Nitric oxide (NO) mediates various physiological and pathological processes, including cell proliferation, differentiation, and inflammation. Protein S-nitrosylation (SNO), a NO-mediated reversible protein modification, leads to changes in the activity and function of target proteins. Recent findings on protein-protein transnitrosylation reactions (transfer of an NO group from one protein to another) have unveiled the mechanism of NO modulation of specific signaling pathways. The intracellular level of S-nitrosoglutathione (GSNO), a major reactive NO species, is controlled by GSNO reductase (GSNOR), a major regulator of NO/SNO signaling. Increasing number of GSNOR-related studies have shown the important role that denitrosylation plays in cellular NO/SNO homeostasis and human pathophysiology. This review introduces recent evidence of GSNO-mediated NO/SNO signaling depending on GSNOR expression or activity. In addition, the applicability of GSNOR as a target for drug therapy will be discussed in this review.
Cell Proliferation ; Drug Therapy ; Homeostasis ; Humans ; Inflammation ; Nitric Oxide ; Oxidoreductases* ; Pathologic Processes ; S-Nitrosoglutathione*

BACKGROUND: Recently, the mechanism involved in nitric oxide (NO)-mediated motor neuron death is under extensive investigation. The role of Cu/Zn superoxide dismutase (SOD) mutation, which is found in about 2% of all ALS patients, has been implicated in selective motor neuron death and it is said to play an important role in NO-mediated motor neuron death. Estrogen is reported to have neuroprotective effect in various neurological diseases. However, neuroprotective effect on estradiol on spinal motor neuron exposed to NO has rarely been studied. METHODS: Motor neuron-neuroblastoma hybrid cell expressing wild-type or mutant (G93A or A4V) SOD gene was treated with 200 micro M Snitrosoglutathione. After 24 hours, cell viability was measured by MTT assay. To see the neuroprotective effect of estradiol, pretreatment with 5 nM or 50 nM 17 beta-estradiols was done 24 hours before S-nitrosoglutathione treatment. RESULTS: S-nitrosoglutathione showed significant neurotoxic effect in all three cell lines. Percentage of cell death was significantly different in each cell line. Both 5 nM and 50 nM estradiols showed neuroprotective effect in G93A cell line. In wild-type cell line, 50 nM estradiol showed neuroprotective effect, but 5 nM estradiol did not. In A4V cell line, estradiol did not showed neuroprotective effect. CONCLUSIONS: This study showed that NO-mediated motor neuron death could be influenced by presence or absence of mutation and type of mutation in SOD gene. Neuroprotective effect of estradiol is also influenced by SOD gene mutation. This study implies that estrogen might be beneficial to some ALS patients.
Cell Death ; Cell Line ; Cell Survival ; Estradiol ; Estrogens* ; Humans ; Hybrid Cells ; Motor Neurons* ; Neuroprotective Agents* ; Nitric Oxide ; S-Nitrosoglutathione ; Superoxide Dismutase* ; Superoxides*

The aim of this study was to investigate the influence of S-nitrosoglutathione (GSNO) on agglutination and nitric oxide (NO) concentration in frozen platelets. The agglutination of platelets was detected by using platelet agglutination apparatus, the level of NO in platelets was detected by the nitrate enzyme reduction method. The results showed that the rates of agglutination in freeze platelets and frozen platelets treated with GSNO were (35.47 ± 2.93) and (24.43 ± 3.07), which were significantly lower than that in fresh liquid platelets (63.44 ± 2.96). The level of NO concentration in frozen platelets was (22.16 ± 6.38), which was significantly lower than that in fresh liquid platelets (31.59 ± 16.88). The level of NO concentration in frozen platelets treated with GSNO was (45.64 ± 6.31), which was significantly higher than that in fresh liquid platelets (P < 0.01). It is concluded that GSNO increases the concentration of NO in frozen platelets, inhibits platelet activation and maintains platelet function, thus GSNO can be used as a frozen protective agent.
Blood Platelets ; drug effects ; Freezing ; Humans ; Nitric Oxide ; metabolism ; Platelet Activation ; drug effects ; Platelet Aggregation ; drug effects ; Platelet Count ; S-Nitrosoglutathione ; pharmacology

BACKGROUND: To determine whether nitric oxide (NO) could inhibit activation of platelets stored in a cold or frozen state, we measured platelet P-selectin expression and platelet-bound fibrinogen in platelet-rich plasma (PRP) with S-nitrosoglutathione (GSNO) (Sigma, USA) by flow cytometry. METHODS: PRP was prepared by centrifuging venous blood collected in a 3.2% sodium citrate tube from 10 healthy donors. It was aliquotted into 4 groups (no cryoprotectant, GSNO, GSNO/dimethyl sulfoxide [DMSO] [Sigma], and DMSO), and stored at room, cold and freezing temperatures for 24 hrs. We performed a flow cytometric analysis of all specimens stained with FITC-fibrinogen and PE-CD62P monoclonal antibodies (Becton Dickinson, USA). The results were compared according to the storage temperature and agonist among 4 groups. RESULTS: GSNO inhibited significantly the activation of frozen platelets, but not in the presence of DMSO. GSNO was also shown to preserve the aggregability of frozen platelets because in the presence of GSNO the delta percent change of P-selectin expression and fibrinogen binding of frozen platelets increased significantly irrelevant to DMSO. CONCLUSIONS: GSNO inhibited the activation of frozen platelets and preserved the platelet aggregability; therefore, it may be used as a protectant for platelet cryopreservation.
Adult ; Blood Platelets/*drug effects/metabolism ; Cryopreservation/*methods ; Female ; Fibrinogen/metabolism ; Flow Cytometry ; Free Radical Scavengers/pharmacology ; Humans ; Male ; Nitric Oxide/metabolism ; Nitric Oxide Donors/*pharmacology ; P-Selectin/metabolism ; Platelet Aggregation/drug effects ; Platelet Aggregation Inhibitors/pharmacology ; S-Nitrosoglutathione/*pharmacology

This study was purposed to explore the influence of S-nitrosoglutathione (GSNO) on membrane glycoprotein of frozen platelet. The levels of membrane glycoprotein on fresh liquid platelets, frozen platelets and frozen platelets with GSNO were measured by flow cytometry. The results showed that the GSNO obviously decreased platelet aggregation, the PAC-1 change in the three groups was not significant. The changes of CD42b and CD62P in fresh liquid platelet group, frozen platelet group and frozen platelets with GSNO were significant different. The change of membrane glycoprotein in above-mentioned three group was not significant. It is concluded that the GSNO inhibits platelet aggregation, maintains the function of platelets and may be used as a cryoprotectant. When frozen platelets were added with GSNO, the molecular rearrangement, structure change and other mechanism may occur in platelets.
Blood Platelets ; drug effects ; Blood Preservation ; methods ; Freezing ; Humans ; P-Selectin ; metabolism ; Platelet Activation ; drug effects ; Platelet Glycoprotein GPIb-IX Complex ; metabolism ; Platelet Membrane Glycoproteins ; metabolism ; S-Nitrosoglutathione ; pharmacology

To investigate the effect of S-nitrosoglutathione (GSNO), a nitric oxide donor, on platelet production from megakaryocytes differentiated from cord blood CD34(+) cells in vitro, the CD34 (+) cells from eight fresh umbilical cord blood samples by a high-gradient magnetic cell sorting (MACS) system were cultured in serum-free medium for 14 d with thrombopoietin (TPO) 50 ng/ml, IL-3 10 ng/ml, stem cell factor (SCF) 50 ng/ml and rHuGM-CSF 20 ng/ml. Then, CD61 (+) cells were purified by MACS system from these CD34 (+) cells, and were cultured in serum-free medium supplemented with TPO 50 ng/ml, IL-3 10 ng/ml and SCF 50 ng/ml in the presence (treatment group) and absence (control group) of GSNO for 30 min or 2 h. Platelet-sized particles were counted by flow cytometry; megakaryocyte structure was detected by scanning electron microscope. Aggregation of the thrombin-induced platelet particle was observed under inversion microscope. cGMP was assessed by commercial ELISA kit. The results showed that, compared with the control group, the number of platelet-sized particles significantly increased (P<0.05) in the treatment group, in which megakaryocytes presented significant pseudopod formation and extensive membrane blebbing. The platelet particle aggregation could be observed under microscope after thrombin induction. cGMP activity was significantly increased after treatment with GSNO (P<0.05). These results propose that GSNO can facilitate platelet production from megakaryocyte, and it may be partly through cGMP pathway.
Antigens, CD34 ; analysis ; Blood Platelets ; cytology ; Cell Differentiation ; drug effects ; Cyclic GMP ; blood ; Female ; Fetal Blood ; cytology ; Hematopoiesis ; drug effects ; Hematopoietic Stem Cells ; cytology ; Humans ; Megakaryocytes ; cytology ; Nitric Oxide ; physiology ; Platelet Aggregation ; drug effects ; Pregnancy ; S-Nitrosoglutathione ; pharmacology