Role Of Endogenous Hydrogen Sulfide Research Articles

Differences in nonoxidative sulfur metabolism between normal human breast MCF-12A and adenocarcinoma MCF-7 cell lines

Recent studies have revealed the role of endogenous hydrogen sulfide (H2S) in the development of breast cancer. The capacity of cells to generate H2S and the activity and expression of the main enzymes (cystathionine beta synthase; CBS, cystathionase γ-lyase; CGL, 3-mercaptopyruvate sulfurtransferase; MPST and thiosulfate sulfurtransferase; TST) involved in H2S metabolism were analyzed using an in vitro model of a non-tumourigenic breast cell line (MCF-12A) and a human breast adenocarcinoma cell line (MCF-7). In both cell lines, MPST, CGL, and TST expression was confirmed at the mRNA (RT-PCR) and the protein (Western Blot) level, while CBS expression was detected only in MCF-7 cells. Elevated levels of GSH, sulfane sulfur and increased CBS and TST activity were presented in the MCF-7 compared to the MCF-12A cells. It appears that cysteine might be mainly a substrate for GSH synthesis in breast adenocarcinoma. Increased capacity of the cells to generate H2S was shown for MCF-12A compared to MCF-7 cell line. Results suggest an important function of CBS in H2S metabolism in breast adenocarcinoma. The presented work may contribute to further research on new therapeutic possibilities for breast cancer - one of the most frequently diagnosed types of cancer among women.

Regulation of Na+/H+ antiport system and nitrogen metabolism by melatonin and endogenous hydrogen sulfide confers resilience to drought and salt stress

The present study was conducted with the purpose of examining the role of endogenous hydrogen sulfide (H2S) in melatonin (Mel)-induced regulation of tolerance mechanisms to drought stress and salinity. The drought stress was induced by maintaining soil water level at 40% and salinity was induced by applying 100 mM sodium chloride (NaCl). The results demonstrate an augmentation in the degree of activation of the antioxidant defense mechanisms in response to drought stress and salinity. However, a significant decline in the K+/Na+ ratio, accompanied by an elevation in reactive oxygen species, lipid peroxidation, and electrolyte leakage, was witnessed as well. A remarkable decrease in the level of hydration and content of chlorophyll was also observed under the imposed stress. The reported findings indicate that the administration of 30 µM Mel effectively regulated the activity of the Na+-H+ antiporter and H+-ATPase. This regulation ultimately led to an increase in K+ influx and a decline in Na+ efflux, resulting in an enhanced K+/Na+ ratio. Application of Mel also demonstrated an improvement in nitrogen metabolism, as well as the enzymatic and non-enzymatic antioxidant defense system. Additionally, Mel application caused a remarkable increase in the accumulation of osmolytes in drought stress and salinity -suffered plants. The synergistic effect of these characteristics resulted in the resilience of the Mel-supplemented plants to drought stress and salinity. However, application of hypotaurine (a scavenger of H2S) counteracted the beneficial impact of Mel which once again resulted in the same condition that was initially created by the imposed stress. These findings imply that Mel, in coordination with H2S was remarkable in inducing plant adaptive responses to drought stress and salinity.

Role of Endogenous Hydrogen Sulfide in Relaxation of the Lymph Node Capsule in LPS-induced Inflammation

Role of Endogenous Hydrogen Sulfide in Relaxation of the Lymph Node Capsule in LPS-induced Inflammation

Disrupted H2S Signaling by Cigarette Smoking and Alcohol Drinking: Evidence from Cellular, Animal, and Clinical Studies.

The role of endogenous hydrogen sulfide (H2S) as an antioxidant regulator has sparked interest in its function within inflammatory diseases. Cigarette and alcohol use are major causes of premature death, resulting from chronic oxidative stress and subsequent tissue damage. The activation of the Nrf2 antioxidant response by H2S suggests that this novel gasotransmitter may function to prevent or potentially reverse disease progression caused by cigarette smoking or alcohol use. The purpose of this study is to review the interrelationship between H2S signaling and cigarette smoking or alcohol drinking. Based on the databases of cellular, animal, and clinical studies from Pubmed using the keywords of H2S, smoking, and/or alcohol, this review article provides a comprehensive insight into disrupted H2S signaling by alcohol drinking and cigarette smoking-caused disorders. Major signaling and metabolic pathways involved in H2S-derived antioxidant and anti-inflammatory responses are further reviewed. H2S supplementation may prove to be an invaluable asset in treating or preventing diseases in those suffering from cigarette or alcohol addiction.

Dose dependent differential effects of toxic metal cadmium in tomato roots: Role of endogenous hydrogen sulfide

In this study, hydroponic experiments were conducted to elucidate mechanism(s) that are associated with differential effects of low (5 μM) and high (25 μM) dose of cadmium (Cd) stress in tomato. Furthermore, emphasis has also been focused on any involvement of endogenous hydrogen sulfide (H2S) in differential behaviour of low and high doses of Cd stress. At low dose of Cd, root growth i.e. root fresh weight, length and fitness did not significantly alter when compared to the control seedlings. Though at low dose of Cd, cellular accumulation of Cd was slightly increased but this was accompanied by higher endogenous H2S and phytochelatins, L-cysteine desulfhydrase (DES) activity, activities of glutathione biosynthetic and AsA-GSH cycle enzymes, and maintained redox status of ascorbate and glutathione. However, addition of hypotaurine (HT, a scavenger of H2S) resulted in greater toxicity, even at low dose of Cd, and these responses resembled with higher dose of Cd stress such as greater decline in root growth, endogenous H2S and phytochelatins, activities of DES, glutathione biosynthesis and AsA-GSH cycle enzymes, disturbed redox status of ascorbate and glutathione which collectively led to higher oxidative stress in tomato roots. Moreover, addition of HT with higher dose of Cd also further enhanced its toxicity. Collectively, the results showed that differential behaviour of low and high dose of Cd stress is mediated by differential regulation of biochemical attributes in which endogenous H2S has a crucial role.

Attenuation of cardiac ischemia-reperfusion injury by sodium thiosulfate is partially dependent on the effect of cystathione beta synthase in the myocardium.

Our early studies have shown that sodium thiosulfate (STS) treatment attenuated the ischemia-reperfusion (IR)-induced injury in an isolated rat heart model by decreasing apoptosis, oxidative stress, and preserving mitochondrial function. Hydrogen sulfide, the precursor molecule is reported to have similar efficacy. This study aims to investigate the role of endogenous hydrogen sulfide in STS-mediated cardioprotection against IR in an isolated rat heart model. D, L-propargylglycine (PAG), an inhibitor of cystathionine γ-lyase was used as endogenous H2S blocker. In addition, we used the hypoxia-reoxygenation (HR) model to study the impact of STS in cardiomyocytes (H9C2) and fibroblast (3T3) cells. STS treatment to animal and cells prior to IR or HR decreased cell injury. The sensitivity of H9C2 and 3T3 cells towards HR (6 h hypoxia followed by 12 h reoxygenation) challenge varies, where, 3T3 cells exhibited higher cell death (54%). Cells treated with PAG prior to STS abrogate the protective effect in 3T3 (cell viability 61%) but not in H9C2 (cell viability 82%). Further evaluation in rat heart model showed partial recovery (46% RPP) of heart from those hearts pretreated with PAG prior to STS condition. In conclusion, we demonstrated that STS-mediated cardioprotection to IR-challenged rat heart is not fully dependent on endogenous H2S level and this dependency may be linked to higher fibroblast content in rat heart.

Role of endogenous hydrogen sulfide in cardiac mitochondrial preservation during ischemia reperfusion injury

Cardio-protective effect of hydrogen sulfide (H2S) against myocardial ischemia reperfusion injury (I/R) via preservation of mitochondria is well documented. But the distinct role of exogenous and endogenous H2S in cardio-protection and its dependency on functional cardiac mitochondria is not understood. The present study was designed to investigate the role of exogenous H2S preconditioning on cardiac mitochondrial subpopulation namely interfibrillar (IFM) and subsarcolemmal (SSM), in attenuating I/R injury in an isolated rat heart model in the absence of endogenous H2S production. The well-known inhibitor of endogenous H2S production DL-propargylglycine (cystathionine gamma lyase inhibitor) used for this purpose. Our previous studies revealed that NaSH (a H2S donor) preconditioning at a dose of 20μM significantly reduced the infarct size and preserved the functional activities of IFM. However, this protective effect significantly declined, when the heart preconditioned with NaSH in the presence of PAG. Apparently, cardiac recovery was improved hemodynamically to a minimal level by increased concentration of NaSH (40μM). The rat heart perfused with 2, 4 dinitrophenol a mitochondrial uncoupler, before NaSH preconditioning to understand the mitochondrial dependency of exogenous NaSH, we found that cardio-protective effect of NaSH was negated even with higher concentration of H2S. The result indicates that exogenous NaSH mediated cardio protection depends on functional cardiac mitochondria. However, no functional difference between the subpopulation i.e., interfibrillar and subsarcolemmal mitochondria observed with NaSH preconditioning in the presence of PAG.

Imbalance of Endogenous Hydrogen Sulfide and Homocysteine in Chronic Obstructive Pulmonary Disease Combined with Cardiovascular Disease.

Background: Considerable studies showed associations between chronic obstructive pulmonary disease (COPD) and cardiovascular disease (CVD), we evaluated the role of endogenous hydrogen sulfide (H2S)/homocysteine (Hcy) in patients with COPD combined with CVD.Methods: Fifty one stable patients with COPD were enrolled (25 COPD, 26 COPD + CVD). Lung function, sputum, peripheral blood samples, serum H2S, Hcy, high-sensitivity C-reactive protein (hs-CRP) and tumor necrosis factor-α (TNF-α) levels were measured. Dyspnea, symptoms and quality of life were quantified by modified Medical Research Council dyspnea scale (mMRC), COPD assessment test (CAT) and St. George’s Respiratory Questionnaire (SGRQ).Results: Compared with COPD group, waist circumference and body mass index (BMI) were higher in COPD + CVD group, mMRC, CAT and activity scores were also higher, high density lipoprotein cholesterol (HDL-C) was lower, total cells, neutrophils (%) in sputum and serum hs-CRP level were higher, whereas macrophages (% ) in sputum was lower. H2S and Hcy levels from COPD + CVD group were higher than those from COPD group, but H2S/Hcy ratio was lower. With increasing COPD severity, H2S level was decreased, however, Hcy level was increased. H2S level was positively correlated with FEV1/FVC, FEV1% predicted, lymphocytes (%) and macrophages (%) in sputum, but negatively correlated with smoking pack-years and neutrophils (%) in sputum. Hcy level was positively correlated with BMI and total cells in sputum. The ratio of H2S/Hcy was also positively correlated with FEV1/FVC, but negatively correlated with total cells in sputum.Conclusion: The imbalance of H2S/Hcy may be involved in the pathogenesis of COPD combined with CVD and provide novel targets for therapy.

Role of endogenous hydrogen sulfide in nerve-evoked relaxation of pig terminal bronchioles

Hydrogen sulfide (H2S) is a gasotransmitter employed for intra- and inter-cellular communication in almost all organ systems. This study investigates the role of endogenous H2S in nerve-evoked relaxation of pig terminal bronchioles with 260 μm medium internal lumen diameter. High expression of the H2S synthesis enzyme cystathionine γ-lyase (CSE) in the bronchiolar muscle layer and strong CSE-immunoreactivity within nerve fibers distributed along smooth muscle bundles were observed. Further, endogenous H2S generated in bronchiolar membranes was reduced by CSE inhibition. In contrast, cystathionine β-synthase expression, another H2S synthesis enzyme, however was not consistently detected in the bronchiolar smooth muscle layer. Electrical field stimulation (EFS) and the H2S donor P-(4-methoxyphenyl)-P-4-morpholinylphosphinodithioic acid (GYY4137) evoked smooth muscle relaxation. Inhibition of CSE, nitric oxide (NO) synthase, soluble guanylyl cyclase (sGC) and of ATP-dependent K+, transient receptor potential A1 (TRPA1) and transient receptor potential vanilloid 1 (TRPV1) channels reduced the EFS relaxation but failed to modify the GYY4137 response. Raising extracellular K+ concentration inhibited the GYY4137 relaxation. Large conductance Ca2+-activated K+ channel blockade reduced both EFS and GYY4137 responses. GYY4137 inhibited the contractions induced by histamine and reduced to a lesser extent the histamine-induced increases in intracellular [Ca2+]. These results suggest that relaxation induced by EFS in the pig terminal bronchioles partly involves the H2S/CSE pathway. H2S response is produced via NO/sGC-independent mechanisms involving K+ channels and intracellular Ca2+ desensitization-dependent pathways. Thus, based on our current results H2S donors might be useful as bronchodilator agents for the treatment of lung diseases with persistent airflow limitation, such as asthma and chronic obstructive lung disease.

Endogenously produced hydrogen sulfide is involved in porcine oocyte maturation in vitro

Hydrogen sulfide, one of three known gasotransmitters, is involved in physiological processes, including reproductive functions. Oocyte maturation and surrounding cumulus cell expansion play an essential role in female reproduction and subsequent embryonic development. Although the positive effects of exogenous hydrogen sulfide on maturing oocytes are well known, the role of endogenous hydrogen sulfide, which is physiologically released by enzymes, has not yet been described in oocytes. In this study, we observed the presence of Cystathionine β-Synthase (CBS), Cystathionine γ-Lyase (CTH) and 3-Mercaptopyruvate Sulfurtransferase (3-MPST), hydrogen sulfide-releasing enzymes, in porcine oocytes. Endogenous hydrogen sulfide production was detected in immature and matured oocytes as well as its requirement for meiotic maturation. Individual hydrogen sulfide-releasing enzymes seem to be capable of substituting for each other in hydrogen sulfide production. However, meiosis suppression by inhibition of all hydrogen sulfide-releasing enzymes is not irreversible and this effect is a result of M-Phase/Maturation Promoting Factor (MPF) and Mitogen-Activated Protein Kinase (MAPK) activity inhibition. Futhermore, cumulus expansion expressed by hyaluronic acid (HA) production is affected by the inhibition of hydrogen sulfide production. Moreover, quality changes of the expanded cumuli are indicated. These results demonstrate hydrogen sulfide involvement in oocyte maturation as well as cumulus expansion. As such, hydrogen sulfide appears to be an important cell messenger during mammalian oocyte meiosis and adequate cumulus expansion.

Role of hydrogen sulfide in early blood-brain barrier disruption following transient focal cerebral ischemia.

We determined the role of endogenous hydrogen sulfide (H₂S) in cerebral vasodilation/hyperemia and early BBB disruption following ischemic stroke. A cranial window was prepared over the left frontal, parietal and temporal cortex in mice. Transient focal cerebral Ischemia was induced by directly ligating the middle cerebral artery (MCA) for two hours. Regional vascular response and cerebral blood flow (CBF) during ischemia and reperfusion were measured in real time. Early BBB disruption was assessed by Evans Blue (EB) and sodium fluorescein (Na-F) extravasation at 3 hours of reperfusion. Topical treatment with DL-propargylglycine (PAG, an inhibitor for cystathionine γ-lyase (CSE)) and aspartate (ASP, inhibitor for cysteine aminotransferase/3-mercaptopyruvate sulfurtransferase (CAT/3-MST)), but not O-(Carboxymethyl)hydroxylamine hemihydrochloride (CHH, an inhibitor for cystathionine β-synthase (CBS)), abolished postischemic cerebral vasodilation/hyperemia and prevented EB and Na-F extravasation. CSE knockout (CSE-/-) reduced postischemic cerebral vasodilation/hyperemia but only inhibited Na-F extravasation. An upregulated CBS was found in cerebral cortex of CSE-/- mice. Topical treatment with CHH didn’t further alter postischemic cerebral vasodilation/hyperemia, but prevented EB extravasation in CSE-/- mice. In addition, L-cysteine-induced hydrogen sulfide (H2S) production similarly increased in ischemic side cerebral cortex of control and CSE-/- mice. Our findings suggest that endogenous production of H2S by CSE and CAT/3-MST during reperfusion may be involved in postischemic cerebral vasodilation/hyperemia and play an important role in early BBB disruption following transient focal cerebral ischemia.

P45 Endogenous hydrogen sulfide suppresses intestinal inflammation through the regulation of macrophage polarization

Introduction Accumulating evidence shows the role of endogenous hydrogen sulfide (H 2 S) in various disease, whereas its role of H 2 S in intestinal inflammation. In addition, recent reports showed that H 2 S modulates NF-kB signaling in macrophages, suggesting that H 2 S could affect macrophage function. In this study, we studied the role of endogenous H 2 S in a dextran sodium sulfate (DSS)-induced colitis model, particularly in relation to macrophage function. Methods Mice were treated with 8% DSS to induce colitis. DL-propargylglycine (PAG), an irreversible CSE (cystathionine gamma-lyase) inhibitor, and sodium sulfide (Na 2 S), an H 2 S donor, were administered intraperitoneally. The respective degrees of mucosal injury were evaluated macroscopically, histologically, and biochemistry. The effect of endogenous H 2 S on macrophage function was evaluated using bone-marrow derived macrophages (BMDMs). Results CSE and CBS (cystathionine beta-synthase) expressions in the colonic mucosa were increased in DSS-treated mice. PAG enhanced DSS-induced colonic damage and myeloperoxidase activity. Meanwhile, Na 2 S counteracted these effects of PAG. Furthermore, PAG-treated BMDMs manifested pro-inflammatory (M1), but not anti-inflammatory (M2) macrophages markers. Conclusion Our results demonstrated that endogenous H 2 S inhibited the development of intestinal inflammation through the regulation of macrophage function. In conclusion, H 2 S may be a novel therapeutic molecule in intestinal inflammation.

Role of endogenous hydrogen sulfide of the spinal cord in the maintenance of diabetic neuropathic pain in rats

Objective To evaluate the role of endogenous hydrogen sulfide in the maintenance of diabetic neuropathic pain (DNP) in rats.Methods The DNP models were established by intraperitoneal injection of stretopzin (STZ) in 24 male SD rats aged 4 weeks (150-170 g),and other 24 rats were intraperitoneally injected with the same volume of normal saline as normal group.All the rats were implanted intrathecal catheter.The rats with DNP were randomly divided into 2 subgroups (n =12 each):saline group (group D) and methemoglobin group (group DM).The rats in normal group were randomly divided into 2 subgroups (n =12 each):saline group (group C) and methemoglobin group (group CM).Three weeks after intraperitoneal injection,the rats were separately intrathecally administered methemoglobin 10 μ mol/L (10 μ1) (group DM),saline 10 μl (group D),methemoglobin 10 μmol/L (10 μl) (group CM) and saline 10 μl (group C),respectively,once a day for 4 days.The paw withdrawal mechanical threshold (PWT) of the rats were measured before and after injection at 1 (T1),2 (T2),3 (T3),4 (T4) days,then the rats were sacrificed at T4 point after the measurement and the lumbar segments of the spinal cord were removed for determination of N-methyl-D-aspartate receptor subunit 2B (NR2B) protein expression (by Western blotting) and NR2B mRNA expression [by reverse transcriptase-polymerase chain reaction (RT-PCR)].Results As compared with group C and group CM,the PWT in group D and group DM was significantly reduced (P ＜ 0.05),while the expression of NR2B protein and mRNA was significantly increased (P ＜ 0.05).As Compared with group D [NR2B mRNA (0.89 ± 0.09),NR2B protein (1.28 ± 0.17)],the PWT in group DM was significantly increased at T2-T4 point before the injection and T1-T4 point after the injection,while the expression of NR2B mRNA (0.69 ±0.11) and NR2B protein (0.98 ± 0.14) in group DM was significantly decreased (P ＜ 0.05).As compared with the point before the injection (3.8 ± 0.9 and 6.1 ± 1.4),the PWT in group DM were significantly increased at T1-T2 point after the injection (7.5 ± 1.8 and 8.7 ± 1.6) (P ＜ 0.05).Conclusion Endogenous hydrogen sulfide in the spinal cord is involved in the maintenance of DNP in rats and the possible mechanisms were related with up-regulation of NR2B in the spinal dorsal horn. Key words: Hydrogen sulfide; N-methyl-D-aspartate receptor subunit 2B; Spinal dorsal horn; Diabete ; Pain

Role of endogenous hydrogen sulfide on hypoxia‐induced hypothermia in hypertensive rats (LB792)

Role of endogenous hydrogen sulfide on hypoxia‐induced hypothermia in hypertensive rats. Joao Paulo J. Sabino1, Renato N. Soriano1, Guillermo A. Ariza Traslaviña2, Luiz G. S. Branco1. 1Dental School and 2Medical School of Ribeirão Preto, University of São Paulo.Hypoxia causes a regulated drop in body temperature (Tb) but the mechanisms involved are not completely understood. The role of hydrogen sulfide (H2S) a gaseous molecule endogenously produced by cystathionine β‐synthase (CBS) has been recently documented during hypoxia‐induced hypothermia in normotensive rats. However, no information exists regarding the role of H2S during hypoxic hypothermia in spontaneous hypertensive rats (SHR). We then evaluated the effect of the H2S donor (Na2S 4 nmol/1µl) and the CBS inhibitor (AOAA 9 nmol/1µl) microinjections into the central nervous system (fourth ventricle) of rats exposed to 10% hypoxia. The Tb was similar in all groups [Saline Wistar group (37.46±0.18 ºC, n= 10), AOAA Wistar group (37.40±0.33 ºC, n= 5), Na2S Wistar group (37.52±0.13 ºC, n= 7), Saline SHR group (37.43±0.43 ºC, n= 8), AOAA SHR group (37.59±0.16 ºC, n= 10) and Na2S SHR group (37.72±0.15 ºC, n= 6)] during basal normoxic condition. Hypoxia induced a typical decrease in Tb in saline injected rats. The decrease in Tb in the Saline SHR group (Δ= ‐1.16±0.11 ºC) was similar to the Saline Wistar group (Δ= ‐0.81±0.20 ºC). In Wistar rats, Na2S administration prevented hypoxia‐induced hypothermia (Δ= ‐0.58±0.16 ºC), whereas AOAA administration significantly increased hypoxia‐induced hypothermia in Wistar rats (Δ= ‐1.60±0.20 ºC), indicating that H2S plays a key role on hypoxia‐induced changes in Tb. Surprisingly, neither Na2S (Δ= ‐1.33±0.20 ºC) nor AOAA (Δ= ‐1.61±0.21 ºC) caused any change in hypoxia‐induced hypothermia in SHR. In conclusion, the H2S prevents the hypoxia‐induced anapyrexia in normotensive rats, but plays no thermoregulatory role during hypoxia exposition in SHR.Financial support: FAPESP (2011/14779‐4).

Hydrogen Sulfide Is an Endogenous Regulator of Aging in Caenorhabditis elegans

To investigate the role of endogenous hydrogen sulfide (H2S) in the control of aging and healthspan of Caenorhabditis elegans. We show that the model organism, C. elegans, synthesizes H2S. Three H2S-synthesizing enzymes are present in C. elegans, namely cystathionine γ lyase (CSE), cystathionine β synthetase, and 3-mercaptopyruvate transferase (MPST or 3-MST). Genetic deficiency of mpst-1 (3-MST orthologue 1), but not cth-2 (CSE orthologue), reduced the lifespan of C. elegans. This effect was reversed by a pharmacological H2S donor (GYY4137). GYY4137 also reduced detrimental age-dependent changes in a range of physiological indices, including pharyngeal contraction and defecation. Treatment of C. elegans with GYY4137 increased the expression of several age-related, stress response, and antioxidant genes, whereas MitoSOX Red fluorescence, indicative of reactive oxygen species generation, was increased in mpst-1 knockouts and decreased by GYY4137 treatment. GYY4137 additionally increased the lifespan in short-lived mev-1 mutants with elevated oxidative stress and protected wild-type C. elegans against paraquat poisoning. The lifespan-prolonging and health-promoting effects of H2S in C. elegans are likely due to the antioxidant action of this highly cell-permeable gas. The possibility that novel pharmacological agents based on the principle of H2S donation may be able to retard the onset of age-related disease by slowing the aging process warrants further study. Our results show that H2S is an endogenous regulator of oxidative damage, metabolism, and aging in C. elegans and provide new insight into the mechanisms, which control aging in this model organism.

Down-Regulation of Hydrogen Sulfide Biosynthesis Accompanies Murine Interstitial Cells of Cajal Dysfunction in Partial Ileal Obstruction

PurposeTo investigate the role of endogenous hydrogen sulfide (H2S) in partial obstruction-induced dysfunction of the interstitial cells of Cajal (ICC) in mice ileum.Materials and MethodsPartial intestinal obstruction was induced surgically in male imprinting control region (ICR) mice. ICC networks were studied by Immunohistochemistry. Electrical activity was recorded by intracellular recording techniques. The expression of ICC phenotype marker c-kit receptor tyrosine kinase (c-kit), membrane binding stem cell factor (mSCF), the endogenous H2S biosynthesis enzymes cystathionine-β-synthase (CBS) and cystathionine-γ-lyase (CSE) was studied by Western blotting. The expression of tumor necrosis factor-α (TNF-α) mRNA was observed by using real-time polymerase chain reaction.ResultsPartial intestinal obstruction resulted in ICC networks were disrupted above obstruction 14 days after the operation. The slow waves of intestinal smooth muscles in the dilated region were significantly suppressed and their amplitude and frequency were reduced, whilst the resting membrane potentials were depolarized. The expression of c-kit and mSCF was significantly decreased, also suggesting the disruption of the ICC network. The expression of TNF-α was significantly increased in the tunica muscularis of the obstructed intestine. Treatment of cultured intestinal smooth muscle cells with TNF-α caused dramatic down regulation of mSCF. The expression of CBS and CSE was significantly decreased in the tunica muscularis of the obstructed intestine. Intraperitoneal injection (i.p) of DL-propargylglycine, an irreversible inhibitor of CSE, and aminooxyacetic acid, an inhibitor of CBS, elevated the expression of TNF-α mRNA in the tunica muscularis of the ileum. Obstruction-induced over expression of TNF-α was significantly improved by supplementation of NaHS, but not the expressions of mSCF and c-kit.ConclusionsThe down regulation of endogenous H2S biosynthesis is related to over expression of TNF-α in obstructed small intestine. TNF-α-mediated mSCF down-regulation is not the only reason of partial intestinal obstruction-induced loss of ICC.

The role of endogenous hydrogen sulfide in pathogenesis of chronotropic dysfunction in rats with cirrhosis

Endogenous hydrogen sulfide is produced by cystathionine-γ-lyase and cystathionine-β-synthase in a variety of tissues and has recently been implicated in the regulation of cardiac functions. Acceleration of the heart rate in response to catecholamines is impaired in patients with cirrhosis. The present study was aimed to examine the role of endogenous hydrogen sulfide in the pathogenesis of chronotropic dysfunction in rats with cirrhosis. Cirrhosis was induced by surgical ligation of bile duct in rats. There was no significant difference in atrial cystathionine-γ-lyase and cystathionine-β-synthase mRNA levels in control and cirrhotic rats as assessed by quantitative RT-PCR. Four weeks after bile duct ligation or sham surgery the atria were isolated and chronotropic responsiveness to adrenergic stimulation was assessed using standard organ bath. Incubation of the atria with propargylglycine (PAG, a cystathionine-γ-lyase inhibitor) and amino-oxyacetic acid (AOAA, a cystathionine-β-synthase inhibitor) was associated with a significant desensitization of chronotropic response to adrenergic stimulation in controls rats. This indicates that endogenous hydrogen sulfide might be involved in modulation of adrenergic signaling in the atrium. Bile duct ligation was associated with impaired chronotropic responsiveness to adrenergic stimulation in comparison with sham-operated rats. In contrast to control group, incubation of the atria with PAG and AOAA was able to partially improve the chronotropic responsiveness to adrenergic stimulation in cirrhotic rats. Our data shows that local inhibition of endogenous hydrogen sulfide in atria has opposite effect in cirrhotic versus control rats and may play a role in physiological modulation of adrenergic signaling in the atrium.

Role of endogenous hydrogen sulfide in neurogenic relaxation of rat corpus cavernosum

Relaxation of corpus cavernosum during penile erection is mediated by a non-adrenergic non-cholinergic (NANC) neurotransmission and by the endothelium via the release of nitric oxide. Hydrogen sulfide (H2S) is an endogenous gaseous mediator which is a potent vasodilator and a neurotransmitter. This study was initiated to characterize the role of H2S in NANC neurogenic transmission in rat corpus cavernosum. The expression of H2S producing enzymes was assessed using RT-PCR as well as Western blotting and showed the expression of cystathionine γ-lyase (CSE) in rat corporal tissue. Homogenates from rat corpus cavernosum convert l-cysteine to H2S and this was partially inhibited by a CSE inhibitor, propargylglycine. Electrical stimulation of corporal tissue strips caused NANC relaxation. This neurogenic relaxation was significantly enhanced by inhibition of CSE by propargylglycine indicating that endogenously produced H2S may have a negative regulatory role in neurogenic relaxation of rat corpus cavernosum. To investigate this further we used physiologically relevant concentrations of exogenous NaHS, and showed that nanomolar concentrations could inhibit corporal relaxation induced by a nitroxyl (HNO) donor (Angeli's salt) but not with nitrosonium (NO+) or NO donors. This suggests that an interaction between endogenously produced H2S and nitroxyl (HNO) might be involved in erectile function.

Role of endogenous hydrogen sulfide on renal damage induced by adriamycin injection

A single injection of adriamycin (ADR) induces marked and persistent proteinuria in rats that progress to glomerular and tubulointerstitial lesions. It has been shown that ADR-induced nephrotoxicity is mediated, at least in part, by oxidative stress that lead to inflammation. Endogenous hydrogen sulfide (H₂S) is synthesized from L-cysteine and is an important signaling molecule in inflammation. This study evaluates the effect of DL-propargylglycine (PAG), an inhibitor of endogenous H₂S formation, on the evolution of renal damage induced by ADR. The rats were injected i.p. with 0.15 M NaCl or PAG (50 mg/kg) 2 h after ADR injection (3.5 mg/kg). Control rats were injected with 0.15 M NaCl or PAG only. Twenty hours urine samples were collected for albuminuria and creatinine measurements on days 1 and 14 after saline or ADR injections and on days 2 and 15 blood samples were collected to measure plasma creatinine, then the rats were killed. The kidneys were removed for H₂S formation evaluation, renal lipid peroxidation and glutathione levels, and histological and immunohistochemical analysis. On day 2 after ADR injection the rats presented increase in oxidative stress associated with neutrophils and macrophages influx in renal tissue. On day 15 the rats also presented increased desmin expression at glomerular edge and vimentin in cortical tubulointerstitium, as well as albuminuria. All these alterations were reduced by PAG injection. The protective effect of PAG on ADR nephrotoxicity was associated to decreased H₂S formation and to restriction of oxidative stress and inflammation in the renal cortex.

Role of endogenous hydrogen sulfide in rats with hepatic cirrhosis at different stages

Role of endogenous hydrogen sulfide in rats with hepatic cirrhosis at different stages