OBJECTIVETo investigate the influence of neonatal maturity on active oxygen metabolism in neutrophils and possible causes of a high susceptibility to bacterial infection in preterm infants.

METHODSThirty-five preterm infants born at a gestation age of 26-32 weeks (< or =32 weeks group, n=15) and at 33-36 weeks (> 32 weeks group, n=20) and 23 full-term infants (control group) were enrolled in this study. The samples of whole cord blood from the two preterm groups and the control group were stimulated in vitro with live bacteria,Staphylococcus aureus ( S. aureus) and Escherichia coli (E. coli) and stained with hydroethedine, an indicator of superoxide. The percentage of neutrophils which produced superoxide and the mean fluorescence intensity for superoxide production were measured by flow cytometry. The incidence of bacterial infection during hospital stay was compared between the two preterm groups.

RESULTSUnder S. aureus or E. coli stimulation, the percentage of neutrophils which produced superoxide in the < or =32 weeks group was significantly lower than that of the > 32 weeks group and the control group (P < 0.01). The percentage of neutrophils which produced superoxide was closely related to gestational age in preterm infants ( y=2.66 x, P < 0.01).There were no significant differences in the blood level of superoxide production in neutrophils among the three groups. The incidence of bacterial infection during hospital stay in the < or =32 weeks group (40%) was significantly higher than that the > 32 weeks group (10%) (P < 0.05).

CONCLUSIONSThe capability of active oxygen metabolism in neutrophils was significantly related to the gestational age in preterm infants. The decreased capability of active oxygen metabolism might be contributed to a higher susceptibility to bacterial infection in preterm infants.

Disease Susceptibility ; Gestational Age ; Humans ; Infant, Newborn ; Infant, Premature ; immunology ; Neutrophils ; metabolism ; Superoxides ; metabolism

OBJECTIVE: To investigate the eliminating ability of catechin to eliminate O2-* and *OH. METHODS: The ability of catechin to clear away O2-* and *OH was respectively measured by faintness chemiluminescence and spin trapping assay. RESULTS: IC50 that catechin eliminated O2-* and *OH was 6.16, 0.59 g/mL respectively, and the eliminating ability of catechin was much stronger than that of the extract from liquorice, rosemary, grape pip, giant knotweed and ginkgo leaf. CONCLUSION Compared with several important natural plants of antioxidants, the eliminating ability of cathechin is the best.
Antioxidants ; pharmacology ; Catechin ; pharmacology ; Free Radical Scavengers ; pharmacology ; Hydroxyl Radical ; metabolism ; Luminescent Measurements ; Superoxides ; metabolism

BACKGROUNDSkeletal muscle has recently been recognized as an endocrine organ that can express, synthesize and secrete a variety of bioactive molecules which exert significant regulatory effects. Hydrogen sulfide (H2S) is endogenously produced in mammalian tissues and participates in a number of physiological and pathophysiological processes. We aimed to verify whether H2S could be endogenously generated and released by rat skeletal muscle, and determine the biological effects of H2S in rat skeletal muscle.

METHODSThe study was divided into two parts: detection of endogenous H2S generation and release in rat skeletal muscle and determination of antioxidative activity of skeletal muscle-derived H2S. H2S content and production in tissues were detected by sensitive sulfur electrode method. The expressions of H2S producing enzymes cystathionine β-synthase, cystathionine γ-lyase and mercaptopyruvate sulfurtransferase were detected by real-time PCR and western blotting and their tissue distributions were observed by immunohistochemical and immunofluorescent analysis. Rat skeletal muscular ischemia-reperfusion (I-R) injury model was created and evaluated by histological analysis under microscope. The malondialdehyde (MDA) contents, hydrogen peroxide levels, superoxide anion and superoxide dismutase (SOD) activities were detected using spectrophotometer.

RESULTSH2S could be endogenously generated and released by skeletal muscle of Sprague-Dawley rats (H2S content: (2.06 ± 0.43) nmol/mg; H2S production: (0.17 ± 0.06) nmol×min(-1)×mg(-1)). Gene and protein expressions of the three H2S producing enzymes were detected in skeletal muscle, as well as the liver and kidney. Endogenous H2S content and production were decreased in skeletal muscles of rats with I-R skeletal muscle injury (P < 0.05). Furthermore, H2S significantly protected rat skeletal muscle against I-R injury and resulted in decreased MDA content, reduced hydrogen peroxide and superoxide anion levels, but increased SOD activity and protein expression in skeletal muscles (all P < 0.01).

CONCLUSIONH2S generation pathway exists in rat skeletal muscle and it acts as an antioxidant in skeletal muscle.

Animals ; Blotting, Western ; Hydrogen Peroxide ; metabolism ; Hydrogen Sulfide ; metabolism ; Immunohistochemistry ; Male ; Malondialdehyde ; metabolism ; Muscle, Skeletal ; metabolism ; Oxidative Stress ; physiology ; Rats ; Rats, Sprague-Dawley ; Superoxide Dismutase ; metabolism ; Superoxides ; metabolism

Neutrophils are the predominant inflammatory cells found in vaginal discharges of patients infected with Trichomonas vaginalis. In this study, we examined superoxide anion (O2(.-)) production by neutrophils activated by T. vaginalis. Human neutrophils produced superoxide anions when stimulated with either a lysate of T. vaginalis, its membrane component (MC), or excretory-secretory product (ESP). To assess the role of trichomonad protease in production of superoxide anions by neutrophils, T. vaginalis lysate, ESP, and MC were each pretreated with a protease inhibitor cocktail before incubation with neutrophils. Superoxide anion production was significantly decreased by this treatment. Trichomonad growth was inhibited by preincubation with supernatants of neutrophils incubated for 3 hr with T. vaginalis lysate. Furthermore, myeloperoxidase (MPO) production by neutrophils was stimulated by live trichomonads. These results indicate that the production of superoxide anions and MPO by neutrophils stimulated with T. vaginalis may be a part of defense mechanisms of neutrophils in trichomoniasis.
Anions/*metabolism ; Female ; Humans ; Neutrophils/enzymology/*metabolism/parasitology ; Peroxidase/metabolism ; Superoxides/*metabolism ; Trichomonas Infections/enzymology/*metabolism/parasitology ; Trichomonas vaginalis/*isolation & purification/physiology

Phagocytosis by inflammatory cells is an essential step and a part of innate immunity for protection against foreign pathogens, microorganism or dead cells. Phagocytosis, endocytotic events sequel to binding particle ligands to the specific receptors on phagocyte cell surface such as Fcgamma recptor (FcgammaR), complement receptor (CR), beta-glucan receptor, and phosphatidylserine (PS) receptor, require actin assembly, pseudopod extension and phagosome closure. Rho GTPases (RhoA, Cdc42, and Rac1) are critically involved in these processes. Abrupt superoxide formation, called as oxidative burst, occurs through NADPH oxidase complex in leukocytes following phagocytosis. NADPH oxidase complex is composed of membrane proteins, p22(PHOX)and gp91(PHOX), and cytosolic proteins, p40(PHOX), p47(PHOX)and p67(PHOX). The cytosolic subunits and Rac-GTP are translocated to the membrane, forming complete NADPH oxidase complex with membrane part subunits. Binding of imunoglobulin G (IgG)- and complement-opsonized particles to FcgammaR and CR of leukocytes induces apoptosis of the cells, which may be due to oxidative burst and accompanying cytochrome c release and casapase-3 activation.
Animals ; Apoptosis/*physiology ; Human ; Macrophages/*cytology/*metabolism ; NADPH Oxidase/metabolism ; Phagocytosis/*physiology ; Reactive Oxygen Species/metabolism ; Superoxides/*metabolism ; Support, Non-U.S. Gov't

OBJECTIVETo explore the effect of 5-HT2A-receptors coupled with superoxide anion (O2-) on respiratory regulation signal transductionin passageway in the medial area of nucleus retrofacialis (mNRF).

METHODSmNRF island was prepared from medullary slices of neonatal SD rats according to Johnson's method and transferred separately into 24-well culture plates with reagents according to protocol, followed by incubation for 60 min at 37 degrees C in a humidified incubator with 5% CO2. Absorbance of 100 microl supernatant was measured by spectrophotometry at 550 nm and the effect of 5-HT and 2,5-dimethoxy-4-iodoamphetamine hydrochloride (DOI, agonist of 5-HT2A-receptors) on O2- generation in the mNRF was observed, along with the inhibition of this effect by ketanserin (antagonist of 5-HT2A-receptors) and alpha-lipoic acid (alpha-LA, a antioxidant).

RESULTS5-HT concentration-response curve demonstrated that absorbance peak occurred at 1 micromol/L without further increment with higher concentration. DOI concentration-response curve showed the absorbance peak at 20 micromol/L without further increment. 5-HT and DOI significantly increased the absorbance with comparable effects. Ketanserin and alpha-LA significantly decreased the absorbance generated by 5-HT and DOI.

CONCLUSIONActivation of 5-HT2A receptors results in obvious O2- production in mNRF, suggesting that 5-HT2A receptors regulate respiratory function in association with O2-.

Animals ; Animals, Newborn ; Medulla Oblongata ; metabolism ; physiology ; Rats ; Rats, Sprague-Dawley ; Receptor, Serotonin, 5-HT2A ; metabolism ; physiology ; Respiratory Center ; metabolism ; physiology ; Superoxides ; metabolism

Production of free radicals of superoxide anion in tissues by cadmium, activities of superoxide dismutase and catalase to protect tissue damages caused by the free radicals and ATPase that plays an important role in energy metabolism at cellular level were investigated. Experiments in viro were conducted with liver, kidney and testicle tissue homogenates of rats adding 0.05~0.50 mM cadmium chloride, and in vivo experiments administering single dose of 5mg of cadmium/kg of body weight in 0.1% cadmium chloride solution intraperitoneally 48 hours prior to evisceration. Production of superoxide radicals in liver and testicle increased with addition of cadmium in vitro, but not in kidney. In vivo experiments, however superoxide radicals slightly increased in liver and kidney but not in testicle. Superoxide dismutase (Cu, Zn-SOD and Mn-SOD), catalase and ATPase (total, (Mg++)- & (Na+)- (K+)-) activity decreased in the presence of cadmium in dose dependent manner. Reduction of these enzyme activities varied not only with dosage of cadmium but also with type of tissue and between in vitro and in vivo experiment.
Adenosine Triphosphatases* ; Animals ; Body Weight ; Cadmium Chloride ; Cadmium* ; Catalase* ; Energy Metabolism ; Free Radicals ; Kidney* ; Liver* ; Rats* ; Superoxide Dismutase* ; Superoxides* ; Testis*

BACKGROUND: The product of oxygen-free radicals inf1ict oxidative injuries on healthy cells. Antioxidants such as superoxide dismutase(SOD), glutathione peroxidase, and reduced glutathione(GSH) are present in almost all cells and play important roles in metabolism, transport, and cellular protection. We measured blood GSH levels in healthy controls and patients with non insulin dependent diabetes mellitus(NIDDM) for evaluation of the clinical usefulness of GSH. METHODS: Erythrocyte GSH levels were measured in fifty healthy controls and thirty NIDDM patients with diabetic retinopathies by Beutler's method. We also tested within-run precision, between-run precision, linearity and recovery rate to evaluate this method measuring erythrocyte GSH levels. RESULTS: The GSH levels (mean +/-SD) of NIDDM patients (5.03+/-0.67mumo1/Hb) were significantly lower than those of healthy control group (6.46+/-0.85mumo1/Hb)(P<0.001). The results of within-run precision and between-run precision when stored at 4degrees Cwere excellent (coefficient of variation were 2.79% and 2.42%, respectively), however, when stored at the room temperature the GSH levels were sharply declined. The linearity and recovery rate were acceptable. CONCLUSIONS: The prescision, linearity, and recovery rate of GSH measurement were excellent. The GSH levels in NIDDM patient group were reduced, and this probably contributes to the defective defense mechanism against increased oxidative stress. Additional measurement of other antioxidants such as superoxide dismutase and glutathione Peroxidase may be required to clarify the pathologic significance of glutathione metabolism in various diseases.
Antioxidants ; Diabetes Mellitus, Type 2 ; Diabetic Retinopathy ; Erythrocytes* ; Glutathione Peroxidase ; Glutathione* ; Humans ; Insulin ; Metabolism ; Oxidative Stress ; Superoxide Dismutase ; Superoxides

BACKGROUNDThe immunomodulatory effects of glucocorticoids (GCs) have been described as bimodal. High concentration of GCs exerts immunosuppressive effects and low levels of GCs are immunopermissive. While the immunosuppressive mechanisms of GCs have been investigated intensely, the immunopermissive effects of GCs remain unclear. A lot of studies showed GCs could exert rapid non-genomic actions. We herein studied the rapid immunopromoting effects of GCs.

METHODSWe observed the rapid (within 30 minutes) effects of corticosterone on respiratory burst of mouse peritoneal macrophages and studied their mechanisms. The superoxide anions were measured by cytochrome C reduction assay. Protein kinase C phosphorylation was measured by Western blotting and membrane fluidity was evaluated by fluorescence polarization measurement.

RESULTSThe 10(-8) mol/L and 10(-7) mol/L corticosterone rapidly increased the superoxide anions production by macrophages, which were insensitive to GC-receptor antagonist, mifepristone, and protein-synthesis inhibitor, cycloheximide. Corticosterone coupled to bovine serum albumin was able to mimic the effects of corticosterone. The effects were independent of protein kinase C pathway and the change in membrane fluidity.

CONCLUSIONSThe results indicate that corticosterone rapidly promote the superoxide anions production by mouse peritoneal macrophages may through non-genomic mechanisms. This study may contribute to understanding the effects of GCs under stress condition and the physiological significance of nongenomic effects of GCs.

Animals ; Corticosterone ; pharmacology ; Macrophages, Peritoneal ; drug effects ; physiology ; Male ; Mice ; Mice, Inbred BALB C ; Respiratory Burst ; drug effects ; Superoxides ; metabolism

Eosinophil degranulation is considered to be a key effector function for the killing of helminthic worms and tissue inflammation at worm-infected lesion sites. However, relatively little data are available with regard to eosinophil response after stimulation with worm-secreted products which contain a large quantity of cysteine proteases. In this study, we attempted to determine whether the degranulation of human eosinophils could be induced by the direct stimulation of the excretory-secretory products (ESP) of Paragonimus westermani, which causes pulmonary paragonimiasis in human beings. Incubation of eosinophils for 3 hr with Paragonimus-secreted products resulted in marked degranulation, as evidenced by the release of eosinophil-derived neurotoxin (EDN) in the culture supernatants. Moreover, superoxide anion was produced by eosinophils after stimulation of the ESP. The ESP-induced EDN release was found to be significantly inhibited when the ESP was pretreated with protease inhibitor cocktail or the cysteine protease inhibitor, E-64. These findings suggest that human eosinophils become degranulated in response to P. westermani-secreted proteases, which may contribute to in vivo tissue inflammation around the worms.
Animals ; *Cell Degranulation ; Cysteine Endopeptidases/metabolism/*physiology ; Eosinophil-Derived Neurotoxin/metabolism ; Eosinophils/*physiology ; Humans ; Paragonimus westermani/*enzymology ; Research Support, Non-U.S. Gov't ; Superoxides/metabolism ; Time Factors