Advanced oxidation protein products: oxidative stress markers and mediators of inflammation in uremia.
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Glycation End Products, Advanced↗
Biomedical subjects
V Witko-Sarsat
Publications and source records attributed to V Witko-Sarsat.
Advanced oxidation protein products: novel uraemic toxins and pro-inflammatory mediators in chronic renal failure?
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Humans↗
Disturbed myeloperoxidase-dependent activity of neutrophils in cystic fibrosis homozygotes and heterozygotes, and its correction by amiloride.
The present study addresses the question of a possible linkage between the cystic fibrosis (CF) genetic autosomal recessive disorder and disturbance in neutrophil function. Neutrophil-dominated chronic airway inflammation is present at an early age in children with CF, even in the absence of detectable infection. As evidenced by extracellular superoxide anion release (measured by lucigenin luminescence) or intracellular hydrogen peroxide production (measured by 2',7'-dichlorofluorescein (DCF) fluorescence), no significant difference in the nicotinamide-adenine dinucleotide phosphate (NADPH) oxidase activity of isolated neutrophils was observed in noninfected CF children (homozygotes), their mothers or fathers (CF heterozygotes), and controls. In contrast, both myeloperoxidase (MPO)-dependent oxygenation activity (measured by luminol luminescence) and chloramine release were increased significantly in both CF homozygotes and heterozygotes as compared with controls. In the presence of either amiloride (a sodium channel inhibitor and sodium/proton antiport blocker) or EIPA (5-ethyl-N-isopropyl-amiloride, a specific inhibitor of the antiport), or choline buffer, intracellular MPO activity was decreased significantly in controls and in the CF homozygotes and heterozygotes, thus bringing intracellular MPO-dependent activity in CF subjects back to the level of controls. Extracellular release of MPO, measured by an ELISA to provide an activity-independent assessment of the enzyme, was increased only in CF homozygotes, and was decreased by amiloride and choline buffer, but not by EIPA. We conclude that a modification of intracellular pH and/or ionic concentrations may be related to the altered MPO enzymatic activity observed in CF neutrophils.
Adolescent↗
Characterization of a recombinant proteinase 3, the autoantigen in Wegener's granulomatosis and its reactivity with anti-neutrophil cytoplasmic autoantibodies.
Using the baculovirus/insect cells system, we have expressed a recombinant proteinase 3 (PR3) -- the neutrophil-derived serine protease autoantigen in Wegener's granulomatosis -- as a glycosylated intracellular and membrane-associated protein. Oligosaccharides accounted for the difference in molecular weights between recombinant (34 kDa) and neutrophil-PR3 (29 kDa). Whereas rabbit-anti-PR3 IgG recognized both recombinant and neutrophil-derived PR3, autoantibodies from Wegener patient sera recognized only neutrophil-derived PR3. Although oligosaccharides were not involved in PR3 epitope recognition, autoantibodies did not recognize the amino acid primary structure of recombinant PR3. Improper disulfide bond formation and/or lack of post-translational events in insect cells, may affect the conformation and/or lack of post-translational events in insect cells, may affect the conformation of PR3, precluding its reactivity with sera from WG patients.
Amidohydrolases↗
Azurocidin, a natural antibiotic from human neutrophils: expression, antimicrobial activity, and secretion.
The azurophil granules of human PMN contain four antibiotic proteins, the serprocidins, which have extensive homology to one another and to serine proteases. Azurocidin, a member of this family, is a 29-kDa glycoprotein with broad spectrum antimicrobial activity and chemotactic activity toward monocytes. Insect cells transfected with a baculovirus vector carrying azurocidin cDNA produced a recombinant azurocidin protein. We purified the recombinant azurocidin protein from the culture medium of the infected cells and showed that it retained the antimicrobial activity of the native neutrophil-derived molecule. In addition, we present evidence that a 49-amino-acid region of the recombinant azurocidin protein is required for its secretion from insect cells.
Amino Acid Sequence↗
Glutathione antioxidant system as a marker of oxidative stress in chronic renal failure.
A profound imbalance between oxidants and antioxidants has been suggested in uremic patients on maintenance hemodialysis. However, the respective influence of uremia and dialysis procedure has not been evaluated. Circulating levels of copper-zinc superoxide dismutase (CuZn SOD), glutathione peroxidase (GSH-Px), and reductase (GSSG-Rd), total GSH and GSSG were determined in a large cohort of 233 uremic patients including 185 undialyzed patients with mild to severe chronic renal failure, and 48 patients treated by peritoneal dialysis or hemodialysis. Compared to controls, erythrocyte GSH-Px and GSSG-Rd activities were significantly increased at the mild stage of chronic uremia (p < .001), whereas erythrocyte CuZn SOD activity was unchanged, total level of GSH and plasma GSH-Px activity were significantly decreased, and GSSG level and GSSG-Rd activity were unchanged. Positive Spearman rank correlations were observed between creatinine clearance and plasma levels of GSH-Px (r = .65, p < .001), selenium (r = .47, p < .001), and GSH (r = .41, p < .001). Alterations in antioxidant systems gradually increased with the degree of renal failure, further rose in patients on peritoneal dialysis and culminated in hemodialysis patients in whom an almost complete abolishment of GSH-Px activity was observed. In conclusion, such disturbances in antioxidant systems that occur from the early stage of chronic uremia and are exacerbated by dialysis provide additional evidence for a resulting oxidative stress that could contribute to the development of accelerated atherosclerosis and other long-term complications in uremic patients.
Antioxidants↗
Advanced oxidation protein products as a novel marker of oxidative stress in uremia.
Evidence suggests an imbalance between antioxidant and oxidant-generating systems resulting in oxidative stress in uremic patients. As plasma proteins are critical targets for oxidants, we developed a novel spectrophotometric assay which allows to detect advanced oxidation protein products (AOPP) in uremic plasma. By size-exclusion chromatography AOPP are retrieved in two distinct peaks at 600 and below 80 kDa in uremic plasma, while no such peaks are found in control plasma. Further biochemical characterization revealed that AOPP are carried by oxidized plasma proteins, especially albumin and do not have oxidant properties. AOPP increased in a dose-dependent manner following in vitro exposure of plasma or purified human serum albumin (HSA) to hypochlorous acid. Advanced glycation end products of human serum albumin (AGE-HSA) also increased AOPP levels. In vivo, plasma level of AOPP was the highest in patients on hemodialysis, followed by those on peritoneal dialysis and by undialyzed patients with advanced chronic renal failure. AOPP levels correlated with plasma concentrations of dityrosine and AGE-pentosidine, as indices of oxidant-mediated protein damage, but not with thiobarbituric reactive substances as lipid peroxidation markers. A close correlation was also found between AOPP and neopterin levels, suggesting that AOPP could be part in the monocyte-mediated inflammatory disorders associated with uremia. In conclusion, we propose the measurement of AOPP as a reliable marker to estimate the degree of oxidant-mediated protein damage in uremic patients and to predict the potential efficacy of therapeutic strategies aimed at reducing such an oxidative stress.
Biomarkers↗
The advanced glycation endproduct pentosidine and monocyte activation in uremia.
RATIONALE: Advanced glycation endproducts (AGEs) contribute to the pathogenesis of vascular complications in diabetes, aging and end-stage renal disease (ESRD). Immune abnormalities in patients with chronic renal failure and those treated by dialysis contribute to high rates of morbidity and mortality. We therefore sought a relationship between a circulating marker of immune dysfunction and plasma levels of the AGE pentosidine. METHOD: We studied non-diabetic patients with mild to advanced renal failure (n = 60), and with ESRD treated by hemodialysis (HD) (n = 44) and peritoneal dialysis (PD) (n = 19). The plasma protein content of the well characterized AGE, pentosidine was measured using HPLC. In the same samples the monocyte activation product neopterin was measured by RIA. RESULTS: Plasma levels of pentosidine and neopterin increased in parallel with the progression of renal failure. Pentosidine and neopterin were highly correlated in all patients even after adjustment for Ccr. This correlation was also present in patients with ESRD. CONCLUSION: These data suggest that the AGE pentosidine is associated with monocyte activation in renal failure, an interaction which may contribute to accelerated rates of complication and death by as yet unknown mechanisms.
Arginine↗
Bimodal distribution of proteinase 3 (PR3) surface expression reflects a constitutive heterogeneity in the polymorphonuclear neutrophil pool.
Proteinase 3, which is known as an intracellular serine protease of neutrophils, was detected at the surface of a subpopulation of freshly isolated PMN. The proportion of PR3-positive and -negative PMN, observed by flow cytometry with anti-PR3 mAbs or ANCA autoantibodies, varies among individuals but is extremely stable for each individual over prolonged time periods. After PMN degranulation by FMLP with cyt. B, membrane PR3 expression increases but the proportion of low and high PR3-expressing cells remains stable. The existence of a subset of PMN which spontaneously expresses PR3 and varies among individuals, may be relevant to the pathogenesis of anti-PR3 ANCA autoantibody-related vasculitis.
Animals↗
Neutrophil-derived long-lived oxidants in cystic fibrosis sputum.
We evaluated long-lived oxidant potential in the sputum of patients with cystic fibrosis (CF) by quantitating the methionine-inhibitable, long-lived oxidant fraction of sputum, referred to as the chloramines. Taurine, the preferred amino acid substrate for chloramine formation, and myeloperoxidase (MPO), the chlorinated oxidant-generating enzyme, were also quantitated. As compared with the sputum of asthmatic subjects, the sputum of CF patients contained high concentrations of chloramines along with high levels of taurine and active MPO. A negative correlation between chloramine and taurine was found in the sputum of CF patients. No correlation was found between the density of Pseudomonas aeruginosa and the level of chloramines, taurine, or MPO. In contrast, respiratory parameters (%FEV or %FVC) and a nutritional index correlated positively with chloramine levels, whereas negative correlations were observed with taurine and MPO. In addition, the effect of antibiotic therapy, which significantly increased chloramine and decreased taurine levels, supported a beneficial effect of chloramines on overall clinical status. Our findings support a dual role of long-lived oxidants at the site of airway inflammation in CF, one component of which is their ability to mediate oxidative stress and the other a beneficial effect that may be partly explained by their inhibitory effect on antiprotease defense systems.
Adolescent↗
Pholasin: a new chemiluminescent probe for the detection of chloramines derived from human phagocytes.
We have previously reported that normal human polymorphonuclear neutrophils (PMN) release taurine-chloramine, a long-lived oxidant, in response to stimulation by phorbol myristate acetate (PMA) or opsonized zymosan in the presence of exogenous taurine. We now describe a new, simple, and highly sensitive method for the detection of chloramines, including taurine-chloramine, using the chemiluminescent probe Pholasin, the luciferin of the mollusc Pholas dactylus. Taurine-chloramine (N-chlorotaurine) detection was assessed with both a colorimetric method (based on the oxidation of potassium iodide) and with the Pholasin-dependent chemiluminescence (CL) method. The taurine-chloramine concentration in PMN supernatants determined using the potassium iodide (KI) method correlated closely with Pholasin-dependent CL. This CL was also assessed in nonoxidative conditions. No taurine-chloramine was detected in supernatants of lymphocytes and PMN from patients with an oxidative burst defect (chronic granulomatous disease, CGD) with the KI method, but Pholasin-dependent CL was consistently observed. The use of methionine, a specific chloramine scavenger in our incubation conditions, allowed us to define a methionine-inhibitable fraction of Pholasin-dependent CL (i.e., chloramine-induced CL).
Child↗
Dialysis-induced oxidative stress: biological aspects, clinical consequences, and therapy.
Oxidative stress, which results from a rupture in the natural balance between pro- and antioxidant systems, is considered as a major factor in dialysis-associated morbidity and mortality. Emerging pharmacologic and dialytic antioxidant therapeutic and dialysis strategies should enable us to reduce the harmful consequences of oxidative stress in dialysis patients. Moreover, since there is increasing evidence of oxidative stress long before the initiation of maintenance dialysis, antioxidant therapeutic strategies should probably be developed very early in the course of renal failure.
Antioxidants↗