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Introduction: Myocardial ischemia/reperfusion (I/R) injury remains a major obstacle in cardiovascular therapies. Hydrogen sulfide (H2S) shows promise for mitigating I/R injury, but conventional delivery methods, such as NaHS injections or adenovirus-mediated CSE gene therapy, face low efficiency and systemic side effects. This study explores the use of angiotensin 1 (AT1) peptide-conjugated CdSe/ZnS quantum dots (QDs) for targeted delivery of cystathionine-γ-lyase (CSE) plasmids to the myocardium, aiming to boost local H2S production and minimize I/R injury. Methods: CdSe/ZnS QDs were conjugated with AT1 peptides to create a nanocarrier system capable of delivering the CSE plasmid specifically to the myocardium. In vivo fluorescence imaging confirmed heart-specific accumulation. Myocardial infarct size, cardiac function, cell death, and oxidative stress were evaluated. Endoplasmic reticulum stress and mitophagy markers, including CHOP/GRP78/eIF2α, were analyzed, and the CHOP gene's role was further assessed using an adenovirus vector. Results: The AT1-conjugated nanocarriers significantly increased CSE expression in the myocardium, as confirmed by fluorescence imaging, without affecting other organs. This localized delivery reduced myocardial infarct size, improved cardiac function, and decreased oxidative stress and cell death. Importantly, a reduction in endoplasmic reticulum stress and mitophagy markers was observed, suggesting that cardioprotection was mediated via the CHOP/GRP78/eIF2α signaling pathway. Reintroduction of CHOP using an adenovirus vector reversed these protective effects, confirming the pathway's involvement. Discussion: This study demonstrates that AT1 peptide-conjugated QDs can effectively deliver CSE plasmids to the heart, providing significant protection against I/R injury through enhanced localized H2S production. This approach offers a promising, targeted, and side-effect-free therapy for myocardial I/R injury, with potential for clinical translation. [ABSTRACT FROM AUTHOR]

Early life stress (ELS) significantly increases the risk of chronic cardiovascular diseases and may cause neuroinflammation. This post hoc study, based on the material available from a previous study showing elevated "serum brain injury markers" in male control animals, examines the effect of sex and/or ELS on the cerebral and cardiac expression of the H2S and oxytocin systems. Following approval by the Regional Council of Tübingen, a randomized controlled study was conducted on 12 sexually mature, uncastrated German Large White swine of both sexes. The control animals were separated from their mothers at 28–35 days, while the ELS group was separated at day 21. At 20–24 weeks, animals underwent anesthesia, ventilation, and surgical instrumentation. An immunohistochemical analysis of oxytocin, its receptor, and the H2S-producing enzymes cystathionine-β-synthase and cystathionine-γ-lyase was performed on hypothalamic, prefrontal cortex, and myocardial tissue samples. Data are expressed as the % of positive tissue staining, and differences between groups were tested using a two-way ANOVA. The results showed no significant differences in the oxytocin and H2S systems between groups; however, sex influenced the oxytocin system, and ELS affected the oxytocin and H2S systems in a sex-specific manner. No immunohistochemical correlate to the elevated "serum brain injury markers" in male controls was identified. [ABSTRACT FROM AUTHOR]

Introduction: Myocardial ischemia/reperfusion (I/R) injury remains a major obstacle in cardiovascular therapies. Hydrogen sulfide (H2S) shows promise for mitigating I/R injury, but conventional delivery methods, such as NaHS injections or adenovirus-mediated CSE gene therapy, face low efficiency and systemic side effects. This study explores the use of angiotensin 1 (AT1) peptideconjugated CdSe/ZnS quantum dots (QDs) for targeted delivery of cystathionine-γ-lyase (CSE) plasmids to the myocardium, aiming to boost local H2S production and minimize I/R injury. Methods: CdSe/ZnS QDs were conjugated with AT1 peptides to create a nanocarrier system capable of delivering the CSE plasmid specifically to the myocardium. In vivo fluorescence imaging confirmed heart-specific accumulation. Myocardial infarct size, cardiac function, cell death, and oxidative stress were evaluated. Endoplasmic reticulum stress and mitophagy markers, including CHOP/GRP78/eIF2a, were analyzed, and the CHOP gene's role was further assessed using an adenovirus vector. Results: The AT1-conjugated nanocarriers significantly increased CSE expression in the myocardium, as confirmed by fluorescence imaging, without affecting other organs. This localized delivery reduced myocardial infarct size, improved cardiac function, and decreased oxidative stress and cell death. Importantly, a reduction in endoplasmic reticulum stress and mitophagy markers was observed, suggesting that cardioprotection was mediated via the CHOP/GRP78/eIF2a signaling pathway. Reintroduction of CHOP using an adenovirus vector reversed these protective effects, confirming the pathway's involvement. Discussion: This study demonstrates that AT1 peptide-conjugated QDs can effectively deliver CSE plasmids to the heart, providing significant protection against I/R injury through enhanced localized H2S production. This approach offers a promising, targeted, and side-effect-free therapy for myocardial I/R injury, with potential for clinical translation. [ABSTRACT FROM AUTHOR]

Epidemiological data suggest that moderate hyperoxemia may be associated with an improved outcome after traumatic brain injury. In a prospective, randomized investigation of long-term, resuscitated acute subdural hematoma plus hemorrhagic shock (ASDH + HS) in 14 adult, human-sized pigs, targeted hyperoxemia (200 < PaO2 < 250 mmHg vs. normoxemia 80 < PaO2 < 120 mmHg) coincided with improved neurological function. Since brain perfusion, oxygenation and metabolism did not differ, this post hoc study analyzed the available material for the effects of targeted hyperoxemia on cerebral tissue markers of oxidative/nitrosative stress (nitrotyrosine expression), blood–brain barrier integrity (extravascular albumin accumulation) and fluid homeostasis (oxytocin, its receptor and the H2S-producing enzymes cystathionine-β-synthase and cystathionine-γ-lyase). After 2 h of ASDH + HS (0.1 mL/kgBW autologous blood injected into the subdural space and passive removal of 30% of the blood volume), animals were resuscitated for up to 53 h by re-transfusion of shed blood, noradrenaline infusion to maintain cerebral perfusion pressure at baseline levels and hyper-/normoxemia during the first 24 h. Immediate postmortem, bi-hemispheric (i.e., blood-injected and contra-lateral) prefrontal cortex specimens from the base of the sulci underwent immunohistochemistry (% positive tissue staining) analysis of oxidative/nitrosative stress, blood–brain barrier integrity and fluid homeostasis. None of these tissue markers explained any differences in hyperoxemia-related neurological function. Likewise, hyperoxemia exerted no deleterious effects. [ABSTRACT FROM AUTHOR]

Early life stress (ELS) significantly increases the risk of chronic cardiovascular diseases and may cause neuroinflammation. This post hoc study, based on the material available from a previous study showing elevated “serum brain injury markers” in male control animals, examines the effect of sex and/or ELS on the cerebral and cardiac expression of the H2S and oxytocin systems. Following approval by the Regional Council of Tübingen, a randomized controlled study was conducted on 12 sexually mature, uncastrated German Large White swine of both sexes. The control animals were separated from their mothers at 28–35 days, while the ELS group was separated at day 21. At 20–24 weeks, animals underwent anesthesia, ventilation, and surgical instrumentation. An immunohistochemical analysis of oxytocin, its receptor, and the H2S-producing enzymes cystathionine-β-synthase and cystathionine-γ-lyase was performed on hypothalamic, prefrontal cortex, and myocardial tissue samples. Data are expressed as the % of positive tissue staining, and differences between groups were tested using a two-way ANOVA. The results showed no significant differences in the oxytocin and H2S systems between groups; however, sex influenced the oxytocin system, and ELS affected the oxytocin and H2S systems in a sex-specific manner. No immunohistochemical correlate to the elevated “serum brain injury markers” in male controls was identified.

Objectives: Cystathionine‐γ‐lyase (CTH) has been proved to involve in inflammation and bone remolding, implying its potential role in the progression of periodontitis. This study was aimed to investigate the function of CTH and its relation to the macrophage polarization in periodontitis. Materials and methods: Bone marrow‐derived macrophages (BMDMs) from wild‐type (WT) and Cth knockout (Cth−/−) mice were stimulated with lipopolysaccharide (LPS) in vitro and pro‐inflammatory cytokines were analyzed by qRT‐PCR. Ligature‐induced periodontitis was established on WT and Cth−/− mice. Histological analysis, tartrate‐resistant acid phosphatase staining, immunostaining, and Western blot were performed to analyze the periodontium destruction and M1 macrophage polarization. Results: Cth expression in BMDMs was upregulated upon increasing LPS stimulation. Deletion of Cth suppressed BMDMs inflammatory response with decreased Il1b, Il6, and Tnf mRNA. Cth−/− mice with periodontitis showed attenuated bone loss and impaired osteoclast differentiation compared with WT. Moreover, Cth knockout hindered M1 macrophage polarization, reduced the expression of IL‐1β, IL‐6, and TNF‐α in periodontally diseased tissue. Conclusion: This study demonstrated that CTH played an important role in regulating the inflammatory responses and periodontitis tissue destruction. Importantly, Cth knockout suppressed M1 macrophages polarization in periodontitis. [ABSTRACT FROM AUTHOR]

The saphenous vein is the conduit of choice for bypass grafting. Unfortunately, the hemodynamic stress associated with the arterial environment of the bypass vein graft leads to the development of intimal hyperplasia (IH), an excessive cellular growth and collagen deposition that results in restenosis and secondary graft occlusion. Hydrogen sulfide (H2S) is a ubiquitous redox-modifying gasotransmitter that inhibits IH. H2S is produced via the reverse trans-sulfuration pathway by three enzymes: cystathionine γ-lyase (CSE), cystathionine β-synthase (CBS) and 3-mercaptopyruvate sulfurtransferase (3-MST). However, the expression and regulation of these enzymes in the human vasculature remains unclear. Here, we investigated the expression of CSE, CBS and 3-MST in segments of native human saphenous vein and large arteries. Furthermore, we evaluated the regulation of these enzymes in vein segments cultured under static, venous (7 mmHg pressure) or arterial (100 mmHg pressure) pressure. CSE was expressed in the media, neointima and intima of the vessels and was negatively regulated by arterial shear stress. Adenoviral-mediated CSE overexpression or RNA interference-mediated CSE knock-down revealed that CSE inhibited primary human VSMC migration but not proliferation. We propose that high shear stress in arteriovenous bypass grafts inhibits CSE expression in both the media and endothelium, which may contribute to increased VSMC migration in the context of IH. [ABSTRACT FROM AUTHOR]

Hydrogen sulfide (H2S) is one of the most important gasotransmitters that affect lifespan and provide resistance to adverse environmental conditions. Here we investigated geroprotective effects of the individual and simultaneous overexpression of genes encoding key enzymes of H2S biosynthesis – cystathionine-β-synthase (CBS) and cystathionine-γ-lyase (CSE) on D. melanogaster model. Simultaneous overexpression of CBS and CSE resulted in additive (in males) and synergistic (in females) beneficial effects on median lifespan. Individual overexpression of CBS was associated with increased thermotolerance and decreased transcription level of genes encoding stress-responsive transcription factors HIF1 and Hsf, while individual overexpression of CSE was associated with increased resistance to paraquat. Simultaneous overexpression of both genes increased resistance to hyperthermia in old females or paraquat in old males. The obtained results suggest sex-specific epistatic interaction of CBS and CSE overexpression effects on longevity and stress resistance. [ABSTRACT FROM AUTHOR]

Cystathionine-γ-lyase (CSE) is expressed in various tissues and generates H2S via an alternative desulfuration reaction. We sought to explore the functions of skeletal muscle CSE using skeletal muscle conditional knockout CSE (MCSEKO) mice. It was found that body weight, muscle morphology, and exercise capacity were not altered in MCSEKO mice compared with littermate wild-type mice. RNA-seq-based transcriptome analysis showed that 275 genes were differentially regulated in skeletal muscle and multiple signaling pathways including insulin signaling and mTOR, PI3K-AKT, and cGMP-PKG signaling pathways were enriched in MCSEKO mice. The intraperitoneal glucose tolerance test and insulin tolerance test showed that glucose tolerance and insulin sensitivity were reduced in MCSEKO mice. Glucose transporter 4 (GLU4) and PKG-1 expression levels and insulin receptor substrate-1(IRS1)/PI3K/Akt signaling pathway were downregulated whilst the mTOR/S6K/S6 pathway was enhanced in MCSEKO mice. These effects were reversed by the H2S supplement. Aerobic treadmill training significantly promoted glucose tolerance and insulin sensitivity and improved GLU4 and PKG-1 levels, promoted IRS1/PI3K/Akt signaling and suppressed mTOR/S6K/S6 signaling pathway in MCSEKO mice. Our data suggest that skeletal muscle CSE/H2S signaling is critical for the maintenance of insulin sensitivity, which is associated with maintaining the balance in PKG, PI3K/Akt, and mTOR/S6K/S6 signaling pathways in skeletal muscle. [ABSTRACT FROM AUTHOR]

The saphenous vein is the conduit of choice for bypass grafting. Unfortunately, the hemodynamic stress associated with the arterial environment of the bypass vein graft leads to the development of intimal hyperplasia (IH), an excessive cellular growth and collagen deposition that results in restenosis and secondary graft occlusion. Hydrogen sulfide (H2S) is a ubiquitous redox-modifying gasotransmitter that inhibits IH. H2S is produced via the reverse trans-sulfuration pathway by three enzymes: cystathionine γ-lyase (CSE), cystathionine β-synthase (CBS) and 3-mercaptopyruvate sulfurtransferase (3-MST). However, the expression and regulation of these enzymes in the human vasculature remains unclear. Here, we investigated the expression of CSE, CBS and 3-MST in segments of native human saphenous vein and large arteries. Furthermore, we evaluated the regulation of these enzymes in vein segments cultured under static, venous (7 mmHg pressure) or arterial (100 mmHg pressure) pressure. CSE was expressed in the media, neointima and intima of the vessels and was negatively regulated by arterial shear stress. Adenoviral-mediated CSE overexpression or RNA interference-mediated CSE knock-down revealed that CSE inhibited primary human VSMC migration but not proliferation. We propose that high shear stress in arteriovenous bypass grafts inhibits CSE expression in both the media and endothelium, which may contribute to increased VSMC migration in the context of IH.

Hydrogen sulfide (H2S) is a gasotransmitter endogenously synthesized by cystathionine-γ-lyase (CSE), cystathionine-β-synthase (CBS), and 3-mercaptopiruvate sulfurtransferase (3-MST) enzymes. H2S exogenous administration prevents the development of hemodynamic impairments after traumatic brain injury (TBI). Since the hypothalamus and the brainstem highly regulate the cardiovascular system, this study aimed to evaluate the effect of NaHS subchronic treatment on the changes of H2S-sythesizing enzymes in those brain areas after TBI and in physiological conditions. For that purpose, animals were submitted to a lateral fluid percussion injury, and the changes in CBS, CSE, and 3-MST protein expression were measured by western blot at days 1, 2, 3, 7, and 28 in the vehicle group, and 7 and 28 days after NaHS treatment. After severe TBI induction, we found a decrease in CBS and CSE protein expression in the hypothalamus and brainstem; meanwhile, 3-MST protein expression diminished only in the hypothalamus compared to the Sham group. Remarkably, i.p. daily injections of NaHS, an H2S donor, (3.1 mg/kg) during seven days: (1) restored CBS and CSE but no 3-MST protein expression in the hypothalamus at day 28 post-TBI; (2) reestablished only CSE in brainstem 7 and 28 days after TBI; and (3) did not modify H2S-sythesizing enzymes protein expression in uninjured animals. Mainly, our results show that the NaHS effect on CBS and CSE protein expression is observed in a time- and tissue-dependent manner with no effect on 3-MST expression, which may suggest a potential role of H2S synthesis in hypothalamus and brainstem impairments observed after TBI. [ABSTRACT FROM AUTHOR]

Simple Summary: Endogenous hydrogen sulfide (H2S) has been implicated in many physiological and pathological processes, particularly in inflammatory responses and adaptive immunity. In this study, we identified the candidate differentially expressed proteins (DEPs) associated with H2S metabolism in Holstein cows with clinical mastitis (CM). The results revealed 17 DEPs included in 44 Gene Ontology (GO) terms and five Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways related to sulfide metabolism and indicated the important role of cystathionine-γ-lyase (CTH)/H2S in CM. Our findings can support research into the function and regulatory mechanism of CTH/H2S in Holstein cows and provide a basis for the prevention and treatment of CM. H2S plays an important role in various inflammatory diseases. However, the role of H2S and synthetic enzymes in Holstein cows with CM is unknown. The aim of this study was to identify DEPs associated with sulfide metabolism and further investigate their roles in dairy cows with CM. From 3739 DEPs generated by data-independent acquisition proteomics, we identified a total of 17 DEPs included in 44 GO terms and five KEGG pathways related to sulfide metabolism, including CTH and cystathionine-β-synthase (CBS). Immunohistochemical and immunofluorescence staining results showed that CTH and CBS proteins were present mainly in the cytoplasm of mammary epithelial cells. Endogenous H2S production in the serum of the CM group was significantly lower than that of the healthy Holstein cows. CTH and CBS mRNA and protein levels in the mammary glands of the CM group were significantly downregulated compared to those of the healthy group. These results indicate that CTH and H2S were correlated with the occurrence and development of CM in Holstein cows, which provides important insights into the function and regulatory mechanism of CTH/H2S in Holstein cows. [ABSTRACT FROM AUTHOR]

Sarcopenia is a gradual and generalized skeletal muscle (SKM) syndrome, characterized by the impairment of muscle components and functionality. Hydrogen sulfide (H2S), endogenously formed within the body from the activity of cystathionine-γ-lyase (CSE), cystathionine- β-synthase (CBS), and mercaptopyruvate sulfurtransferase, is involved in SKM function. Here, in an in vitro model of sarcopenia based on damage induced by dexamethasone (DEX, 1 μM, 48 h treatment) in C2C12-derived myotubes, we investigated the protective potential of exogenous and endogenous sources of H2S, i.e., glucoraphanin (30 μM), L-cysteine (150 μM), and 3-mercaptopyruvate (150 μM). DEX impaired the H2S signalling in terms of a reduction in CBS and CSE expression and H2S biosynthesis. Glucoraphanin and 3-mercaptopyruvate but not L-cysteine prevented the apoptotic process induced by DEX. In parallel, the H2S-releasing molecules reduced the oxidative unbalance evoked by DEX, reducing catalase activity, O2− levels, and protein carbonylation. Glucoraphanin, 3-mercaptopyruvate, and L-cysteine avoided the changes in myotubes morphology and morphometrics after DEX treatment. In conclusion, in an in vitro model of sarcopenia, an impairment in CBS/CSE/H2S signalling occurs, whereas glucoraphanin, a natural H2S-releasing molecule, appears more effective for preventing the SKM damage. Therefore, glucoraphanin supplementation could be an innovative therapeutic approach in the management of sarcopenia. [ABSTRACT FROM AUTHOR]

In the porcine model discussed in this review, the acute subdural hematoma was induced by subdural injection of autologous blood over the left parietal cortex, which led to a transient elevation of the intracerebral pressure, measured by bilateral neuromonitoring. The hematoma-induced brain injury was associated with albumin extravasation, oxidative stress, reactive astrogliosis and microglial activation in the ipsilateral hemisphere. Further proteins and injury markers were validated to be used for immunohistochemistry of porcine brain tissue. The cerebral expression patterns of oxytocin, oxytocin receptor, cystathionine-γ-lyase and cystathionine-β-synthase were particularly interesting: these four proteins all co-localized at the base of the sulci, where pressure-induced brain injury elicits maximum stress. In this context, the pig is a very relevant translational model in contrast to the rodent brain. The structure of the porcine brain is very similar to the human: the presence of gyri and sulci (gyrencephalic brain), white matter to grey matter proportion and tentorium cerebelli. Thus, pressure-induced injury in the porcine brain, unlike in the rodent brain, is reflective of the human pathophysiology.

BackgroundThe hydrogen sulfide (H2S) and the oxytocin/oxytocin receptor (OT/OTR) systems interact in the central nervous and cardiovascular system. As a consequence of osmotic balance stress, H2S stimulates OT release from the paraventricular nuclei (PVN) in the hypothalamic regulation of blood volume and pressure. Hemorrhagic shock (HS) represents one of the most pronounced acute changes in blood volume, which, moreover, may cause at least transient brain tissue hypoxia. Atherosclerosis is associated with reduced vascular expression of the main endogenous H2S producing enzyme cystathionine-γ-lyase (CSE), and, hence, exogenous H2S administration could be beneficial in these patients, in particular after HS. However, so far cerebral effects of systemic H2S administration are poorly understood. Having previously shown lung-protective effects of therapeutic Na2S2O3 administration in a clinically relevant, long-term, porcine model of HS and resuscitation we evaluated if these protective effects were extended to the brain.MethodsIn this study, available unanalyzed paraffin embedded brain sections (Na2S2O3N = 8 or vehicle N = 5) of our recently published HS study were analyzed via neuro-histopathology and immunohistochemistry for the endogenous H2S producing enzymes, OT, OTR, and markers for brain injury and oxidative stress (glial fibrillary acidic protein (GFAP) and nitrotyrosine).ResultsNeuro-histopathological analysis revealed uninjured brain tissue with minor white matter edema. Protein quantification in the hypothalamic PVN showed no significant inter-group differences between vehicle or Na2S2O3 treatment.ConclusionsThe endogenous H2S enzymes, OT/OTR co-localized in magnocellular neurons in the hypothalamus, which may reflect their interaction in response to HS-induced hypovolemia. The preserved blood brain barrier (BBB) may have resulted in impermeability for Na2S2O3 and no inter-group differences in the PVN. Nonetheless, our results do not preclude that Na2S2O3 could have a therapeutic benefit in the brain in an injury that disrupts the BBB, e.g., traumatic brain injury (TBI) or acute subdural hematoma (ASDH).

Significance: Hydrogen sulfide (H2S) plays critical roles in redox biology, and its regulatory effects are tightly controlled by its cellular location and concentration. The imbalance of H2S is believed to contribute to some pathological processes. Recent Advances: Downregulation of H2S requires chemical tools such as inhibitors of H2S-producing enzymes and H2S scavengers. Recent efforts have discovered some promising inhibitors and scavengers. These advances pave the road toward better understanding of the functions of H2S. Critical Issues: Precise H2S downregulation is challenging. The potency and specificity of current inhibitors are still far from ideal. H2S-producing enzymes are involved in complex sulfur metabolic pathways and ubiquitously present in biological matrices. The inhibition of these enzymes can cause unwanted side effects. H2S scavengers allow targeted H2S clearance, but their options are still limited. In addition, the scavenging process often results in biologically active by-products. Future Directions: Further development of potent and specific inhibitors for H2S-producing enzymes is needed. Scavengers that can rapidly and selectively remove H2S while generating biocompatible by-products are needed. Potential therapeutic applications of scavengers and inhibitors are worth exploring. Antioxid. Redox Signal. 36, 294–308. [ABSTRACT FROM AUTHOR]

We have previously shown that diallyl disulfide (DADS) protects mice against cerulein-induced acute pancreatitis (AP) and associated lung injury. However, the molecular mechanisms underlying its effect and the components involved have not been studied. We hypothesized that DADS may reduce TNF-α, CSE expression, H2S production, STAT3, and NF-κB activation and induce SOCS3 expression through peroxisome proliferator-activated receptor γ (PPAR-γ) pathway in cerulein-induced mice. Male Swiss mice were treated with hourly intraperitoneal injections of cerulein (50 µg/kg) for 6 h. Diallyl disulfide (200 μg/kg) was administered in the presence or absence of PPAR-γ antagonist GW9662 (0.3 mg/kg) (i.p) 1 h after the induction of AP. Our findings revealed that DADS blocked TNF-α, CSE expression, H2S production, and STAT3, and NF-κB activation was reversed by GW9662. Furthermore, GW9662 abrogated DADS-induced SOCS3 expression. The results show for the first that DADS-induced anti-inflammatory effect in acute pancreatitis is regulated through PPAR-γ. [ABSTRACT FROM AUTHOR]

Cystathionine-γ-lyase (CSE) is expressed in various tissues and generates H2S via an alternative desulfuration reaction. We sought to explore the functions of skeletal muscle CSE using skeletal muscle conditional knockout CSE (MCSEKO) mice. It was found that body weight, muscle morphology, and exercise capacity were not altered in MCSEKO mice compared with littermate wild-type mice. RNA-seq-based transcriptome analysis showed that 275 genes were differentially regulated in skeletal muscle and multiple signaling pathways including insulin signaling and mTOR, PI3K-AKT, and cGMP-PKG signaling pathways were enriched in MCSEKO mice. The intraperitoneal glucose tolerance test and insulin tolerance test showed that glucose tolerance and insulin sensitivity were reduced in MCSEKO mice. Glucose transporter 4 (GLU4) and PKG-1 expression levels and insulin receptor substrate-1(IRS1)/PI3K/Akt signaling pathway were downregulated whilst the mTOR/S6K/S6 pathway was enhanced in MCSEKO mice. These effects were reversed by the H2S supplement. Aerobic treadmill training significantly promoted glucose tolerance and insulin sensitivity and improved GLU4 and PKG-1 levels, promoted IRS1/PI3K/Akt signaling and suppressed mTOR/S6K/S6 signaling pathway in MCSEKO mice. Our data suggest that skeletal muscle CSE/H2S signaling is critical for the maintenance of insulin sensitivity, which is associated with maintaining the balance in PKG, PI3K/Akt, and mTOR/S6K/S6 signaling pathways in skeletal muscle.

Pathogenic fungi are recognized as a progressive threat to humans, particularly those with the immunocompromised condition. The growth of fungi is controlled by several factors, one of which is signaling molecules, such as hydrogen sulfide (H2S), which was traditionally regarded as a toxic gas without physiological function. However, recent studies have revealed that H2S is produced enzymatically and endogenously in several species, where it serves as a gaseous signaling molecule performing a variety of critical biological functions. However, the influence of this endogenous H2S on the biological activities occurring within the pathogenic fungi, such as transcriptomic and phenotypic alternations, has not been elucidated so far. Therefore, the present study was aimed to decipher this concern by utilizing S-propargyl-cysteine (SPRC) as a novel and stable donor of H2S and Saccharomyces cerevisiae as a fungal model. The results revealed that the yeast could produce H2S by catabolizing SPRC, which facilitated the growth of the yeast cells. This implies that the additional intracellularly generated H2S is generated primarily from the enhanced sulfur-amino-acid-biosynthesis pathways and serves to increase the growth rate of the yeast, and presumably the growth of the other fungi as well. In addition, by deciphering the implicated pathways and analyzing the in vitro enzymatic activities, cystathionine-γ-lyase (CYS3) was identified as the enzyme responsible for catabolizing SPRC into H2S in the yeast, which suggested that cystathionine-γ-lyase might play a significant role in the regulation of H2S-related transcriptomic and phenotypic alterations occurring in yeast. These findings provide important information regarding the mechanism underlying the influence of the gaseous signaling molecules such as H2S on fungal growth. In addition, the findings provide a better insight to the in vivo metabolism of H2S-related drugs, which would be useful for the future development of anti-fungal drugs.

Pathogenic fungi are recognized as a progressive threat to humans, particularly those with the immunocompromised condition. The growth of fungi is controlled by several factors, one of which is signaling molecules, such as hydrogen sulfide (H2S), which was traditionally regarded as a toxic gas without physiological function. However, recent studies have revealed that H2S is produced enzymatically and endogenously in several species, where it serves as a gaseous signaling molecule performing a variety of critical biological functions. However, the influence of this endogenous H2S on the biological activities occurring within the pathogenic fungi, such as transcriptomic and phenotypic alternations, has not been elucidated so far. Therefore, the present study was aimed to decipher this concern by utilizing S -propargyl-cysteine (SPRC) as a novel and stable donor of H2S and Saccharomyces cerevisiae as a fungal model. The results revealed that the yeast could produce H2S by catabolizing SPRC, which facilitated the growth of the yeast cells. This implies that the additional intracellularly generated H2S is generated primarily from the enhanced sulfur-amino-acid-biosynthesis pathways and serves to increase the growth rate of the yeast, and presumably the growth of the other fungi as well. In addition, by deciphering the implicated pathways and analyzing the in vitro enzymatic activities, cystathionine-γ-lyase (CYS3) was identified as the enzyme responsible for catabolizing SPRC into H2S in the yeast, which suggested that cystathionine-γ-lyase might play a significant role in the regulation of H2S-related transcriptomic and phenotypic alterations occurring in yeast. These findings provide important information regarding the mechanism underlying the influence of the gaseous signaling molecules such as H2S on fungal growth. In addition, the findings provide a better insight to the in vivo metabolism of H2S-related drugs, which would be useful for the future development of anti-fungal drugs. [ABSTRACT FROM AUTHOR]

H2S plays an important role in various inflammatory diseases. However, the role of H2S and synthetic enzymes in Holstein cows with CM is unknown. The aim of this study was to identify DEPs associated with sulfide metabolism and further investigate their roles in dairy cows with CM. From 3739 DEPs generated by data-independent acquisition proteomics, we identified a total of 17 DEPs included in 44 GO terms and five KEGG pathways related to sulfide metabolism, including CTH and cystathionine-β-synthase (CBS). Immunohistochemical and immunofluorescence staining results showed that CTH and CBS proteins were present mainly in the cytoplasm of mammary epithelial cells. Endogenous H2S production in the serum of the CM group was significantly lower than that of the healthy Holstein cows. CTH and CBS mRNA and protein levels in the mammary glands of the CM group were significantly downregulated compared to those of the healthy group. These results indicate that CTH and H2S were correlated with the occurrence and development of CM in Holstein cows, which provides important insights into the function and regulatory mechanism of CTH/H2S in Holstein cows.

Sp1-CSE-H 2 S pathway plays an important role in homocysteine-metabolism, whose disorder can result in hyperhomocysteinemia. H 2 S deficiency in hyperhomocysteinemia has been reported, while the underlying mechanism and whether it in turn affects the progress of hyperhomocysteinemia are unclear. This study focused on the post-translational modification of Sp1/CSE and revealed four major findings: (1) Homocysteine-accumulation augmented CSE's nitration, inhibited its bio-activity, thus caused H 2 S deficiency. (2) H 2 S deficiency inhibited the S-sulfhydration of Sp1, down-regulated CSE and decreased H 2 S further, which in turn weakened CSE's own S-sulfhydration. (3) CSE was S-sulfhydrated at Cys84, Cys109, Cys172, Cys229, Cys252, Cys307 and Cys310, among which the S-sulfhydration of Cys172 and Cys310 didn't affect its enzymatic activity, while the S-sulfhydration of Cys84, Cys109, Cys229, Cys252 and Cys307 was necessary for its bio-activity. (4) H 2 S deficiency trapped homocysteine-metabolism into a vicious cycle, which could be broken by either blocking nitration or restoring S-sulfhydration. This study detected a new mechanism that caused severe hyperhomocysteinemia, thereby provided new therapeutic strategies for hyperhomocysteinemia. Image 1 • Hcy augmented the nitration of CSE, blunted its bio-activity and caused H 2 S deficiency. • Deficiency of Sp1's S-sulfhydration down-regulated CSE, lowered H 2 S level further, which weakened CSE's S-sulfhydration. • CSE was S-sulfhydrated at Cys84, Cys109, Cys172, Cys229, Cys252, Cys307 and Cys310 under physiological conditions. • H 2 S deficiency trapped HHcy into a vicious cycle, blocking nitration or restoring S-sulfhydration could break this cycle. [ABSTRACT FROM AUTHOR]

The pathophysiology of sepsis-induced myocardial dysfunction is not resolved to date and comprises inflammation, barrier dysfunction and oxidative stress. Disease-associated reduction of tissue cystathionine-γ-lyase (CSE) expression, an endogenous H2S-producing enzyme, is associated with oxidative stress, barrier dysfunction and organ injury. CSE-mediated cardio-protection has been suggested to be related the upregulation of oxytocin receptor (OTR). CSE can also mediate glucocorticoid receptor (GR) signaling, which is important for normal heart function. A sepsis-related loss of cardiac CSE expression associated with impaired organ function has been reported previously. The aim of this current post hoc study was to investigate the role of cardiac GR and OTR after polymicrobial sepsis in a clinically relevant, resuscitated, atherosclerotic porcine model. Anesthetized and instrumented FBM (Familial Hypercholesterolemia Bretoncelles Meishan) pigs with high fat diet-induced atherosclerosis underwent poly-microbial septic shock (n = 8) or sham procedure (n = 5), and subsequently received intensive care therapy with fluid and noradrenaline administration for 24 h. Cardiac protein expression and mRNA levels were analyzed. Systemic troponin, a marker of cardiac injury, was significantly increased in septic animals in contrast to sham, whereas OTR and GR expression in septic hearts were reduced, along with a down-regulation of anti-inflammatory GR target genes and the antioxidant transcription factor NRF2. These results suggest a potential interplay between GR, CSE, and OTR in sepsis-mediated oxidative stress, inflammation and cardiac dysfunction.

Abstract Background Both the hydrogen sulfide/cystathionine-γ-lyase (H2S/CSE) and oxytocin/oxytocin receptor (OT/OTR) systems have been reported to be cardioprotective. H2S can stimulate OT release, thereby affecting blood volume and pressure regulation. Systemic hyper-inflammation after blunt chest trauma is enhanced in cigarette smoke (CS)-exposed CSE−/− mice compared to wildtype (WT). CS increases myometrial OTR expression, but to this point, no data are available on the effects CS exposure on the cardiac OT/OTR system. Since a contusion of the thorax (Txt) can cause myocardial injury, the aim of this post hoc study was to investigate the effects of CSE−/− and exogenous administration of GYY4137 (a slow release H2S releasing compound) on OTR expression in the heart, after acute on chronic disease, of CS exposed mice undergoing Txt. Methods This study is a post hoc analysis of material obtained in wild type (WT) homozygous CSE−/− mice after 2-3 weeks of CS exposure and subsequent anesthesia, blast wave-induced TxT, and surgical instrumentation for mechanical ventilation (MV) and hemodynamic monitoring. CSE−/− animals received a 50 μg/g GYY4137-bolus after TxT. After 4h of MV, animals were exsanguinated and organs were harvested. The heart was cut transversally, formalin-fixed, and paraffin-embedded. Immunohistochemistry for OTR, arginine-vasopressin-receptor (AVPR), and vascular endothelial growth factor (VEGF) was performed with naïve animals as native controls. Results CSE−/− was associated with hypertension and lower blood glucose levels, partially and significantly restored by GYY4137 treatment, respectively. Myocardial OTR expression was reduced upon injury, and this was aggravated in CSE−/−. Exogenous H2S administration restored myocardial protein expression to WT levels. Conclusions This study suggests that cardiac CSE regulates cardiac OTR expression, and this effect might play a role in the regulation of cardiovascular function.

Background/Aims: Sepsis is a severe and complicated syndrome that is characterized by dysregulation of host inflammatory responses and organ failure. Cystathionine-γ-lyase (CSE)/ hydrogen sulfide (H2S) has potential anti-inflammatory activities in a variety of inflammatory diseases. NADPH oxidase 4 (Nox4), a member of the NADPH oxidases, is the major source of reactive oxygen species (ROS) and its expression is increased in sepsis, but its function in CSE-mediated anti-inflammatory activities remains unknown. Methods: Macrophages were either transfected with CSE, Nox4 siRNA or transduced with lentiviral vector encoding CSE or Nox4, and then stimulated with lipopolysaccharide (LPS). The expression of inflammatory mediators and signaling pathway activation were measured by quantitative PCR (qPCR), ELISA, and immunoblotting. LPS-induced shock severity in WT, Nox4 knockdown and CSE knockout (CSE-/-) mice was assessed. Results: Here we showed that CSE and Nox4 were upregulated in macrophage and mouse in response to LPS. After LPS stimulation, the inflammatory responses were significantly ameliorated by lentiviral Nox4 shRNA knockdown, but were exacerbated by lentiviral overexpressing Nox4. Furthermore, Nox4 mediated inflammation through PI3K/Akt and p-p38 mitogen-activated protein kinase signal pathway. Notably, CSE knockout served to amplify the inflammatory cascade by increasing Nox4-ROS signaling activation in septic mice and macrophage. Similarly, the enhanced production of inflammatory mediators by macrophages was reduced by CSE overexpression. Conclusion: Thus, we demonstrated that CSE/H2S attenuated LPS-induced sepsis against oxidative stress and inflammation damage probably largely through mediated Nox4 pathway.

Aim of the Study: This study was designed to explore the relative susceptibility of in vitro fertilization (IVF)-conceived mice to global cerebral ischemic injury with the possible role of hydrogen sulphide and enzymes responsible for its production. Materials and Methods: IVF was carried to obtain pups, which were allowed to grow to the age of eight weeks. Thereafter, male mice were subjected to 20 min of global ischemia and 24 h of reperfusion. The mice obtained from other groups including normal mating, superovulation but normal mating and normal mating but embryo implantation were also subjected to global ischemia-reperfusion (I/R) injury. Results: IVF-derived mice exhibited significant more injury in response to I/R injury in comparison to other groups assessed in terms of impairment in locomotor activity, development of motor in coordination, neurological severity score, cerebral infarction and apoptosis markers (caspase-3 activity and Bcl-2 expression). Moreover, there was a relative decrease in the brain levels of hydrogen sulphide (H2S) and its biosynthetic enzymes viz. cystathionine-β-synthase and cystathionine-γ-lyase. Interestingly, the levels of H2S and cystathionine-γ-lyase were significantly low in IVF-derived mice in basal conditions also, i.e. before subjecting to I/R injury and these biochemical alterations were associated with the behavioural deficits in mice, even before subjecting to I/R injury. Conclusion: It is concluded that in vitro fertilization-derived mice are more susceptible to global cerebral I/R injury, which may be possibly due to decreased levels of hydrogen sulphide and its biosynthetic enzymes viz., cystathionine-β-synthase and cystathionine-γ-lyase. [ABSTRACT FROM AUTHOR]

The pathophysiology of sepsis-induced myocardial dysfunction is not resolved to date and comprises inflammation, barrier dysfunction and oxidative stress. Disease-associated reduction of tissue cystathionine-γ-lyase (CSE) expression, an endogenous H2S-producing enzyme, is associated with oxidative stress, barrier dysfunction and organ injury. CSE-mediated cardio-protection has been suggested to be related the upregulation of oxytocin receptor (OTR). CSE can also mediate glucocorticoid receptor (GR) signaling, which is important for normal heart function. A sepsis-related loss of cardiac CSE expression associated with impaired organ function has been reported previously. The aim of this current post hoc study was to investigate the role of cardiac GR and OTR after polymicrobial sepsis in a clinically relevant, resuscitated, atherosclerotic porcine model. Anesthetized and instrumented FBM (Familial Hypercholesterolemia Bretoncelles Meishan) pigs with high fat diet-induced atherosclerosis underwent poly-microbial septic shock (n = 8) or sham procedure (n = 5), and subsequently received intensive care therapy with fluid and noradrenaline administration for 24 h. Cardiac protein expression and mRNA levels were analyzed. Systemic troponin, a marker of cardiac injury, was significantly increased in septic animals in contrast to sham, whereas OTR and GR expression in septic hearts were reduced, along with a down-regulation of anti-inflammatory GR target genes and the antioxidant transcription factor NRF2. These results suggest a potential interplay between GR, CSE, and OTR in sepsis-mediated oxidative stress, inflammation and cardiac dysfunction. [ABSTRACT FROM AUTHOR]

H2S causes vasorelaxation however there is considerable heterogeneity in the reported pharmacological mechanism of this effect. This study examines the contribution of endogenously released H2S in the regulation of vascular tone and the mechanism of H2S-induced vasorelaxation in small resistance-like arteries. Mesenteric arteries from C57 and eNOS−/− mice were mounted in myographs to record isometric force. Vasorelaxation responses to NaHS were examined in the presence of various inhibitors of vasorelaxation pathways. Expression and activity of the H2S-producing enzyme, cystathionine-γ-lyase (CSE), were also examined. CSE was expressed in vascular smooth muscle and perivascular adipose cells from mouse mesenteric artery. The substrate for CSE, l-cysteine, caused a modest vasorelaxation (35%) in arteries from C57 mice and poor vasorelaxation (10%) in arteries from eNOS−/− mice that was sensitive to the CSE inhibitor dl-propargylglycine. The fast H2S donor, NaHS, elicited a full and biphasic vasorelaxation response in mesenteric arteries (EC50 (1) 8.7 μM, EC50 (2) 0.6 mM), which was significantly inhibited in eNOS−/− vessels (P < 0.05), unaffected by endothelial removal, or blockers at any point in the NO via soluble guanylate cyclase and cGMP (NO-sGC-cGMP) vasorelaxation pathway. Vasorelaxation to NaHS was significantly inhibited by blocking K+ channels of the KCa and KV subtypes and the Cl−/HCO3− exchanger (P < 0.05). Further experiments showed that NaHS can significantly inhibit voltage-gated Ca2+ channel function (P < 0.05). The vasorelaxant effect of H2S in small resistance-like arteries is complex, involving eNOS, K+ channels, Cl−/HCO3− exchanger, and voltage-gated Ca2+ channels. CSE is present in the smooth muscle and periadventitial adipose tissue of these resistance-like vessels and can be activated to cause modest vasorelaxation under these in vitro conditions. [ABSTRACT FROM AUTHOR]

Background/Aims: Chelerythrine (CHE), a benzophenanthridine alkaloid, is a potent, selective, and cell-permeable protein kinase C (PKC) inhibitor. The purpose of the present study was to evaluate the effect of CHE on myocardial recovery after renal ischemia/reperfusion (I/R)-induced myocardial injury (RI/RMI) in a streptozocin (STZ)-induced diabetic rat model. Methods: Diabetes mellitus (DM) rats preconditioned with CHE and D, L-propargylglycine (PAG) were subjected to renal I/R. The extent of cardiac morphologic lesions and the biochemical markers of cardiorenal function and oxidative stress were detected utilizing hematoxylin-eosin staining, commercial kits, and enzyme-linked immunoassay, respectively. The expressions of cystathionine-γ-lyase (CSE), PKC-α, PKC-β2, and nuclear factor-kappa B (NF-κB) in the rat myocardial tissue were measured utilizing western blotting. Results: Renal I/R treatment resulted in myocardial injury. CHE-preconditioning promoted the recovery from myocardial damage by ameliorating the biochemical parameters of myocardial injury, reducing oxidative stress, increasing the H2S level, up-regulating the expression of CSE, and down-regulating the expressions of PKC-α, PKC-β2, and NF-κB. Conclusion: These findings suggest that CHE-pretreatment may exert a protective effect on the myocardium against RI/RMI by activating endogenous CSE/H2S via PKC/NF-κB pathway in STZ-induced diabetic rats. Further studies are needed defining underlying mechanisms.

Cardiac hypertrophy is the most prevalent compensatory heart disease that ultimately leads to spontaneous heart failure. Mounting evidence suggests that microRNAs (miRs) and endogenous hydrogen sulfide (H 2 S) play a crucial role in the regulation of cardiac hypertrophy. In this study, we aimed to investigate whether inhibition of miR-27a could protect against cardiac hypertrophy by modulating H 2 S signaling. We established a model of cardiac hypertrophy by obtaining hypertrophic tissue from mice subjected to transverse aortic constriction (TAC) and from cells treated with angiotensin-II. Molecular alterations in the myocardium were quantified using quantitative real time PCR (qRT-PCR), Western blotting, and ELISA. Morphological changes were characterized by hematoxylin and eosin (HE) staining and Masson's trichrome staining. Functional myocardial changes were assessed using echocardiography. Our results demonstrated that miR-27a levels were elevated, while H 2 S levels were reduced in TAC mice and myocardial hypertrophy. Further luciferase and target scan assays confirmed that cystathionine-γ-lyase (CSE) was a direct target of miR-27a and was negatively regulated by it. Notably, enhancement of H 2 S expression in the heart was observed in mice injected with recombinant adeno-associated virus vector 9 (rAAV9)-anti-miR-27a and in cells transfected with a miR-27a inhibitor during cardiac hypertrophy. However, this effect was abolished by co-transfection with CSE siRNA and the miR-27a inhibitor. Conversely, injecting rAAV9-miR-27a yielded opposite results. Interestingly, our findings demonstrated that glucagon-like peptide-1 (GLP-1) agonists could mitigate myocardial damage by down-regulating miR-27a and up-regulating CSE. In summary, our study suggests that inhibition of miR-27a holds therapeutic promise for the treatment of cardiac hypertrophy by increasing H 2 S levels. Furthermore, our findings unveil a novel mechanism of GLP-1 agonists involving the miR-27a/H 2 S pathway in the management of cardiac hypertrophy., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Thyroid hormones have a role in the regulation of hydrogen sulfide (H2S) biosynthesis. In this study, we determined the effects of hyperthyroidism on H2S levels in various tissues and messenger RNA (mRNA) expression of cystathionine‐β‐synthase (CBS), cystathionine‐γ‐lyase (CSE), and 3‐mercaptopyruvate sulfurtransferase (3‐MST) in the liver and muscles of the rat. Sixteen male Wistar rats were divided into the hyperthyroid and the control groups. Hyperthyroidism was induced by adding l‐thyroxine (12 mg/L) to drinking water for a period of 21 days. H2S concentrations in serum, liver, aorta, heart, and soleus muscles, as well as mRNA expressions of CBS, CSE, and 3‐MST in these tissues were measured at Day 21. Hyperthyroid rats had lower H2S levels in the serum compared with controls (14.7 ± 1.4 vs. 25.7 ± 1.6 µmol/L, p < 0.001). Compared with controls, hyperthyroid rats had lower levels of H2S in the aorta (89%), heart (80%), and soleus (103%) muscles, but higher levels in the liver (35%). Hyperthyroidism decreased the ratio of CBS/CSE mRNA expression in the liver and the CSE/CBS mRNA expression in the muscles by decreasing CBS levels in liver (34% cf. controls) and CSE levels in the aorta, heart, and soleus muscles (respectively, 51%, 7%, and 52% cf.). In addition, hyperthyroidism decreased the mRNA expression of 3‐MST in the liver (51%) and aorta (33%), and increased it in the heart (300%) and soleus muscle (182%). In conclusion, hyperthyroidism increased H2S levels in the liver and decreased it in muscles; these effects are at least in part due to increases and decreases in expression of CSE in the liver and muscles, respectively. These data indicate an association between thyroid hormone status and gene expression of the H2S‐producing enzymes in the rat. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • Celecoxib reverses the hypertensive, nephrotoxic, and CSE/H 2 S inhibitory effects of CSA. • The celecoxib effects are replicated by Na 2 S and abrogated by DL-PAG". • Celecoxib or Na 2 S ameliorates the inflammatory, oxidative, and apoptotic insults of CSA. Abstract We recently reported that celecoxib, a selective cyclooxygenase-2 (COX2) inhibitor, counteracts the adverse circulatory and renal actions of cyclosporine (CSA). Despite the seemingly advantageous nature of this interaction particularly in clinical settings that necessitate the combined use of the two drugs such as immune-related arthritis, the underlying mechanism remains elusive. This prompted us to test the hypothesis that the facilitation of the cystathionine-γ-lyase (CSE)/hydrogen sulfide (H 2 S) signaling accounts for such favorable effects of celecoxib on CSA nephrotoxicity. The data showed that the 10-day co-treatment of rats with celecoxib (10 mg/kg/day) ameliorated the hypertensive and biochemical and renal structural damages caused by CSA (20 mg/kg/day). Celecoxib also reversed the CSA-evoked (i) reductions in the tubular and glomerular protein expression of CSE and levels of H 2 S, prostaglandin E2 (PGE2), and total antioxidant capacity (TAC), and (ii) increases in inflammatory (tumor necrosis factor-α, TNF-α), fibrotic (transforming growth factor-β1, TGF-β1) and apoptotic (caspase-3) cytokines. These celecoxib effects disappeared when rats were treated concomitantly with the CSE inhibitor DL-propargylglycine (DL-PAG), indicating the importance of the CSE-derived H 2 S in mediating the renoprotective action of celecoxib. This view is bolstered by the observation that the beneficial hemodynamic and renal actions of celecoxib were replicated after supplementation of rats with sodium sulfide (Na 2 S, H 2 S donor). Together, the increased abundance of renal CSE and H 2 S and subsequent dampening down of inflammatory, fibrotic, oxidant, and apoptotic pathways play pivotal roles in the capacity of celecoxib to compromise the troublesome hypertensive and nephrotoxic insults caused by CSA in rats. [ABSTRACT FROM AUTHOR]

Highlights • Hypoxia increased carotid body (CB) H 2 S levels, sensory nerve activity, and elevated [Ca2+] i in glomus cells. • Anoxia had no effect on CB H 2 S levels but produced only weak sensory excitation and robust elevation of [Ca2+] i in glomus cells. • Inhibition of H 2 S synthesis or genetic deletion of cystathionine-γ-lyase (CSE) prevented the effects of hypoxia but not anoxia on the CB. • Hypoxia stimulated breathing in wild-type but not in CSE knockout mice, whereas anoxia inhibited breathing in both strains of mice. • Hypoxia and anoxia are not similar stimuli for studying CB physiology, and H 2 S contributes to CB response to hypoxia but not to anoxia. Abstract The role of cystathionine-γ-lyase (CSE) derived H 2 S in the hypoxic and anoxic responses of the carotid body (CB) were examined. Experiments were performed on Sprague-Dawley rats, wild type and CSE knockout mice on C57BL/6 J background. Hypoxia (pO 2 = 37 ± 3 mmHg) increased the CB sensory nerve activity and elevated H 2 S levels in rats. In contrast, anoxia (pO 2 = 5 ± 4 mmHg) produced only a modest CB sensory excitation with no change in H 2 S levels. DL-propargylglycine (DL-PAG), a blocker of CSE, inhibited hypoxia but not anoxia-evoked CB sensory excitation and [Ca2+] i elevation of glomus cells. The inhibitory effects of DL-PAG on hypoxia were seen: a) when it is dissolved in saline but not in dimethyl sulfoxide (DMSO), and b) in glomus cells cultured for18 h but not in cells either soon after isolation or after prolonged culturing (72 h) requiring 1–3 h of incubation. On the other hand, anoxia-induced [Ca2+] i responses of glomus cell were blocked by high concentration of DL-PAG (300μM) either alone or in combination with aminooxyacetic acid (AOAA; 300μM) with a decreased cell viability. Anoxia produced a weak CB sensory excitation and robust [Ca2+] i elevation in glomus cells of both wild-type and CSE null mice. As compared to wild-type, CSE null mice exhibited impaired CB chemo reflex as evidenced by attenuated efferent phrenic nerve responses to brief hyperoxia (Dejours test), and hypoxia. Inhalation of 100% N 2 (anoxia) depressed breathing in both CSE null and wild-type mice. These observations demonstrate that a) hypoxia and anoxia are not analogous stimuli for studying CB physiology and b) CSE-derived H 2 S contributes to CB response to hypoxia but not to that of anoxia. [ABSTRACT FROM AUTHOR]

Endogenous gas transmitters, hydrogen sulfide (H2S), carbon monoxide (CO) and nitric oxide (NO) are important signaling molecules known to exert multiple biological functions. In recent years, the role of H2S, CO and NO in regulation of cardiovascular, neuronal and digestive systems physiology and pathophysiology has been emphasized. Possible link between these gaseous mediators and multiple diseases as well as potential therapeutic applications has attracted great attention from biomedical scientists working in many fields of biomedicine. Thus, various pharmacological tools with ability to release CO or H2S were developed and implemented in experimental animal in vivo and in vitro models of many disorders and preliminary human studies. This review was designed to review signaling functions, similarities, dissimilarities and a possible cross-talk between H2S and CO produced endogenously or released from chemical donors, with special emphasis on gastrointestinal digestive system pathologies prevention and treatment.

Background/Aims: Sepsis is a severe and complicated syndrome that is characterized by dysregulation of host inflammatory responses and organ failure. Cystathionine-γ-lyase (CSE)/ hydrogen sulfide (H2S) has potential anti-inflammatory activities in a variety of inflammatory diseases. NADPH oxidase 4 (Nox4), a member of the NADPH oxidases, is the major source of reactive oxygen species (ROS) and its expression is increased in sepsis, but its function in CSE-mediated anti-inflammatory activities remains unknown. Methods: Macrophages were either transfected with CSE, Nox4 siRNA or transduced with lentiviral vector encoding CSE or Nox4, and then stimulated with lipopolysaccharide (LPS). The expression of inflammatory mediators and signaling pathway activation were measured by quantitative PCR (qPCR), ELISA, and immunoblotting. LPS-induced shock severity in WT, Nox4 knockdown and CSE knockout (CSE-/-) mice was assessed. Results: Here we showed that CSE and Nox4 were upregulated in macrophage and mouse in response to LPS. After LPS stimulation, the inflammatory responses were significantly ameliorated by lentiviral Nox4 shRNA knockdown, but were exacerbated by lentiviral overexpressing Nox4. Furthermore, Nox4 mediated inflammation through PI3K/Akt and p-p38 mitogen-activated protein kinase signal pathway. Notably, CSE knockout served to amplify the inflammatory cascade by increasing Nox4-ROS signaling activation in septic mice and macrophage. Similarly, the enhanced production of inflammatory mediators by macrophages was reduced by CSE overexpression. Conclusion: Thus, we demonstrated that CSE/H2S attenuated LPS-induced sepsis against oxidative stress and inflammation damage probably largely through mediated Nox4 pathway. [ABSTRACT FROM AUTHOR]

Summary: Hydrogen sulfide (H2S) is generated from l‐cysteine by multiple enzymes including cystathionine‐γ‐lyase (CSE), and promotes nociception by targeting multiple molecules such as Cav3.2 T‐type Ca2+ channels. Bladder pain accompanying cyclophosphamide (CPA)‐induced cystitis in mice has been shown to involve the functional upregulation of the CSE/H2S/Cav3.2 pathway. Therefore, we investigated whether NF‐κB, as an upstream signal of the CSE/H2S system, contributes to bladder pain in mice with CPA‐induced cystitis. Bladder pain‐like nociceptive behaviour was observed in CPA‐treated mice, and referred hyperalgesia was evaluated by the von Frey test. Isolated bladder weights were assessed to estimate bladder swelling, and protein levels were measured by Western blotting. CPA, administered intraperitoneally, induced nociceptive behaviour, referred hyperalgesia and increased bladder weights in mice. β‐Cyano‐ l‐alanine, a reversible selective CSE inhibitor, prevented CPA‐induced nociceptive behaviour, referred hyperalgesia, and, in part, increases in bladder weight. CPA markedly increased phosphorylated NF‐κB p65 levels in the bladder, an effect that was prevented by pyrrolidine dithiocarbamate (PDTC), an NF‐κB inhibitor. PDTC and curcumin, which inhibits NF‐κB signals, abolished CPA‐induced nociceptive behaviour, referred hyperalgesia and, in part, increases in bladder weight. CPA caused the overexpression of CSE in the bladder, and this was prevented by PDTC or curcumin. The CPA‐induced activation of NF‐κB signals appeared to cause CSE overexpression in the bladder, contributing to bladder pain and in part swelling, possibly through H2S/Cav3.2 signaling. Therefore, NF‐κB‐inhibiting compounds including curcumin may be useful for the treatment of cystitis‐related bladder pain. [ABSTRACT FROM AUTHOR]

Overproduction of inflammatory mediators contributes to uncontrolled inflammation during endotoxin shock. Cystathionine-γ-lyase (CSE), an enzyme involved in hydrogen sulfide (H 2 S) biosynthesis, has potential anti-inflammatory activity in a variety of inflammatory diseases. Jumonji domain-containing protein 3 (JMJD3), a histone 3 Lys27 (H3K27) demethylase, has been implicated in macrophage activation, but its function in CSE-mediated anti-inflammatory activities remains unknown. In the present study CSE was found to be upregulated in macrophages and mouse lipopolysaccharide (LPS) challenge models. LPS stimulation also enhanced the activation of JMJD3 and decreased H3K27me3 levels. JMJD3 knockdown upregulated H3K27me3 levels and attenuated the LPS-mediated inflammatory response. CSE knockout amplified the inflammatory cascade by increasing JMJD3 expression in septic mice. Similarly, enhanced production of inflammatory mediators by macrophages was mitigated by CSE overexpression via inhibition of JMJD3 expression. This is the first report indicating that inflammation enhanced CSE/H 2 S system biosynthesis, that in turn attenuated the LPS-triggered inflammatory response by regulating JMJD3 expression. Thus, the CSE/H 2 S system represents an epigenetic-based modification mechanism to prevent uncontrolled inflammation. [ABSTRACT FROM AUTHOR]

Cystathionine-γ-lyase (CSE) is a multifunctional enzyme, and hydrogen sulfide (H 2 S) is one of its products. CSE and H 2 S have recently been proposed to be critical signaling molecules in hypoxia-induced excitation of carotid body (CB) glomus cells and the chemosensory response. Because the role of H 2 S in arterial chemoreception is still debated, we further examined the role of CSE by studying the effects of hypoxia on TASK K + channel activity, cell depolarization, [Ca 2+ ] i and ventilation using CSE +/+ and CSE −/− mice. As predicted, hypoxia reduced TASK activity and depolarized glomus cells isolated from CSE +/+ mice. These effects of hypoxia were not significantly altered in glomus cells from CSE −/− mice. Basal [Ca 2+ ] i and hypoxia-induced elevation of [Ca 2+ ] were also not significantly different in glomus cells from CSE +/+ and CSE −/− mice. In whole-body plethysmography, hypoxia (10%O 2 ) increased minute ventilation in both CSE +/+ and CSE −/− mice equally well, and no significant differences were found in either males or females when adjusted by body weight. Together, these results show that deletion of the CSE gene has no effects on hypoxia-induced changes in TASK, cell depolarization, [Ca 2+ ] i and ventilation, and therefore do not support the idea that CSE/H 2 S signaling is important for CB chemoreceptor activity in mice. [ABSTRACT FROM AUTHOR]

Hydrogen sulfide (H2S) plays an important role in smooth muscle relaxation. Here, we investigated the expression of enzymes in H2S synthesis and the mechanism regulating colonic smooth muscle function by H2S. Expression of cystathionine-γ-lyase (CSE), but not cystathionine-β-synthase (CBS), was identified in the colonic smooth muscle of rabbit, mouse, and human. Carbachol (CCh)-induced contraction in rabbit muscle strips and isolated muscle cells was inhibited by l-cysteine (substrate of CSE) and NaHS (an exogenous H2S donor) in a concentration-dependent fashion. H2S induced S-sulfhydration of RhoA that was associated with inhibition of RhoA activity. CCh-induced Rho kinase activity also was inhibited by l-cysteine and NaHS in a concentration-dependent fashion. Inhibition of CCh-induced contraction by l-cysteine was blocked by the CSE inhibitor, dl-propargylglycine (DL-PPG) in dispersed muscle cells. Inhibition of CCh-induced Rho kinase activity by l-cysteine was blocked by CSE siRNA in cultured cells and DL-PPG in dispersed muscle cells. Stimulation of Rho kinase activity and muscle contraction in response to CCh was also inhibited by l-cysteine or NaHS in colonic muscle cells from mouse and human. Collectively, our studies identified the expression of CSE in colonic smooth muscle and determined that sulfhydration of RhoA by H2S leads to inhibition of RhoA and Rho kinase activities and muscle contraction. The mechanism identified may provide novel therapeutic approaches to mitigate gastrointestinal motility disorders. [ABSTRACT FROM AUTHOR]

Background/Aims: Chelerythrine (CHE), a benzophenanthridine alkaloid, is a potent, selective, and cell-permeable protein kinase C (PKC) inhibitor. The purpose of the present study was to evaluate the effect of CHE on myocardial recovery after renal ischemia/reperfusion (I/R)-induced myocardial injury (RI/RMI) in a streptozocin (STZ)-induced diabetic rat model. Methods: Diabetes mellitus (DM) rats preconditioned with CHE and D, L-propargylglycine (PAG) were subjected to renal I/R. The extent of cardiac morphologic lesions and the biochemical markers of cardiorenal function and oxidative stress were detected utilizing hematoxylin-eosin staining, commercial kits, and enzyme-linked immunoassay, respectively. The expressions of cystathionine-γ-lyase (CSE), PKC-α, PKC-β2, and nuclear factor-kappa B (NF-κB) in the rat myocardial tissue were measured utilizing western blotting. Results: Renal I/R treatment resulted in myocardial injury. CHE-preconditioning promoted the recovery from myocardial damage by ameliorating the biochemical parameters of myocardial injury, reducing oxidative stress, increasing the H2S level, up-regulating the expression of CSE, and down-regulating the expressions of PKC-α, PKC-β2, and NF-κB. Conclusion: These findings suggest that CHE-pretreatment may exert a protective effect on the myocardium against RI/RMI by activating endogenous CSE/H2S via PKC/NF-κB pathway in STZ-induced diabetic rats. Further studies are needed defining underlying mechanisms. [ABSTRACT FROM AUTHOR]

Excessive proliferation of vascular smooth muscle cells (SMC) is an important contributor to the progression of atherosclerosis. Inhibition of proliferation can be achieved by endogenously produced and exogenously supplied nitrogen monoxide, commonly known as nitric oxide (NO). We report herein the dichotomous effects of two isomeric families of secondary amines, precursors to the N -nitrosated NO-donors, on HASMC proliferation. The syntheses of these two families were carried out using two equivalents of homologous, aliphatic monoamines and 2,6-difluoro-3-nitrobenzonitrile (2,6-DFNBN, O family) or 2,4-difluoro-5-nitrobenzonitrile (2,4-DFNBN, P family). The secondary amines belonging to the P family inhibited HASMC proliferation at all concentrations, whereas the O family induced HASMC proliferation at low concentrations, and exhibited inhibitory properties at high concentrations. A probable explanation of these behaviors is proposed herein. l -homocysteine (HCY) is known to induce HASMC proliferation at low concentrations (<1 mM) and inhibit HASMC proliferation at higher concentrations (>2.5 mM). Our findings suggest that these two families of amines inhibit cystathionine-γ-lyase (CSE) to varying extents, which directly results in altered levels of intracellular HCY and consequent changes in HASMC proliferation. [ABSTRACT FROM AUTHOR]

The chronic articular disease osteoarthritis (OA) is characterized by osteophyte generation, subchondral bone remodeling, and cartilage deterioration. Low levels of H2S catalyzed by cystathionine-γ-lyase (CSE) encoded by Cthhas neuroprotective, cardioprotective, anti-apoptotic, and anti-inflammatory effects thus, Cth is being developed as a potential therapy for the management of the pathogenesis and symptoms of osteoarthritis. Single-cell RNA sequencing (scRNA-seq) and immunohistochemistry of human cartilage revealed that the expression of CTH was decreased in OA patients. We found that Cthoverexpression decrease IL-1β-induced overactivation of the NF-κB signaling pathway. In vivo, Cthoverexpression relieved pain response and cartilage damage in the anterior cruciate ligament transection (ACLT) rat model. In vitro, CSE alleviated chondrocytes catabolism, inflammation, apoptosis, and senescence, and suppressed the NF-κB pathway. We postulate that CSE has therapeutic effects in suppressing inflammation and degeneration in OA and should be further investigated clinically., 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 © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

Alcohol is the most socially accepted addictive drug. Alcohol consumption is associated with some health problems such as neurological, cognitive, behavioral deficits, cancer, heart, and liver disease. Mechanisms of alcohol-induced toxicity are presently not yet clear. One of the mechanisms underlying alcohol toxicity has to do with its interaction with amino acid homocysteine (Hcy), which has been linked with brain neurotoxicity. Elevated Hcy impairs with various physiological mechanisms in the body, especially metabolic pathways. Hcy metabolism is predominantly controlled by epigenetic regulation such as DNA methylation, histone modifications, and acetylation. An alteration in these processes leads to epigenetic modification. Therefore, in this review, we summarize the role of Hcy metabolism abnormalities in alcohol-induced toxicity with epigenetic adaptation and their influences on cerebrovascular pathology. [ABSTRACT FROM AUTHOR]

Background/Aim: H2S is a novel vasoactivator. To verify the hypothesis that H2S may act as an endothelium-derived hyperpolarizing factor (EDHF) in the rat cerebrovasculature, the role of H2S in endothelium-derived relaxing factor (EDRF)-mediated responses was investigated. Methods: Cystathionine-γ-lyase (CSE) was knocked down with an siRNA technique. Artery diameter, hyperpolarization and Ca2+-activated K+ (KCa) current were measured. Results: CSE knockdown was indicated by a decrease in protein and mRNA expression in the rat middle cerebral artery (MCA) and cerebral basilar artery (CBA). Acetylcholine (ACh) induced significant hyperpolarization and vasodilation in endothelium-intact MCA and CBA. Removal of the endothelium abolished these responses. The nitric oxide (NO) synthase inhibitor L-NAME, but not the PGI2 production inhibitor indomethacin, significantly inhibited ACh-induced hyperpolarization and vasodilation in the CBA. In the presence of L-NAME and indomethacin, ACh-induced hyperpolarization and vasodilation in the MCA and CBA were attenuated. The non-NO/PGI2-mediated responses were abolished by the KCa channel blockers charybdotoxin and apamin. In the cerebral arteries from the CSE knockdown rat, non-NO/PGI2-mediated responses were significantly attenuated, and the remaining responses were abolished by charybdotoxin and apamin or the CSE inhibitor propargylglycine. CSE knockdown did not affect L-NAME-sensitive responses in the CBA. Sodium hydrosulfide (NaHS) augmented the KCa current in CBA vascular smooth muscle cells. Conclusion: EDHF-mediated responses in rat cerebral arteries were due to H2S activating the KCa channel. [ABSTRACT FROM AUTHOR]

The etiology of achalasia remains largely unknown. Considerable evidence reveals that the lower esophageal sphincter dysfunction is due to the lack of inhibitory neurotransmitter, secondary to esophageal neuronal inflammation or loss. Recent studies suggest hydrogen sulfide may act as an inhibitory transmitter in gastrointestinal tract, but study about hydrogen sulfide in human esophagus still lack. The aim of the study was to investigate if hydrogen sulfide synthesis enzymes could be detected in human esophagus and if the synthesis of the endogenous hydrogen sulfide could be affected in achalasia patients. Tissue samples in cardia, lower esophageal sphincter, 2 cm and 4 cm above lower esophageal sphincter were obtained from achalasia patients undergoing peroral endoscopic myotomy. Control tissues in lower esophageal sphincter were obtained from esophageal carcinoma patients. Expression of cystathionine-β-synthase and cystathionine-γ-lyase in lower esophageal sphincter of achalasia patients and control were detected by immunohistochemical staining. In addition, expression of cystathionine-β-synthase and cystathionine-γ-lyase were compared among different parts of esophagus in achalasia patients. Compared with control, the expression of cystathionine-β-synthase and cystathionine-γ-lyase in lower esophageal sphincter of achalasia patients was significantly reduced (χ2 = 11.429, P = 0.010). The expression of cystathionine-β-synthase and cystathionine-γ-lyase were lower in lower esophageal sphincter than that in 2 cm and 4 cm above lower esophageal sphincter, respectively (all P < 0.05). In conclusion, the expression of hydrogen sulfide synthesis enzymes, cystathionine-β-synthase and cystathionine-γ-lyase, can be detected in human esophagus and is reduced in patients with achalasia, which implicates the involvement of the two hydrogen sulfide synthesis enzymes in the pathophysiology of achalasia. [ABSTRACT FROM AUTHOR]

Whether high oxygen is harmful to the vascular function is unclear. The present study examined if high oxygen modifies vasodilator effect of cysteine via enhanced oxidative stress and thromboxane production . Rat mesenteric arteries with endothelium at 95 or 50 % oxygen were subjected to isometric force recordings, measurement of thromboxane B levels, determination of superoxide and peroxynitrite levels and evaluation of NADPH oxidase subunit protein expression, respectively. L-cysteine (0.01-3 mM) constricted or dilated arteries at 95 and 50 % oxygen, respectively. Thromboxane receptor antagonist SQ-29,548 (1 μM) abolished the constriction at 95 % oxygen. L-cysteine (3 mM) increased levels of thromboxane B in arteries upon 95 % oxygen application. L-cysteine relaxed arteries treated with superoxide inhibitor tiron (2 mM) or NADPH oxidase inhibitor gp91ds-tat (1 μM) irrespective of the oxygen concentration while ATP-sensitive K channel inhibitor glibenclamide (1 μM) and cystathionine-γ-lyase (CSE) inhibitor DL-propargylglycine (10 mM) similarly abolished the relaxation. L-cysteine (3 mM) with 95 % oxygen augmented levels of superoxide as well as nitrotyrosine within the artery, concomitantly with enhanced membrane protein expression of NADPH oxidase subunit p47phox. The higher concentration of oxygen attenuates L-cysteine-induced vasodilation via superoxide production mediated by NADPH oxidase along with thromboxane A production, resulting in vasoconstriction. The increased levels of superoxide, as well as peroxynitrite, coexist with the impaired vasodilation related to ATP-sensitive K channels and CSE. Higher oxygen with plasma cysteine may cause oxidative stress and vasoconstrictor prostanoid production in blood vessels. [ABSTRACT FROM AUTHOR]