Sulfur dioxide inhibits mast cell degranulation by sulphenylation of galectin-9 at cysteine 74.

Objectives: Mast cell (MC) degranulation is a key process in allergic reactions and inflammatory responses. Aspartate aminotransferase 1 (AAT1)-derived endogenous sulfur dioxide (SO2) is an important regulator of MC function. However, the mechanism underlying its role in MC degranulation remains unclear. This study aimed to investigate the mechanism by which endogenous SO2 controlled MC degranulation. Methods: HMC-1 and Rat basophilic leukemia cell MC line (RBL-2H3) were used in the cell experiments. SO2 content was detected by in situ fluorescent probe. MC degranulation represented by the release rate of MC ß-hexosaminidase was determined using a colorimetric assay. Sulfenylation of galectin-9 (Gal-9) in MCs and purified protein was detected using a biotin switch assay. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to determine the exact sulfenylation sites of Gal-9 by SO2. Animal models of passive cutaneous anaphylaxis (PCA) and hypoxia-driven pulmonary vascular remodeling were used to investigate the effect of SO2 on mast cell activation in vivo. Site-directed mutation of Gal-9 was conducted to confirm the exact site of SO2 and support the significance of SO2/Gal-9 signal axis in the regulation of MC degranulation. Results: Degranulation was increased in AAT1-knockdowned MCs, and SO2 supplementation reversed the increase in MC degranulation. Furthermore, deficiency of endogenous SO2 contributed to IgE-mediated degranulation in vitro. Besides, SO2 inhibited IgE-mediated and hypoxia-driven MC degranulation in vivo. Mechanistically, LC-MS/MS analysis and site-directed mutation results showed that SO2 sulfenylated Gal-9 at cysteine 74. Sulfenylation of the 74th cysteine of Gal-9 protein was required in the SO2-inhibited MC degranulation under both physiological and pathophysiological conditions. Conclusion: These findings elucidated that SO2 inhibited MC degranulation via sulfenylating Gal-9 under both physiological and pathophysiological conditions, which might provide a novel treatment approach for MC activation-related diseases.

Endogenous sulfur dioxide deficiency as a driver of cardiomyocyte senescence through abolishing sulphenylation of STAT3 at cysteine 259.

OBJECTIVE: Cardiomyocyte senescence is an important contributor to cardiovascular diseases and can be induced by stressors including DNA damage, oxidative stress, mitochondrial dysfunction, epigenetic regulation, etc. However, the underlying mechanisms for the development of cardiomyocyte senescence remain largely unknown. Sulfur dioxide (SO2) is produced endogenously by aspartate aminotransferase 2 (AAT2) catalysis and plays an important regulatory role in the development of cardiovascular diseases. The present study aimed to explore the effect of endogenous SO2 on cardiomyocyte senescence and the underlying molecular mechanisms. APPROACH AND RESULTS: We interestingly found a substantial reduction in the expression of AAT2 in the heart of aged mice in comparison to young mice. AAT2-knockdowned cardiomyocytes exhibited reduced SO2 content, elevated expression levels of Tp53, p21Cip/Waf, and p16INk4a, enhanced SA-ß-Gal activity, and elevated level of γ-H2AX foci. Notably, supplementation with a SO2 donor ameliorated the spontaneous senescence phenotype and DNA damage caused by AAT2 deficiency in cardiomyocytes. Mechanistically, AAT2 deficiency suppressed the sulphenylation of signal transducer and activator of transcription 3 (STAT3) facilitated its nuclear translocation and DNA-binding capacity. Conversely, a mutation in the cysteine (Cys) 259 residue of STAT3 blocked SO2-induced STAT3 sulphenylation and subsequently prevented the inhibitory effect of SO2 on STAT3-DNA-binding capacity, DNA damage, and cardiomyocyte senescence. Additionally, cardiomyocyte (cm)-specific AAT2 knockout (AAT2cmKO) mice exhibited a deterioration in cardiac function, cardiomegaly, and cardiac aging, whereas supplementation with SO2 donors mitigated the cardiac aging and remodeling phenotypes in AAT2cmKO mice. CONCLUSION: Downregulation of the endogenous SO2/AAT2 pathway is a crucial pathogenic mechanism underlying cardiomyocyte senescence. Endogenous SO2 modifies STAT3 by sulphenylating Cys259, leading to the inhibition of DNA damage and the protection against cardiomyocyte senescence.

Gasotransmitter-Mediated Cysteinome Oxidative Posttranslational Modifications: Formation, Biological Effects, and Detection.

Significance: Gasotransmitters, including nitric oxide (NO), hydrogen sulfide (H2S) and sulfur dioxide (SO2), participate in various cellular processes via corresponding oxidative posttranslational modifications (oxiPTMs) of specific cysteines. Recent Advances: Accumulating evidence has clarified the mechanisms underlying the formation of oxiPTMs derived from gasotransmitters and their biological functions in multiple signal pathways. Because of the specific existence and functional importance, determining the sites of oxiPTMs in cysteine is crucial in biology. Recent advances in the development of selective probes, together with upgraded mass spectrometry (MS)-based proteomics, have enabled the quantitative analysis of cysteinome. To date, several cysteine residues have been identified as gasotransmitter targets. Critical Issues: To clearly understand the underlying mechanisms for gasotransmitter-mediated biological processes, it is important to identify modified targets. In this review, we summarize the chemical formation and biological effects of gasotransmitter-dependent oxiPTMs and highlight the state-of-the-art detection methods. Future Directions: Future studies in this field should aim to develop the next generation of probes for in situ labeling to improve spatial resolution and determine the dynamic change of oxiPTMs, which can lay the foundation for research on the molecular mechanisms and clinical translation of gasotransmitters. Antioxid. Redox Signal. 40, 145-167.

Gut Microecology May Be Involved in the Pathogenesis of Hashimoto Thyroiditis by Reducing Production of Hydrogen Sulfide.

CONTEXT: Hashimoto thyroiditis (HT) is related to intestinal microbiota alteration, but the causal relationship remains unclear. Hydrogen sulfide (H2S) is a microbiota-derived metabolite. We speculated that abnormal intestinal microbiota might limit H2S production capacity, promoting HT pathogenesis. OBJECTIVE: This work aimed to illustrate that the intestinal microbiota plays important roles in HT pathogenesis via microbiota-derived H2S levels. METHODS: We collected feces from HT patients and healthy donors for fecal microbiota transplantation (FMT). Thirty-six female CBA/J mice were randomly assigned to 4 groups: experimental autoimmune thyroiditis (EAT) group, EAT + Healthy group, EAT + HT group, and EAT + HT + H2S group. 16S ribosomal RNA sequencing was performed to examine gut microbiota alterations and the H2S production pathway. Serum TgAb and H2S levels were assayed by enzyme-linked immunosorbent assay and H2S-selective sensors, respectively. T-cell subpopulations in the spleen were detected by flow cytometry. RESULTS: The gut microbiota was different after FMT among the EAT, EAT + Healthy, and EAT + HT groups. The thyroiditis score assessed by hematoxylin and eosin staining was higher in the EAT + HT group than that in the EAT and EAT + HT + H2S groups. Helper T (Th1) and Th17 cell differentiation ratios were increased in the EAT + HT group compared to the other 3 groups. Serum H2S levels were decreased and the dissimilatory sulfate reduction (DSR) pathway was attenuated in the EAT + HT group compared to the EAT + Healthy group. CONCLUSION: H2S alleviated thyroiditis severity and related immune disorders, which were aggravated by the FMT from HT patients. The attenuated DSR pathway in the gut microbiota from HT patients might be involved in thyroiditis pathogenesis.

Serum uric acid predicts therapeutic response to midodrine hydrochloride in children with vasovagal syncope: a pilot study.

Serum uric acid (UA) level has been proven to be related to several cardiovascular and metabolic diseases. In the present study, we examined if baseline serum UA level could predict the therapeutic efficacy of midodrine hydrochloride on vasovagal syncope (VVS) in children. The pediatric VVS patients who received midodrine hydrochloride from November 2008 to October 2022 were enrolled. After a median treatment duration of 3 months, the therapeutic effect was evaluated. According to the patients' responses to midodrine hydrochloride, which was determined by the recurrence of syncope, they were divided into effective and ineffective groups. The baseline variables were explored using univariable and multivariate logistic analysis. The predictive efficacy was assessed by receiver operating characteristic curve (ROC), precision-recall curve (PR), Hosmer-Lemeshow test, calibration curve, and decision curve analysis (DCA). Totally, 53 participants were included in the study. Among the 51 patients who were successfully followed up, 29 (56.9%) responded to midodrine hydrochloride (effective group), and the other 22 (43.1%) failed to respond to midodrine hydrochloride (ineffective group). The participants in effective group had lower baseline serum UA level than those in ineffective group (276.5 ± 73 µmol/L vs. 332.7 ± 56 µmol/L, p = 0.004). Multivariable logistic analysis showed that serum UA was associated with the therapeutic response (odds ratio (OR): 0.985, 95% confidence interval (CI): 0.974-0.997, p = 0.01). ROC analysis indicated that using baseline serum UA < 299 µmol/L as a threshold value yielded a sensitivity of 77.3% and a specificity of 79.3% in predicting the treatment response to midodrine hydrochloride. The area under the PR curve was 0.833. Hosmer-Lemeshow test yielded a p value of 0.58, and calibration plot indicated that the model was well-fitted. DCA demonstrated that treatment decision depending on the baseline serum UA level resulted in a favorable net benefit.   Conclusion: This pilot study suggested that the baseline serum UA level could be taken as a predictor of therapeutic effect of midodrine hydrochloride on VVS in children. What is Known: • Empirical and unselected use of midodrine hydrochloride has an unfavorable therapeutic effect on VVS in children. Serum uric acid (UA) is closely linked to cardiovascular events. What is New: • A low baseline serum UA level successfully predicts the therapeutic effectiveness of midodrine hydrochloride on VVS in children.

L-cystathionine protects against oxidative stress and DNA damage induced by oxidized low-density lipoprotein in THP-1-derived macrophages.

Introduction: Oxidative stress in monocyte-derived macrophages is a significant pathophysiological process in atherosclerosis. L-cystathionine (L-Cth) acts as a scavenger for oxygen free radicals. However, the impact of L-Cth on macrophage oxidative stress during atherogenesis has remained unclear. This study aimed to investigate whether L-Cth affects oxidative stress in THP-1-derived macrophages and its subsequent effects on DNA damage and cell apoptosis. Methods: We established a cellular model of oxLDL-stimulated macrophages. The content of superoxide anion, H2O2, NO, and H2S in the macrophage were in situ detected by the specific fluorescence probe, respectively. The activities of SOD, GSH-Px, and CAT were measured by colorimetrical assay. The protein expressions of SOD1, SOD2, and iNOS were detected using western blotting. The DNA damage and apoptosis in the macrophage was evaluated using an fluorescence kit. Results: The results demonstrated that oxLDL significantly increased the content of superoxide anion and H2O2, the expression of iNOS protein, and NO production in macrophages. Conversely, oxLDL decreased the activity of antioxidants GSH-Px, SOD, and CAT, and downregulated the protein expressions of SOD1 and SOD2 in macrophages. However, treatment with L-Cth reduced the levels of superoxide anion, H2O2, and NO, as well as the protein expression of iNOS induced by oxLDL. Moreover, L-Cth treatment significantly enhanced GSH-Px, SOD, and CAT activity, and upregulated the expressions of SOD1 and SOD2 proteins in macrophages treated with oxLDL. Furthermore, both L-Cth supplementation and activation of endogenous L-Cth production suppressed DNA damage and cell apoptosis in oxLDL-injured macrophages, whereas inhibition of endogenous L-Cth exacerbated the deleterious effects of oxLDL. Conclusion: These findings suggest that L-Cth exerts a pronounced inhibitory effect on the oxidative stress, subsequent DNA damage and cell apoptosis in oxLDL-stimulated THP-1 monocytes. This study deepens our understanding of the pathogenesis of macrophage-related cardiovascular pathology.

Hydrogen Sulfide Regulates Macrophage Function in Cardiovascular Diseases.

Significance: Hydrogen sulfide (H2S) is an endogenous gasotransmitter that plays a vital role in immune system regulation. Recently, the regulation of macrophage function by H2S has been extensively and actively recognized. Recent Advances: The mechanisms by which endogenous H2S controls macrophage function have attracted increasing attention. The generation of endogenous H2S from macrophages is mainly catalyzed by cystathionine-γ-lyase. H2S is involved in the macrophage activation and inflammasome formation, which contributes to macrophage apoptosis, adhesion, chemotaxis, and polarization. In addition, H2S has redox ability and interacts with reactive oxygen species to prevent oxidative stress. Moreover, H2S epigenetically regulates gene expression. Critical Issues: In this article, the generation of endogenous H2S in macrophages and its regulatory effect on macrophage function are reviewed. In addition, the signal transduction targeting macrophages by H2S is also addressed. Finally, the potential therapeutic effect of H2S on macrophages is discussed. Future Directions: Further experiments are required to explore the involvement of endogenous H2S in the regulation of macrophage function in various physiological and pathophysiological processes and elucidate the mechanisms involved. Regarding the clinical translation of H2S, further exploration of the application of H2S in inflammation-related diseases is needed. Antioxid. Redox Signal. 38, 45-56.

Endogenous Hydrogen Sulfide Persulfidates Caspase-3 at Cysteine 163 to Inhibit Doxorubicin-Induced Cardiomyocyte Apoptosis.

Doxorubicin (DOX) is an efficient antitumor anthracycline drug, but its cardiotoxicity adversely affects the prognosis of the patients. In this study, we explored whether endogenous gasotransmitter hydrogen sulfide (H2S) could protect against DOX-induced cardiomyocyte apoptosis and its mechanisms. The results indicated that DOX significantly downregulated endogenous H2S production and endogenous synthetase cystathionine γ-lyase (CSE) expression and obviously stimulated the apoptosis in H9C2 cells. The supplement of H2S donor sodium hydrosulfide (NaHS) or overexpression of CSE inhibited DOX-induced H9C2 cell apoptosis. DOX enhanced the activities of caspase family members in cardiomyocytes, while NaHS attenuated DOX-enhanced caspase-3, caspase-2, and caspase-9 activities by 223.1%, 73.94%, and 52.29%, respectively. Therefore, taking caspase-3 as a main target, we demonstrated that NaHS or CSE overexpression alleviated the cleavage of caspase-3, suppressed caspase-3 activity, and inhibited the cleavage of poly ADP-ribose polymerase (PARP). Mechanistically, we found that H2S persulfidated caspase-3 in H9C2 cells and human recombinant caspase-3 protein, while the thiol-reducing agent dithiothreitol (DTT) abolished H2S-induced persulfidation of caspase-3 and thereby prevented the antiapoptotic effect of H2S on caspase-3 in H9C2 cells. The mutation of caspase-3 C148S and C170S failed to block caspase-3 persulfidation by H2S in H9C2 cells. However, caspase-3 C163S mutation successfully abolished the effect of H2S on caspase-3 persulfidation and the corresponding protection of H9C2 cells. Collectively, these findings indicate that endogenous H2S persulfidates caspase-3 at cysteine 163, inhibiting its activity and cardiomyocyte apoptosis. Sufficient endogenous H2S might be necessary for the protection against myocardial cell apoptosis induced by DOX. The results of the study might open new avenues with respect to the therapy of DOX-stimulated cardiomyopathy.

The expression of hydrogen sulfide-producing enzymes in primary and lung metastatic osteosarcoma.

BACKGROUND: Hydrogen sulfide (H2S) is a novel gas transmitter signaling molecule. H2S is synthesized by cystathionine ß-synthase (CBS), cystathionine γ-lyase (CSE), and 3-mercaptopyruvate sulfurtransferase (MST). There have been no reports about the roles of these enzymes in osteosarcoma and its metastases. We detected H2S synthase expression levels in human primary osteosarcoma and lung metastatic osteosarcoma. METHODS: Immunohistochemistry was performed in primary osteosarcoma (n=19), lung metastatic osteosarcoma (n=11), osteoblastoma (n=10) and bony callus (n=2). The expression of CBS, CSE, and MST was defined as negative, moderately positive and strongly positive. RESULTS: MST staining was moderately to strongly positive in all cases. CSE staining was negative in 94.7% (18/19) of primary osteosarcoma cases and 90.9% (10/11) of lung metastatic osteosarcoma cases. CBS staining was strongly positive in 68.4% (13/19) of primary osteosarcoma cases, moderately positive in 15.8% (3/19) of cases, and negative in 15.8% (3/19) of cases. In lung metastatic osteosarcoma, the proportions of negative, moderately positive and strongly positive cases were 63.6% (7/11), 18.2% (2/11) and 18.2% (2/11), respectively. CONCLUSIONS: CBS and CSE expression, especially CSE expression, decreased in both primary osteosarcoma and lung metastatic osteosarcoma, which may suggest that CBS and CSE play roles in osteoblast cell malignant transformation and osteosarcoma progression. These enzymes could be used as new prognostic assessment factors and may represent new therapeutic targets for osteosarcoma and metastasis prevention.

Nitric oxide inhibits endothelial cell apoptosis by inhibiting cysteine-dependent SOD1 monomerization.

Endothelial cell apoptosis is an important pathophysiology in many cardiovascular diseases. The gasotransmitter nitric oxide (NO) is known to regulate cell survival and apoptosis. However, the mechanism underlying the effect of NO remains unclear. In this research, by targeting cytosolic copper/zinc superoxide dismutase (SOD1) monomerization, we aimed to explore how NO inhibited endothelial cell apoptosis. We showed that treatment with the NO synthase (NOS) inhibitor nomega-nitro-l-arginine methyl ester hydrochloride (L-NAME) significantly decreased the endogenous NO content of endothelial cells, facilitated the formation of SOD1 monomers, inhibited dismutase activity, and promoted reactive oxygen species (ROS) accumulation in human umbilical vein endothelial cells (HUVECs); by contrast, supplementation with the NO donor sodium nitroprusside (SNP) upregulated NO content, prevented the formation of SOD1 monomers, enhanced dismutase activity, and reduced ROS accumulation in L-NAME-treated HUVECs. Mechanistically, tris(2-carboxyethyl) phosphine hydrochloride (TCEP), a specific reducer of cysteine thiol, increased SOD1 monomer formation, thus preventing the NO-induced increase in dismutase activity and the decrease in ROS. Furthermore, SNP inhibited HUVEC apoptosis caused by the decrease in endogenous NO, whereas TCEP abolished this protective effect of SNP. In summary, our data reveal that NO protects endothelial cells against apoptosis by inhibiting cysteine-dependent SOD1 monomerization to enhance SOD1 activity and inhibit oxidative stress.

Endogenous Taurine Downregulation Is Required for Renal Injury in Salt-Sensitive Hypertensive Rats via CBS/H2S Inhibition.

Although taurine is known to exert an antihypertensive effect, it is unclear whether it is involved in the mechanism for hypertension-related target organ injury. To reveal the role of endogenous taurine in renal injury formation during salt-sensitive hypertension and clarify its mechanisms, both salt-sensitive Dahl rats and salt-resistant SS-13BN rats were fed a high-salt diet (8% NaCl) and given 2% taurine for 6 weeks. Rat systolic blood pressure (SBP) was measured by the tail-cuff method and artery catheterization. Kidney ultrastructure was observed under an electron microscope. Taurine content and mRNA and protein levels of taurine synthases, cysteine dioxygenase type 1 (CDO1) and cysteine sulfinic acid decarboxylase (CSAD), were decreased in Dahl rats fed a high-salt diet. However, taurine supplementation and the resulting increase in renal taurine content reduced the increased SBP and improved renal function and structural damage in high-salt diet-fed Dahl rats. In contrast, taurine did not affect SS-13BN SBP and renal function and structure. Taurine intervention increased the renal H2S content and enhanced cystathionine-ß-synthase (CBS) expression and activity in Dahl rats fed a high-salt diet. Taurine reduced the renin, angiotensin II, and aldosterone contents and the levels of oxidative stress indices in Dahl rat renal tissues but increased antioxidant capacity, antioxidant enzyme activity, and protein expression. However, taurine failed to achieve this effect in the renal tissue of SS-13BN rats fed a high-salt diet. Pretreatment with the CBS inhibitor HA or renal CBS knockdown inhibited H2S generation and subsequently blocked the effect of taurine on renin, superoxide dismutase 1 (SOD1), and superoxide dismutase 2 (SOD2) levels in high-salt-stimulated Dahl renal slices. In conclusion, the downregulation of endogenous taurine production resulted in a decrease in the renal CBS/H2S pathway. This decrease subsequently promoted renin-angiotensin-aldosterone system (RAAS) activation and oxidative stress in the kidney, ultimately contributing to renal injury in salt-sensitive Dahl rats.

Endogenous sulfur dioxide is a novel inhibitor of hypoxia-induced mast cell degranulation.

Introduction: Mast cell (MC) degranulation is an important step in the pathogenesis of inflammatory reactions and allergies; however, the mechanism of stabilizing MC membranes to reduce their degranulation is unclear. Methods: SO2 content in MC culture supernatant was measured by HPLC-FD. The protein and mRNA expressions of the key enzymes aspartate aminotransferase 1 (AAT1) and AAT2 and intracellular AAT activity were detected. The cAMP level in MCs was detected by immunofluorescence and ELISA. The release rate of MC degranulation marker ß-hexosaminidase was measured. The expression of AAT1 and cAMP, the MC accumulation and degranulation in lung tissues were detected. Objectives: To exam whether an endogenous sulfur dioxide (SO2) pathway exists in MCs and if it serves as a novel endogenous MC stabilizer. Results: We firstly show the existence of the endogenous SO2/AAT pathway in MCs. Moreover, when AAT1 was knocked down in MCs, MC degranulation was significantly increased, and could be rescued by a SO2 donor. Mechanistically, AAT1 knockdown decreased the cyclic adenosine monophosphate (cAMP) content in MCs, while SO2 prevented this reduction in a dose-independent manner. Pretreatment with the cAMP-synthesizing agonist forskolin or the cAMP degradation inhibitor IBMX significantly blocked the increase in AAT1 knockdown-induced MC degranulation. Furthermore, in hypoxia-stimulated MCs, AAT1 protein expression and SO2 production were markedly down regulated, and MC degranulation was activated, which were blunted by AAT1 overexpression. The cAMP synthesis inhibitor SQ22536 disrupted the suppressive effect of AAT1 overexpression on hypoxia-induced MC degranulation. In a hypoxic environment, mRNA and protein expression of AAT1 was significantly reduced in lung tissues of rats. Supplementation of SO2 elevated the cAMP level and reduced perivascular MC accumulation and degranulation in lung tissues of rats exposed to a hypoxic environment in vivo. Conclusion: SO2 serves as an endogenous MC stabilizer via upregulating the cAMP pathway under hypoxic circumstance.

Endogenous SO2-dependent Smad3 redox modification controls vascular remodeling.

Sulfur dioxide (SO2) has emerged as a physiological relevant signaling molecule that plays a prominent role in regulating vascular functions. However, molecular mechanisms whereby SO2 influences its upper-stream targets have been elusive. Here we show that SO2 may mediate conversion of hydrogen peroxide (H2O2) to a more potent oxidant, peroxymonosulfite, providing a pathway for activation of H2O2 to convert the thiol group of protein cysteine residues to a sulfenic acid group, aka cysteine sulfenylation. By using site-centric chemoproteomics, we quantified >1000 sulfenylation events in vascular smooth muscle cells in response to exogenous SO2. Notably, ~42% of these sulfenylated cysteines are dynamically regulated by SO2, among which is cysteine-64 of Smad3 (Mothers against decapentaplegic homolog 3), a key transcriptional modulator of transforming growth factor ß signaling. Sulfenylation of Smad3 at cysteine-64 inhibits its DNA binding activity, while mutation of this site attenuates the protective effects of SO2 on angiotensin II-induced vascular remodeling and hypertension. Taken together, our findings highlight the important role of SO2 in vascular pathophysiology through a redox-dependent mechanism.

Hydrogen sulfide regulates insulin secretion and insulin resistance in diabetes mellitus, a new promising target for diabetes mellitus treatment? A review.

BACKGROUND: Insulin resistance and impaired insulin secretion lead to disorders of glucose metabolism, which contributes to the development of diabetes. Hydrogen sulfide (H2S), a novel gasotransmitter, is found to play important roles in regulation of glucose metabolism homeostasis. AIM OF REVIEW: This study aimed to summarize and discuss current data about the function of H2S in insulin secretion and insulin resistance regulation as well as the underlying mechanisms. KEY SCIENTIFIC CONCEPTS OF REVIEW: H2S could be endogenously produced in islet ß cells, liver, adipose, skeletal muscles, and the hypothalamus, and regulates local and systemic glucose metabolism. It is reported that H2S suppresses insulin secretion, promotes or reduces the apoptosis of islet ß cells. It plays important roles in the regulation of insulin sensitivity in insulin responsive tissues. H2S inhibits glucose uptake and glycogen storage, and promotes or inhibits gluconeogenesis, mitochondrial biogenesis and mitochondrial bioenergetics in the liver. In adipose tissue, several investigators indicated that H2S promoted glucose uptake in adipocytes, while other studies reported that H2S inhibits this process. H2S has also been shown to promote adipogenesis, inhibit lipolysis, and regulate adiponectin and MCP-1 secretion from adipocytes. In skeletal muscle, H2S increases glucose uptake and improves insulin sensitivity. It is also observed that H2S modulates circadian-clock genes in muscle. Hypothalamic CBS/H2S pathway reduces obesity and improves insulin sensitivity via the brain-adipose interaction. Most studies indicated plasma H2S levels decreased in diabetic patients. However, the mechanisms by which H2S regulates systemic glucose metabolism remain unclear. Whether H2S acts as a new promising target for diabetes mellitus treatment merits further studies.

Persulfidation of transcription factor FOXO1 at cysteine 457: A novel mechanism by which H2S inhibits vascular smooth muscle cell proliferation.

INTRODUCTION: The proliferation of vascular smooth muscle cells (VSMCs) is an important physiological and pathological basis for many cardiovascular diseases. Endogenous hydrogen sulfide (H2S), the third gasotransmitter, is found to preserve vascular structure by inhibiting VSMC proliferation. However, the mechanism by which H2S suppresses VSMC proliferation has not been fully clear. OBJECTIVES: This study aimed to explore whether H2S persulfidates the transcription factor FOXO1 to inhibit VSMC proliferation. METHODS: After the proliferation of VSMC A7r5 cells was induced by endothelin-1 (ET-1), FOXO1 phosphorylation and proliferating cell nuclear antigen (PCNA) expression were detected by Western blotting, the degree of FOXO1 nuclear exclusion and PCNA fluorescent signals in the nucleus were detected by immunofluorescence, and the persulfidation of FOXO1 was measured through a biotin switch assay. RESULTS: The results showed that ET-1 stimulation increased cell proliferation, FOXO1 phosphorylation and FOXO1 nuclear exclusion to the cytoplasm in the cells. However, pretreatment with NaHS, an H2S donor, successfully abolished the ET-1-induced increases in the VSMC proliferation, FOXO1 phosphorylation, and FOXO1 nuclear exclusion to the cytoplasm. Mechanistically, H2S persulfidated the FOXO1 protein in A7r5 and 293T cells, and the thiol reductant DTT reversed this effect. Furthermore, the C457S mutation of FOXO1 abolished the H2S-induced persulfidation of FOXO1 in the cells and the subsequent inhibitory effects on FOXO1 phosphorylation at Ser256, FOXO1 nuclear exclusion to the cytoplasm and cell proliferation. CONCLUSION: Thus, our findings demonstrated that H2S might inhibit VSMC proliferation by persulfidating FOXO1 at Cys457 and subsequently preventing FOXO1 phosphorylation at Ser256.