Morpho-functional modifications of human neutrophils induced by aqueous cigarette smoke extract: comparison with chemiluminescence activity.

Cigarette smoking plays an important role as a cause of morbidity and mortality in humans, involving respiratory, cardiovascular, digestive and reproductive systems. Tobacco smoke contains a large number of molecules, some of which are proven carcinogens. Although not fully understood, polymorphonuclear leukocytes seem to play a crucial role in the mechanisms by which tobacco smoke compounds are implicated in smoke-related diseases. In this paper the effects of an aqueous cigarette smoke extract on the expression of adhesion molecules of polymorphonuclear leukocytes together with the changes in the cell morphology have been related to the chemiluminescence activity. The results obtained show that polymorphonuclear leukocytes treated with aqueous cigarette smoke extract are significantly impaired, as suggested by the changes of chemiluminescence activity, of membrane receptors (CD18, CD62), myeloperoxidase expression and of cell morphology. Altogether the present data indicate that treated polymorphonuclear leukocytes are ineffectively activated and therefore unable to phagocytize zymosan particles.

Salivary thiols and enzyme markers of cell damage in periodontal disease.

OBJECTIVES: Recent studies describe the potential use of biochemical markers in the evaluation of the severity of periodontitis; moreover, patients suffering from periodontitis frequently complain of halitosis (breath malodour), mainly depending on volatile compounds (e.g. hydrogen sulphide, methyl mercaptan, etc.) produced by anaerobic metabolism of oral bacteria and involving sulphur-containing amino acids. In this study, salivary sulphur compounds, such as cysteine, cysteinylglycine and glutathione and some markers of cellular damage (lactate dehydrogenase and aspartate amino transferase), were measured in periodontitis patients and correlated with the periodontal probing pocket's depth. DESIGN AND METHODS: Twenty-two periodontitis patients and forty control subjects were studied for the salivary activities of lactate dehydrogenase and aspartate aminotransferase and cysteine, cysteinylglycine and glutathione concentrations. The periodontitis patients were divided into two subgroups based on the severity of periodontal disease, expressed as median periodontal probing pocket depth (> or <5 mm). Enzyme activities were measured by using an automated clinical analyzer; cysteine, cysteinylglycine and glutathione concentrations were measured by HPLC equipped with fluorescence detector. RESULTS: A statistically significant increase of the salivary parameters level (cysteine, cysteinylglycine, glutathione, aspartate aminotransferase and lactate dehydrogenase) was found in the patient subgroup with periodontal probing pocket depth >5 mm, the salivary cysteine concentrations showing the most significant correlation. CONCLUSIONS: Salivary cysteine, a direct precursor of hydrogen sulphide, could be considered reliable markers for the oral tissue damage severity in periodontitis patients.

Alterations of energy metabolism and glutathione levels of HL-60 cells induced by methacrylates present in composite resins.

OBJECTIVES: Methacrylic compounds such as 2-hydroxyethyl methacrylate (HEMA), triethylene glycol dimethacrylate (TEGDMA) and bisphenol A glycerolate (1 glycerol/phenol) dimethacrylate (Bis-GMA) are largely present in auto- or photopolymerizable composite resins. Since the polymerization reaction is never complete, these molecules are released into the oral cavity tissues and biological fluids where they could cause local adverse effects. The aim of this work was to verify the hypothesis that the biological effects of HEMA, TEGDMA and Bis-GMA - at a non-cytotoxic concentration - depend on the interaction with mitochondria and exert consequent alterations of energy metabolism, GSH levels and the related pathways in human promyelocytic cell line (HL-60). METHODS: The biological effects of methacrylic monomers were determined by analyzing the following parameters: GSH concentration, glucose-6-phosphate dehydrogenase (G6PDH) and glutathione reductase (GR) activity, oxygen and glucose consumption and lactate production along with cell differentiation and proliferation. RESULTS: All monomers induced both cellular differentiation and decrease in oxygen consumption. Cells treated with TEGDMA and Bis-GMA showed a significant enhancement of glucose consumption and lactate production. TEGDMA and HEMA induced GSH depletion stimulating G6PDH and GR activity. CONCLUSIONS: All the monomers under study affect the metabolism of HL-60 cells and show differentiating activity. Since alterations in cellular metabolism occurred at compound concentrations well below cytotoxic levels, the changes in energy metabolism and glutathione redox balance could be considered as potential mechanisms for inducing clinical and sub-clinical adverse effects and thus providing useful parameters when testing biocompatibility of dental materials.

Decreased chemiluminescence in leukocyte adhesion deficiency presenting with recurrent sepsis, amoebiasis and Candida albicans urinary tract infection.

Leukocyte adhesion deficiency (LAD) is a rare disorder of cellular immunity, generally due to various mutations producing reduced or altered expression of membrane integrins. The authors report a case of LAD due to integrins expression imbalance. LAD was suspected after recurrent sepsis, fungal infection and amoebiasis with persistent leukocytosis. Neutrophils were studied with chemiluminescence showing decreased functional activity: up to now, this seems the first chemiluminescence study of neutrophil function and the first report of amoebiasis at the onset in LAD.

Lactic dehydrogenase and glycolytic activity of human platelets in uncomplicated pregnancy.

Lactic dehydrogenase (LDH) of isolated platelets was determined in a group of 103 normally pregnant and 19 unpregnant women. LDH activity is significantly increased in the last trimester of pregnancy while the isoenzyme pattern does not vary in relation to LDH increase. Moreover, glucose consumption and lactic acid production are linearly correlated to platelet LDH activity.

Antimicrobial and antioxidant activities of Feijoa sellowiana fruit.

The present study was designed to evaluate the antibacterial and antioxidant activities of an aqueous extract from the tropical Feijoa sellowiana Berg. fruit which is widely used for human food. The extract was tested against gram-positive and gram-negative bacteria by a broth dilution test and on human whole blood leukocytes, as well as isolated neutrophils using a chemiluminescence (CL) assay. The extract inhibited bacterial growth; Pseudomonas aeruginosa, Enterobacter aerogenes and Enterobacter cloacae were the most sensitive. The fruit extract significantly decreased CL emission from human whole blood phagocytes and isolated polymorphonuclear leukocytes whether they were activated or not by soluble or phagocytic stimuli. F. sellowiana showed both antibacterial and antioxidant properties and therefore its extract might be used as a new multifaceted drug.

Is homocysteine a pro-oxidant?

High plasma homocysteine concentrations have been found to be associated with atherosclerosis and thrombosis of arteries and deep veins. The oxidative damage mediated by hydrogen peroxide production during the metal-catalyzed oxidation of homocysteine is to date considered to be one of the major pathophysiological mechanisms for this association. In this work, a very sensitive and accurate method was employed to measure the effective production of H2O2 during homocysteine oxidation. Furthermore, the interaction of homocysteine with powerful oxidizing species (hypochlorite, peroxynitrite, ferrylmyoglobin) was evaluated in order to ascertain the putative pro-oxidant role of homocysteine. Our findings indicate that homocysteine does not produce H2O2 in a significant amount (1/4000 mole/mole ratio of H2O2 to homocysteine). Moreover, homocysteine strongly inhibits the oxidation of luminol and dihydrorhodamine by hypochlorite or peroxynitrite and rapidly reduces back ferrylmyoglobin, the oxidizing species, to metmyoglobin. All these results should, in our opinion, lead to a rethinking of the commonly held view that homocysteine oxidation is one of the main causative mechanisms of cardiovascular damage.

A fast chemiluminescent method for H(2)O(2) measurement in exhaled breath condensate.

BACKGROUND: Breath condensate can give useful information on volatile compounds produced at alveolar level. Actual concentration of H(2)O(2) in breath condensate is dependent on its production at alveolar level and on the efficacy of the detoxifying systems, catalase, glutathione peroxidase, etc. METHODS: In the present paper, a simple chemiluminescent method for the determination of the H(2)O(2) collected in exhaled breath is shown and data of both smokers and nonsmokers volunteers are presented. RESULTS: The chemiluminescent response is linear up to 100 micromol/l H(2)O(2). The analytical sensitivity is about 0.01 micromol/l. Most of the nonsmokers have a H(2)O(2) content lower than 0.05 micromol/l, while smokers have a content ranging from 0.1 to 0.6 micromol/l.

The priming effect of gemfibrozil on reactive oxygen metabolism of phagocytic leucocytes. An intriguing side effect.

Gemfibrozil is an antihyperlipidaemia agent used in therapy to reduce the occurrence of coronary heart disease. Considering the biochemical and pharmacological peculiarities of this class of drugs, we investigated the influence of a single oral therapeutic dose of gemfibrozil on the reactive oxygen metabolism of phagocytic leucocytes. Analysis was carried out adopting a chemiluminescence assay. Results clearly indicate that gemfibrozil, acting as a primer, significantly enhances the induced reactive-oxygen-species (ROS) production by overall blood phagocytes (increment of Stimulation Index (SI) = +52% with respect to time 0 values; P < 0.01), by polymorphonuclear leucocytes (increment of SI = +28% with respect to control values; P < 0.01) and by monocytes (increment of SI = +83% with respect control values; P < 0.001), when these cells are stimulated by phorbol myristate acetate. This iatrogenic derangement of ROS metabolism could explain, at least in part, the occurrence of some side effects that occur in treatment with fibrates.

Purine metabolites and malondialdehyde in platelets of diabetic patients.

The concentration of some of the purine nucleotides and their metabolites together with that of malondialdehyde (MDA) have been measured in resting and stimulated platelets of type 1 and type 2 diabetic patients. While control platelets show a net decrease of guanosine triphosphate (GTP) (3.1 vs. 2.3 nmol per 10(9) platelets) and guanosine diphosphate (GDP) (3.0 vs. 2.0 nmol per 10(9) platelets) and a significant increase of adenosine (0.04 vs. 0.55 nmol per 10(9) platelets) with platelet stimulation, platelets of type 1 and type 2 diabetic patients have a lesser change of these metabolites (GTP, 2.6 vs. 2.4; GDP, 2.3 vs. 2.4; adenosine, 0.04 vs. 0.30 (P < 0.05 vs. control) nmol per 10(9) platelets in type 1 diabetics; GTP, 2.4 vs. 2.7; GDP, 2.4 vs. 2.1; adenosine, 0.08 vs. 0.32 (P < 0.05 vs. control) nmol per 10(9) platelets in type 2 diabetics). These results indicate that the change (stimulated minus resting) of GTP, GDP and adenosine in diabetic platelets is significantly different from that of controls (P < 0.001). Moreover, the amount of MDA produced during platelet activation seems to be lower than controls only in type 2 diabetes (1.81 vs. 2.86 nmol per 10(9) platelets, P < 0.05). These results seem to indicate that a difference in the pattern of platelet nucleotides could be an important feature even in well-controlled diabetes, while MDA is probably modified only in association with the late vascular complications of diabetes.

Chemiluminescence of bronchoalveolar macrophages: effect of adherence to plastic cuvette.

The chemiluminescence of alveolar macrophages was determined in presence of luminol or lucigenin, either before or after the macrophage isolation by adherence to plastic. In presence of luminol, the purified and non-purified macrophages behave almost in the same way, and their chemiluminescence activity is stimulated both by zymosan and phorbol ester. On the other hand, the lucigenin chemiluminescence of non-purified macrophages is considerably higher than that obtained in presence of luminol, but is almost completely insensitive to the zymosan or phorbol ester stimulation. Moreover, the adherence purification considerably inhibits the lucigenin chemiluminescence. These results can be indicative of functionally different macrophage states.

Effect of smoking one cigarette on antioxidant metabolites in the saliva of healthy smokers.

Concentrations of glutathione, uric acid and total antioxidant activity, expressed as Trolox (a water-soluble vitamin E analogue) equivalent, were measured in the saliva of healthy non-smokers and smokers before and just after smoking a single cigarette. There was no statistically significant difference between smokers and non-smokers in uric acid concentrations and total radical-trapping antioxidant capacity, but glutathione concentrations were significantly (p < 0.05) higher in smokers. Smoking of a single cigarette induced a significant reduction in glutathione concentration (p < 0.05). Salivary antioxidant power may affect individual sensitivity toward tobacco stress.

In vitro effect of different ubiquinones on the scavenging of biologically generated O2-.

The ability of different homologues of Coenzyme Q to quench O2- was tested in vitro with three experimental systems known to generate O2-. Two of them were biological generators, namely the xanthine-xanthine oxidase system and the cyanide-insensitive NADPH oxidase of polymorphonuclear leucocytes. The third was a chemical generator of O2-, the NADH-phenazine methosulphate-nitroblue tetrazolium mixture. Short-side-chain ubiquinones were found to be the most potent scavengers of O2-, being effective at concentrations as low as 10(-7) M. This finding might be ascribed to the relatively greater water-solubility of the lower homologues of CoQ. We postulate that CoQ10 may well exert such an O2- -scavenging mechanisms in vivo where it is inserted in its natural phospholipid environment.

Reactive oxygen species production in circulating polymorphonuclear leukocytes in psoriasis.

Metabolic burst of polymorphonuclear leukocytes was evaluated by means of a luminol-enhanced chemiluminescence assay. Patients affected with active psoriasis showed a higher response than patients with chronic disease and controls when the neutrophils were stimulated with opsonized zymosan or phorbol myristate acetate.