the inflammatory response in sepsis. Thus the aim of this study was to investigate if H2S regulates sepsis-associated systemic inflammation and production of proinflammatory mediators via the activation of NF-kappaB. Male Swiss mice were subjected to cecal ligation and puncture (CLP)-induced sepsis and treated with dl-propargylglycine (PAG; 50 mg/kg ip), NaHS (10 mg/kg ip), or saline. PAG, an inhibitor of H2S formation, was administered either 1 h before or 1 h after CLP, whereas NaHS, an H2S donor, was given at the time of CLP. Some normal mice were given NaHS (10 mg/kg ip) to induce lung inflammation with or without pretreatment with the NF-kappaB inhibitor BAY 11-7082. Eight hours after CLP, both prophylactic and therapeutic administration of PAG significantly reduced the mRNA and protein levels of IL-1beta, IL-6, TNF-alpha, monocyte chemotactic protein-1, and macrophage inflammatory protein-2 in lung and liver coupled with decreased activation and translocation of NF-kappaB in lung and liver. Inhibition of H2S formation also significantly reduced lung permeability and plasma alanine aminotransferase activity. In contrast, injection of NaHS significantly aggravated sepsis-associated systemic inflammation and increased NF-kappaB activation. In addition, H2S-induced lung inflammation was blocked by BAY 11-7082. Therefore, H2S upregulates the production of proinflammatory mediators and exacerbates the systemic inflammation in sepsis through a mechanism involving NF-kappaB activation.

We have shown earlier that H(2)S acts as a mediator of inflammation. In this study, we have investigated the involvement of substance P and neurogenic inflammation in H(2)S-induced lung inflammation. Intraperitoneal administration of NaHS (1-10 mg/kg), an H(2)S donor, to mice caused a significant increase in circulating levels of substance P in a dose-dependent manner. H(2)S alone could also cause lung inflammation, as evidenced by a significant increase in lung myeloperoxidase activity and histological evidence of lung injury. The maximum effect of H(2)S on substance P levels and on lung inflammation was observed 1 h after NaHS administration. At this time, a significant increase in lung levels of TNF-alpha and IL-1beta was also observed. In substance P-deficient mice, the preprotachykinin-A knockout mice, H(2)S did not cause any lung inflammation. Furthermore, pretreatment of mice with CP-96345 (2.5 mg/kg ip), an antagonist of the neurokinin-1 (NK(1)) receptor, protected mice against lung inflammation caused by H(2)S. However, treatment with antagonists of NK(2), NK(3), and CGRP receptors did not have any effect on H(2)S-induced lung inflammation. Depleting neuropeptide from sensory neurons by capsaicin (50 mg/kg sc) significantly reduced the lung inflammation caused by H(2)S. In addition, pretreatment of mice with capsazepine (15 mg/kg sc), an antagonist of the transient receptor potential vanilloid-1, protected mice against H(2)S-induced lung inflammation. These results demonstrate a key role of substance P and neurogenic inflammation in H(2)S-induced lung injury in mice.

Endogenous hydrogen sulfide (H(2)S) is naturally synthesized in various types of mammalian cells from l-cysteine in a reaction catalyzed by two enzymes, cystathionine-gamma-lyase (CSE) and/or cystathionine-beta-synthase. The latest studies have implied that H(2)S functions as a vasodilator and neurotransmitter. However, so far there is little information about the role played by H(2)S in systemic inflammation such as sepsis. Thus the aim of this study was to investigate the potential role of endogenous H(2)S in cecal ligation and puncture (CLP)-induced sepsis. Male Swiss mice were subjected to CLP-induced sepsis and treated with saline (ip), dl-propargylglycine (PAG, 50 mg/kg ip), a CSE inhibitor, or sodium hydrosulfide (NaHS; 10 mg/kg ip). PAG was administered either 1 h before or 1 h after the induction of sepsis, whereas NaHS was given at the same time of CLP. CLP-induced sepsis significantly increased the plasma H(2)S level and the liver H(2)S synthesis 8 h after CLP compared with sham operation. Induction of sepsis also resulted in a significant upregulation of CSE mRNA in liver. On the other hand, prophylactic as well as therapeutic administration of PAG significantly reduced sepsis-associated systemic inflammation, as evidenced by myeloperoxidase activity and histological changes in lung and liver, and attenuated the mortality of CLP-induced sepsis. Injection of NaHS significantly aggravated sepsis-associated systemic inflammation. Therefore, the effect of inhibition of H(2)S formation and administration of NaHS suggests that H(2)S plays a proinflammatory role in regulating the severity of sepsis and associated organ injury.

Animal production units produce and store many contaminants on-site, including organic dust (OD) and hydrogen sulfide (H2S). Workers in these settings report various respiratory disease symptoms. Both OD and H2S have shown to induce lung inflammation. However, impact of co-exposure to both H2S and OD has not been investigated. Therefore, we tested a hypothesis that pre-exposure to H2S modulates the innate inflammatory response of the lungs to organic dust. In a mouse model of H2S and organic dust extract (ODE) exposure, we assessed lung inflammation quantitatively. We exposed human airway epithelial and monocytic cells to medium or H2S alone or H2S followed by ODE and measured cell viability, oxidative stress, and other markers of inflammation. Exposure to 10 ppm H2S followed by ODE increased the lavage fluid leukocytes. However, exposure to 10 ppm H2S alone resulted in changes in tight junction proteins, an increase in mRNA levels of tlr2 and tlr4 as well as ncf1, ncf4, hif1α, and nrf2. H2S alone or H2S and ODE exposure decreased cell viability and increased reactive nitrogen species production. ODE exposure increased the transcripts of tlr2 and tlr4 in both in vitro and in vivo models, whereas increased nfkbp65 transcripts following exposure to ODE and H2S was seen only in in vitro model. H2S alone and H2S followed by ODE exposure increased the levels of IL-1β. We conclude that pre-exposure to H2S modulates lung innate inflammatory response to ODE.