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Resume Introduction L’erythropoietine possede, a fortes doses, des proprietes antioxydantes et antiapoptotiques. Quel est l’impact de la prise d’Epo aux doses habituellement utilisees sur le stress oxydant a l’exercice ? Patients et methodes Trente-six rats Wistar ont ete repartis en quatre groupes : C (controles), CE (epuises), T (traites), TE (traites et epuises). Les groupes T et TE sont traites par Epo recombinante humaine (rhuEpo [EPREX®]) a la dose de 50 UI/kg trois fois par semaine pendant trois semaines. Les rats des groupes C et CE sont traites par serum physiologique. Apres une periode d’habituation sur tapis roulant, les rats des groupes CE et TE subissent une epreuve d’effort a epuisement a 25 m/min avec une pente de 15 %. Nous avons mesure l’activite NADPH oxydase musculaire, le taux des enzymes antioxydantes intraerythrocytaires (glutathion peroxydase [GPx] et superoxyde dismutase [SOD]) et les marqueurs de peroxydation lipidique plasmatiques (malondialdehyde [MDA] et F2 isoprostanes [F2IsoP]). Synthese des faits Le traitement a l’EPO augmente l’hematocrite de 11,9 % (46,2 vs 55,1) sans augmenter le temps de course jusqu’a epuisement.. L’exercice augmente l’activite NADPH oxydase musculaire (p = 0,0245), augmente les taux d’isoprostanes plasmatique et l’activite GPx intraerythrocytaire (p = 0,0049) alors que le MDA plasmatique et l’activite SOD intraerythrocytaire ne sont pas modifies. L’effet de l’exercice n’est pas modifie par la rhuEPO. L’activite NADPH oxydase musculaire tend a diminuer chez les rats traites. Conclusion Ni l’EPO, ni l’augmentation des capacites de transport de l’oxygene qui en resulte n’ont d’effet sur les marqueurs sanguins de stress oxydant apres un exercice a epuisement chez le rat. La quantite d’enzymes antioxydantes circulantes est toutefois augmentee.

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7-Dehydrocholesterol (7-DHC), a normal cholesterol precursor, accumulates in plasma and tissues of patients affected by the Smith-Lemli-Opitz (SLO) syndrome, a recessive autosomic disease characterized by mental retardation and physical malformations. In these patients, we suspected that 7-DHC interfered with α-tocopherol (α-T) determination because this steroid could be coeluted with α-tocopherol acetate (α-TA), an internal standard commonly used for α-T determination, and because the conjugated dienic structure of 7-DHC strongly absorbs at 292 nm. The aim of this work was (i) to characterize the interfering material and (ii) to evaluate the possibility of a rapid screening test for SLO syndrome identification from 7-DHC determination by reverse-phase chromatography high-performance liquid chromatography (HPLC). Human plasmas (control and SLO syndrome) were extracted with hexane after ethanol protein precipitation and addition of ascorbic acid as protective agent plus α-TA or δ-tocopherol (δ-T) as internal standards. The dry hexane extract was used for HPLC, and trimethylsilyl ethers derivatives of the extracts were submitted to gas chromatography (GC) and gas chromatography-mass spectrometry (GC-MS). The material interfering in the α-T determination was unambiguously identified to 7-DHC by retention time (RT) relative to standard, RT relative to δ-T, and analysis of the eluted material by GC and by GC-MS. In α-T determination, the interference can be eliminated by using δ-T as internal standard instead of α-TA. Rapid detection and evaluation of 7-DHC in plasma appear to be possible by HPLC under the conditions described; comparison with the GC reference method suggests that the rapid HPLC determination of 7-DHC in plasma can be used within the 1–40 mg/dL range, values commonly found in SLO syndrome.

Plasmapheresis and low-density lipoprotein (LDL)-apheresis are recognized procedures for the treatment of hyperlipidemia resistant to diet and lipid-lowering drugs and provide information on cholesterol synthesis in hypercholesterolemic patients. However, cholesterol synthesis after acute cholesterol removal from plasma has never been investigated in normocholesterolemic patients. In this study, cholesterol synthesis was evaluated in three normocholesterolemic patients by determination of plasma lathosterol, lathosterol-to-cholesterol ratio, and plasma mevalonic acid. In a short-term kinetic study, samples were collected before and after plasmapheresis and every 6 hours during 24 hours. In the second part of the study, cholesterol synthesis was evaluated daily for 3 days. In normocholesterolemic patients, cholesterol returns to basal levels in 3 days. However, cholesterol removal did not result in a significant increase in lathosterol-to-cholesterol ratio or in plasma mevalonic acid, despite a slight increase in lathosterol. In contrast, when repeated plasma exchanges induced a dramatic hypocholsterolemia (

Introduction – objectifs Meme si l’atteinte tubulaire liee au tenofovir disoproxil fumarate (TDF) est bien connue, son evolution est peu documentee. Une etude transversale realisee sur 1 158 adultes infectes par le VIH et ayant un debit de filtration glomerulaire estime (DFGe) superieur a 60 ml/min/1,73 m2 a permis de depister une proteinurie de type tubulaire (PT) chez 107 de ces patients (pts) et de preciser les facteurs associes, dont le TDF (AIDS 2013 ; 27 : 1 295–302). Les objectifs sont de decrire l’evolution de cette PT et d’evaluer l’impact de l’arret eventuel du TDF. Materiels et methodes Etude longitudinale prospective observationnelle de 107 pts VIH+ presentant une PT (ratio proteinurie/creatininurie > 200 mg/g et ratio albuminurie/proteinurie Resultats Pour les 81 pts analyses (hommes : 80,2 % ; âge median 51 ans ; sous TDF a l’inclusion : 59 pts), la duree mediane de suivi est de 32 mois [IQR : 29,2–33,3], avec une mediane de 9 visites [IQR : 8–11]. Apres deux ans de suivi, 15/75 pts (20 %) ont une persistance de la PT initiale. Les facteurs associes a cette persistance sont la poursuite du TDF [OR 9,0 ; IC95 : 1,9–41,4 ; p = 0,01], et une valeur de proteinurie a l’inclusion plus elevee (> mediane 290 mg/g) [OR 6,0 ; IC95 : 1,3–28,0 ; p = 0,02]. Les 27 pts qui ont arrete le TDF (27/59) avaient a l’inclusion un DFGe plus bas et une proteinurie plus elevee, que ceux qui l’ont poursuivi. L’arret du TDF est associe a une diminution significative du ratio proteinurie/creatininurie, mais n’a pas d’impact sur le DFGe. Conclusion La persistance, rare, d’une proteinurie de type tubulaire concerne principalement les patients poursuivant le TDF et/ou ayant une valeur initiale de proteinurie elevee.

A new automated immunoassay low-mid volume (or = 250 immunoassays/day) chemiluminescent analyzer, Abbott Architect i1000sR, was evaluated by seven laboratories around the world (4 in Europe, one each in Canada, Japan, and the U.S.A.) to demonstrate equivalent performance for key operating characteristics (e.g., precision, turn around time, limit of detection, functional sensitivity, and linearity).The laboratories followed standard protocols to assess precision, limit of detection (LoD), functional sensitivity, assay linearity, method comparison, and sample carryover. Turn around time for three stat assays (beta-hCG, BNP, and CK-MB) and the time required to complete workloads of 50 and 100 tests with a mixture of 75% routine tests and 25% stat tests was also evaluated.Total precision was typically5% CV for nine immunoassays. Analytical performance met design goals and demonstrated equivalency to package insert data for assays on market and in use for an existing high volume immunoassay system. Stat turn around times were consistent with the fixed analytical time of 15.6 minutes and met the expectations of the laboratories. Measured test throughput ranged from 47 - 54 tests per hour and demonstrated that the analyzer was fit for the intended purpose of supporting a laboratory that performsor = 250 immunoassays per day.A multisite, international analyzer familiarization study is a practical means of confirming that a new instrument meets both a manufacturer's design specifications and users' real world expectations and provides a pragmatic test for the system. The experience of investigators at seven sites around the world indicates that a new fully automated chemiluminescent system is suitable for use.

Oxidative stress is commonly observed in chronic renal failure patients resulting from an unbalance between overproduction of reactive oxygen species and impairement of defense mechanisms. Proteins appear as potential targets of uremia-induced oxidative stress and may undergo qualitative modifications. Proteins could be directly modified by reactive oxygen species which leads to amino acid oxydation and cross-linking. Proteins could be indirectly modified by reactive carbonyl compounds produced by glycoxidation and lipo-peroxidation. The resulting post-traductional modifications are known as carbonyl stress. In addition, thiols could be oxidized or could react with homocystein leading to homocysteinylation. Finally, tyrosin could be oxidized by myeloperoxidase leading to advanced oxidative protein products (AOPP). Oxidatively modified proteins are increased in chronic renal failure patients and may contribute to exacerbate the oxidative stress/inflammation syndrome. They have been involved in long term complications of uremia such as amyloidosis and accelerated atherosclerosis.

SummaryIn rheumatoid arthritis (RA) there are currently no good indicators to predict a clinical response to rituximab. The purpose of this study was to monitor and determine the role of peripheral blood cytokine profiling in differentiating between a good versus poor response to rituximab in RA. Blood samples were collected at baseline and at 3 months from 46 RA patients who were treated with rituximab. Responders are defined by the presence of three of four American College of Rheumatology criteria: ≥ 20% decrease in C-reactive protein, visual analogical score of disease activity, erythrocyte sedimentation rate and improvement of the disease activity score (28) (four values) by ≥ 1·2 obtained at 3 months. Twelve cytokines were measured from serum collected on days 0 and 90 by proteomic array, including interleukin-6 (IL-6), tumour necrosis factor-α, IL-1a, IL-1b, IL-2, IL-8, interferon-γ, IL-4, IL-10, monocyte chemoattractant protein-1, epidermal growth factor and vascular growth factor. We showed that C-reactive protein and IL-6 levels decrease significantly at 3 months in the responder group compared with baseline. At day 90 we identified a cytokine profile which differentiates responders and non-responders. High serum levels of two proinflammatory cytokines, monocyte chemoattractant protein-1 and epidermal growth factor, were significantly higher in the responder group at day 90 compared with non-responders. However, we were not able to identify a baseline cytokine profile predictive of a good response at 3 months. These findings suggest that cytokine profiling by proteomic analysis may be a promising tool for monitoring rituximab and may help in the future to identify responder RA patients.

In general, blood gas analysers can also determine the value of haematocrite by measuring the blood's conductivity. The question to ask is whether this value is reliable. In this study, hematocrit obtained via conductivity from 6 different pieces of equipment were compared with those measured using the gold standard method, which is microcentrifugation. By interpreting the results of 320 arterial blood samples taken in the intensive care unit DAR "B" we can see that the reliability between two measurements on the same piece of equipment is very good, in general0.95 whatever the equipment. The reliability between the means of the two measurements and the gold standard is slightly lower but remains very satisfactory, most often between 0.8 and 0.9. The Gem Premier 3000 (IL) analyser and the Roche OMNI S gave the best reliability compared with centrifugation. The Spearman coefficients between the mean values of the analysers and those of centrifugation were high, with the exception of the Rapidpoint 405. They are all statistically different from zero (p0.0001).

Since 2005, international guidelines propose a stadification for chronic renal failure based on the glomerular filtration rate (GFR) value. The performance of the creatinine-based equations allowing the estimation of GFR and the bias of the creatinine measurements is, more than ever, a crucial issue. The consequences for the clinical biologists are of importance. First, the Cockcroft-Gault formula must be replaced by the four variable-MDRD equation. Second, the biologists must chose from the "175" and the "186" versions of the MDRD equation. The first one fits the creatinine methods which are traceable to the reference method (liquid or gas chromatography coupled to mass spectrometry). The second equation must be used for creatinine methods, which are not traceable to the reference method. Today, only some enzymatic methods can prove that they are traceable to the reference method. For the colorimetric methods, future is inclear.

During the last years, GFR estimation has received substantial attention with a focus on comparing results of new formulas with GFR measurements, and standardization of creatinine assays. Calibration of creatinine should improve performances. However, frequently used equations have lower precision in high GFR populations. This is the reason why a continuous effort in improving predicting equations is still needed. The use of calibrated creatinine, the onset of new GFR markers such as cystatin C, and pooling data across many study populations are underway to develop better prediction.

SummaryIn rheumatoid arthritis (RA) there are currently no useful indicators to predict a clinical response to tumour necrosis factor-α (TNF-α) blockade. The purpose of this study was to determine the role of peripheral blood cytokine profiling in differentiating between a good versus poor response to etanercept in RA. Peripheral blood samples were collected at baseline and at 3 months from 33 patients with active disease who were treated twice weekly by etanercept therapy. Responders are defined by the presence of three of four American College of Rheumatology criteria: ≥20% decrease in C-reactive protein (CRP), visual analogue score of disease activity, erythrocyte sedimentation rate and improvement of the disease activity score (28; four values) by ≥1·2 obtained at 3 months. Twelve cytokines were measured from serum collected on days 0 and 90 by proteomic array (protein biochip array, Investigator Evidence, Randox France), including interleukin (IL)-6, TNF-α, IL-1a, IL-1b, IL-2, IL-8, interferon-γ, IL-4, IL-10, monocyte chemoattractant protein (MCP)-1, epidermal growth factor (EGF) and vascular endothelium growth factor. Our results showed that high serum levels of MCP-1 and EGF were associated with a response to etanercept. In addition, the increase of two combined parameters CRP and EGF was predictive of a response to etanercept treatment at 3 months (sensitivity: 87·5% and specificity: 75%, accuracy: 84·4%). These findings suggest that cytokine profiling by proteomic analysis before treatment initiation may help to identify a responder patient to TNF-α blocking agents in RA.

Electrolyte disorders are frequently observed in nephrology and intensive care unit department and Na determination is routinely performed in biochemistry laboratories. Three methods are currently available. Flame photometry remains the reference method. With this method the serum sample is diluted before the actual measurement is obtained. Results are expressed as molarity (per Liter of plasma). Potentiometric methods have an increasing importance due to the advances in ion sensitive (selective) electrodes (ISE). Whereas the instruments for routine chemical analysis typically use indirect potentiometry (involving te dilution of samples) to measure sodium levels, the equipment for measuring arterial blood gases use direct potentiometry without any dilution. Thus, results obtained with indirect potentiometry are expressed in molarity (per liter of plasma) while results obtained with direct potentiometry are initially given in morality (per kg of plasma water) then converted in molarity. Analytical performances are in all cases satisfactory and therefore all the methods could be used in both normal and pathological ranges. Methods involving sample dilution such as flame photometry or indirect potentiometry, the serum sodium value would be expected to be low in case of decrease plasma water (pseudohyponatremia). By contrast, with direct potentiometry where no sample dilution takes place, no interference would be expected since the activity of sodium in the water phase only is being measured. Thus, the classical pseudohyponatremia observed with hyperlipemia or paraproteinemia are not further observed with direct potentiometry. These differences in methodology should be taken into account to explain discrepancies between results obtained with classical biochemistry analyser and with blood gas apparatus.

Residual renal function (RRF) contributes to the achievement of treatment adequacy in CKD-5 patients. It may facilitate patients' acceptance of renal replacement therapy (RRT) in minimizing dietary and fluid restriction. It has been confirmed to improve dialysis patient outcomes. Attempts to preserve RKF in incident CKD-5 patients are still subject to controversies. In this review we analyze the role of RRT in dialysis patient. What are the positive and the beneficial effects of maintaining RRF? What are the negative and the risks of maintaining a RRF? At what expense the maintenance of RRF is achieved? Preservation of RRF is undoubtedly an interesting means to enhance the efficacy of renal replacement therapy and reduce dietary fluid restriction. However, maintainance of RRF should not be considered as a goal of dialysis adequacy in dialysis patients but rather a means of optimizing RRT. Further, preservation of RRF should be considered as a permanent trade-off between patient comfort and chronic fluid volume overload with its deleterious effects.

An evaluation process directed by the biochemistry department of the UHC of Montpellier about acquisition of blood gas analysers for a point of care testing in 10 different medical care services. The values obtained by the different measurement machines have been compared taking into account the intra and inter measures variabilities. In this work, the results of 4 machines tested: Rapid-point 405, GEM Premier 3000, i-STAT 1 (série 300) and ABL 77 are compared with one i-STAT (série 200) and one OMNI S already available in the laboratory. The study lasted one month (november 2004). Each week, the two machines of the laboratory was compared with a new analyser randomly chosen between the four machines tested. In this study, including 21000 values, the statistical tests have been applied under the hypothesis that none of the measurement machine could be a "gold standard". Beside the classical research of correlation between the different values, we added the research of concordance between machines and the application of imperfect gold standard method. A high level of statistic concordance was observed between the different analysers. So the equipment has been selected using biological, analytic and computing criteria.

Capillary zone electrophoresis of serum proteins is increasingly gaining impact in clinical laboratories. During 2003, we compared the fully automated capillary electrophoresis (CE) system from Beckman (Paragon CZE 2000) with the method agarose gel electrophoresis Sebia (Hydrasis-Hyris, AGE). This new study focused on the evaluation of analytical performance and a comparison including 115 fresh routine samples (group A) and a series of 97 frozen pathologic sera with suspicion of monoclonal protein (group B). Coefficients of variation (CVs %) for the five classical protein fractions have been reported to be consistenly9% in within-run and10% in between-run imprecision studies with the Paragon 2000 system. The results of the comparison study (group A) demonstrated a good correlation between the CE system and AGE, except for beta-globulin (r = 0.65). Among the 97 pathologic serum samples (group B), there were 90 in which we detected a monoclonal protein by immunofixation (IF) (immunosubtraction (IS) was not used). AGE and Paragon 2000 failed to detect 7 and 12 monoclonal proteins, respectively, leading to a concordance to 92% for AGE and 87% for Paragon 2000 for identifying electrophoretic abnormalities in this group. Beta-globulin abnormalities and M paraprotein were well detected with Paragon 2000. Only 81% (21 vs 26) of the gammopathies were immunotyped with IS by two readers blinded to the IF immunotype. The Paragon 2000 is a reliable alternative to conventional agarose gel electrophoresis combining the advantages of full automation (rapidity, ease of use and cost) with high analytical performance. Qualified interpretation of results requires an adaptation period which could further improve concordance between the methods. Recently, this CE system has been improved by the manufacturer (Beckman) concerning the migration buffer and detection of beta-globulin abnormalities.

Monitoring beta2-microglobulin (beta2M) in biological fluids has gained considerable interest in pathologies such as haematologic malignancies, renal diseases, and chronic inflammatory diseases. Due to limitations of the RIA in the routine laboratory, we measure beta2M with non-isotopic methods. 189 patients suffering from myeloma (n=66), end stage renal failure (n=54) or inflammation (n=69) were included in this study. beta2M was determined in serum, urine and dialysate using an immunoenzymometric assay with chemiluminescence detection [Immulite Diagnostic Products Corporation (DPC), La Garenne Colombes, France] and an immunoturbidimetric assay (Olympus, Rungis, France). The data were compared with a radioimmunoassay (Immunotech, Marseille, France) taken as a reference. Using serum samples, the immunoenzymometric assay with chemiluminescence detection and the immunoturbidimetric assay have reliable analytical performances. Values obtained with serum samples are highly correlated with the radioimmunoassay (DPC/RIA r2=0.84; Olympus/RIA r2=0.94) whatever the type of pathology; however an over-estimation which could be related to cross reactivity with beta2M fragments was observed with the RIA method as suggested by crossover calibration and recovery studies. Values obtained with urinary samples (n=96) are closely related to those obtained with the RIA (DPC/RIA r2 = 0.98; Olympus/RIA: r2=0.99). Despite the low levels observed in dialysate (n=57) good correlations were observed between Olympus vs DPC (r2=0.85). By contrast, the two non-isotope methods are poorly related with the RIA method (DPC vs RIA r2=0.47 and Olympus vs RIA r2=0.54). In conclusion, the immunoenzymometric assay with chemiluminescence detection or the immunoturbidimetric assay could be used in the routine laboratory in order to determine beta2M in plasma, urine and dialysate.

The POLA (Pathologies Oculaires Liées à L'Age) Study is a population-based study of cataract and age-related macular degeneration and their risk factors being carried out among 2,584 residents of Sète, southern France, aged 60-95 years. Recruitment took place between June 1995 and July 1997. Cataract classification was based on a standardized lens examination by slit lamp, according to Lens Opacities Classification System III. This paper presents results obtained from cross-sectional analysis of the first phase of the study. In polytomous logistic regression analyses, an increased risk of cataract was found for female sex (cataract surgery: odds ratio (OR) = 3.03; cortical cataract: OR = 1.67), brown irises (cortical, nuclear, and mixed cataracts: OR = 1.61), smoking (cataract surgery: OR = 2.34 for current smokers and OR = 3.75 for former smokers), known diabetes of 10 or more years' duration (posterior subcapsular, cortical, and mixed cataracts and cataract surgery: OR = 2.72), use of oral corticosteroids for at least 5 years (posterior subcapsular cataract: OR = 3.25), asthma or chronic bronchitis (cataract surgery: OR = 2.04), cancer (posterior subcapsular cataract: OR = 1.92), and cardiovascular disease (cortical cataract: OR = 1.96). Decreased risk of cataract was found with higher education (all types of cataract and cataract surgery: OR = 0.59), hypertension (cataract surgery: OR = 0.57), and high plasma retinol levels (nuclear and mixed cataracts and cataract surgery: OR = 0.75 for a 1-standard-deviation increase). Most of the risk factors identified in this study confirm the findings of other studies. The association of cataract with plasma retinol level requires further investigation.

To give the levels of antioxidant nutrients in relation to age-related macular degeneration (AMD).Pathologies Oculaires Liees a l'Age is a population-based study on cataract and AMD and their risk factors, carried out on 2584 inhabitants of Sete, France. Age-related macular degeneration was defined by findings from fundus photographs according to an international classification. Biological measurements were taken from fasting blood samples.After multivariate adjustment, plasma alpha-to-copherol levels showed a weak negative association with late AMD (P = .07). Lipid-standardized plasma alpha-tocopherol levels showed a significant negative association with late AMD (P= .003): the risk of late AMD was reduced by 82% in the highest quintile compared with the lowest. Similarly, lipid-standardized plasma alpha-tocopherol levels were inversely associated with early signs of AMD (odds ratio, 0.72 [95% confidence interval, 0.53-0.98]; P=.04). No associations were found with plasma retinol and ascorbic acid levels or with red blood cell glutathione values.These results suggest that vitamin E may provide protection against AMD. Only randomized interventional studies could prove the protective effect of vitamin E on AMD.

Chronic rejection remains the single most important cause of renal allograft loss after the first year post-transplant. We performed a matched case control study within our cohort of 471 renal allograft recipients, comparing 66 patients with histologically proven chronic rejection with 66 controls. Analysis of immunological (transfusion, sensitisation, HLA matching, number of transplantation, number of acute rejections (AR), immunosuppression) and non-immunological (donors and recipients age and sex, CMV disease, post-transplant acute tubular necrosis, cold ischemia) factors which could predict the occurrence of chronic rejection (CR) was performed, using Wilcoxon rank test, Mac Nemar test and Cox model. Univariate analysis showed that potential risk factors for CR are: donor age45 years (p = 0.05), recipient age40 years (p = 0.008), CMV disease (p = 0.03), number of acute rejection episodes (p = 0.009), retransplantation (p = 0.002). Multivariate analysis showed that only the following factors significantly increased the risk of CR: AR episodes (p = 0.01) with an odds-ratio at 3.5 (95% CI = 1.3-3.9) for the second acute rejection episode and at 6.5 (95% CI = 1.5-29.4) for the third acute rejection episode, donor age45 years (p = 0.03) with an odds-ratio at 3.5 (95% CI = 1.1-10.6). Our data suggest that better matching at donor recipient age and more potent immunosuppressive protocols resulting in no acute rejection may improve the long term graft survival. They also show that the use of old donors (45 years), as a response to organ shortage is detrimental for long term renal function.

Oxidative mechanisms may play an important role in the etiology of cataract and age-related macular degeneration (AMD). The authors present the level of two antioxidant enzymes in relation to cataract and AMD.Population-based, cross-sectional study on cataract and AMD and their risk factors.This study includes 2584 participants recruited among the residents of the town of Sète (in the south of France), who were 60 years of age or older.Cataract was defined on the basis of slit-lamp examination, according to the Lens Opacities Classification System III, and AMD on the basis of fundus photographs according to an international classification. Biologic measurements were made centrally from blood samples for which the patient fasted.The presence of early and late AMD and of subcapsular, cortical, nuclear, and mixed cataracts was assessed and related to the levels of plasma glutathione peroxidase and erythrocyte superoxide dismutase.After multivariate adjustment, higher levels of plasma glutathione peroxidase (pIGPx) were significantly associated with a ninefold increase in late AMD prevalence, a sixfold increase in cortical cataract, and a twofold increase in nuclear and mixed cataracts. High levels of erythrocyte superoxide dismutase (SOD) activity were not associated with late AMD and early signs of AMD but were associated with a twofold increase in nuclear cataract.The authors show here, for the first time, a strong association of high levels of pIGPx with age-related eye diseases. High levels of SOD also are associated with increased risk of nuclear cataract. More data are needed at the biochemical and epidemiologic levels for a better understanding of these findings.

Malondialdehyde assay is the most generally used test in the appreciation of the role of oxidative stress in disease. Malondialdehyde is one of several products formed during the radical induced decomposition of polyunsaturated fatty acids. Most often, malondialdehyde assay used its reactivity at high temperature and low pH, towards thiobarbituric acid. This reaction is very sensitive but its specificity, even with improvement of pre-analytical (sampling, preservatives), and analytical stages (fluorescence, HPLC) is still a matter of debate. At present, the concept of "thiobarbituric acid reactive substances" (TBARS) have merged and progressively replaced the initial malondialdehyde assay. In this review, we presented the main results concerning the assays of TBARS and malondialdehyde in blood and different biological medium. In the future, oxidative stress appreciation will need the precise analytical determination of different molecules triggered by free radicals. The TBARS assay should be considered as a global test, allowing a global approach of lipoperoxidation whereas specific determination of malondialdehyde can only appreciate one of the end-product formed during oxidative stress.

The histological picture of chronic rejection with endothelial lesions and vascular hyperplasia resembles the early arteriosclerotic lesions. As increasing evidence suggests a role for oxidative stress in arteriosclerosis, we examined whether chronic rejection in renal transplant recipients was associated with increased oxidative stress markers.We investigated lipid metabolism and oxidative stress in 77 renal transplant recipients. Group I patients (n=34; 48+/-2 years old, 12 women, 22 men) had no clinical or histological signs of chronic rejection, whereas group II patients (n=43; 47+/-3 years old, 15 women, 28 men) had histologically proven chronic rejection. All patients were treated with cyclosporine and steroids. Lipid metabolism was evaluated by determining total cholesterol, triglycerides, high-density lipoprotein cholesterol, apolipoproteins AI and B, and lipoprotein (a). Oxidative stress was evaluated by determining: (i) the end product of lipid peroxidation, malonyldialdehyde (MDA), and erythrocyte polyunsaturated fatty acids; (ii) the nonenzymatic antioxidant system: erythrocyte alpha-tocopherol and glutathione; and (iii) the enzymatic antioxidant system: erythrocyte superoxide dismutase and plasma glutathione peroxidase. Results were compared with those of a control group (38 healthy volunteers).Compared with controls, renal transplant recipients had significantly increased total cholesterol, triglyceride, and apolipoprotein B levels; they also had, in association with these lipid abnormalities, a significant increase in MDA and a significant decrease in erythrocyte polyunsaturated fatty acids, as well as a significant decrease in enzymatic and nonenzymatic antioxidant defense mechanisms. In contrast to lipid disturbances, where no difference was observed between groups I and II, markers of oxidative stress were significantly higher in group II compared with group I (MDA: 1.87+/-0.43 and 1.62+/-0.31 nmol/ml, respectively, P0.05). The red blood cell antioxidative defense mechanisms were significantly decreased in group II compared with controls (erythrocyte alpha-tocopherol: 0.61+/-0.38 and 1.08+/-0.31 mg/L, respectively, P0.01; superoxide dismutase: 1.08+/-0.2 and 1.32+/-0.31 U/mg Hb, respectively, P0.01).Our data show that oxidative stress with a decrease in antioxidant defenses is associated with kidney transplantation. In addition, oxidative stress markers are particularly increased in transplant recipients with chronic rejection, which suggests that oxidative stress may participate in the development and/or progression of vascular lesions observed in these patients.

Cardiovascular diseases represent the first cause of mortality in chronic renal failure patients treated by hemodialysis. Alterations in lipid metabolism and oxidative stress are recognized as vascular risk factors. Their corrections could be of interest for atherosclerosis prevention. In order to evaluate interest of an therapeutic intervention, we have analyzed oxidative metabolism in hemodialysis patients by determining the production of oxygen reactive species (ROS), the level of defense mechanisms, and the balance between nitric oxide (NO) and ROS, responsible for anti- or proxidant effects of NO. During dialysis sessions performed with cellulosic membrane (Cuprophan) an increase in hydroperoxide production by platelets was noted (12 HETE) (5.62 +/- 0.94 pg); similarly, superoxide anion (O2(0)-) production by monocytes (fluorescence index: 115 +/- 24) and by polynuclear cells (fluorescence index: 115 +/- 24) was enhanced. On the other hand, anti-oxidant defenses were significantly reduced with a decrease in RBC SOC activity (0.92 +/- 0.06 U/mg Hg) and in RBC vitamin E (0.7 +/- 0.07 mg/l) concentration. We have demonstrated a profound alteration in the L-arginine/NO pathway consequently to an accumulation of NO synthases inhibitors or activators. The necessity to reduce the production of ROS during dialysis sessions justifies the use of more biocompatible membranes, such as modified cellulosic or synthetic membranes, decreasing leucocyte activation. In addition, NO synthetase inhibitors can be preferentially eliminated by convection. Finally, a supplementation with an exogenous anti-oxidant, such as oral vitamin E (500 mg/day for 6 months) normalizes RBC vitamin E levels and concomitantly allows a decrease in MDA concentrations In conclusion, oxidative metabolism alterations observed in hemodialysis are multifactorial: preventive measures include the use of a more biocompatible material, the reequilibrium of the NO/ROS balance, and supplementation with exogenous anti-oxidants.

Plasmapheresis and low-density lipoprotein (LDL)-apheresis are recognized procedures for the treatment of hyperlipidemia resistant to diet and lipid-lowering drugs and provide information on cholesterol synthesis in hypercholesterolemic patients. However, cholesterol synthesis after acute cholesterol removal from plasma has never been investigated in normocholesterolemic patients. In this study, cholesterol synthesis was evaluated in three normocholesterolemic patients by determination of plasma lathosterol, lathosterol-to-cholesterol ratio, and plasma mevalonic acid. In a short-term kinetic study, samples were collected before and after plasmapheresis and every 6 hours during 24 hours. In the second part of the study, cholesterol synthesis was evaluated daily for 3 days. In normocholesterolemic patients, cholesterol returns to basal levels in 3 days. However, cholesterol removal did not result in a significant increase in lathosterol-to-cholesterol ratio or in plasma mevalonic acid, despite a slight increase in lathosterol. In contrast, when repeated plasma exchanges induced a dramatic hypocholesterolemia (1 mmol/liter), an acute but transient stimulation of cholesterol synthesis was observed (lathosterol/cholesterol ratio and MVA, respectively, increase from 8.2 to 22.3 and from 28 nmol/liter to 98 nmol/liter). This study shows that cholesterol synthesis is not stimulated by plasmapheresis in normocholesterolemic patients but is enhanced in dramatic hypocholesterolemic patients (1 mmol/liter).

Lipoprotein glomerulopathy is defined by the presence of lipidic deposits in the capillary lumen giving them a dialted and microaneurysmal aspect and the presence of quantitative and/or qualitative alterations of plasma apolipoprotein E. We describe here the long-term follow-up of a young female patient who presented with corticoresistant nephrotic syndrome in 1979 and progressed to chronic renal failure requiring dialysis in 1990. The three renal biopsies performed during the follow-up showed markedly enlarged capillary loops due to intra-luminal fibrinolipidic material deposition forming true intracapillary thrombi. She received a cadaver renal transplant in 1993. One year after transplantation, nephrotic syndrome reappeared and graft biopsy showed recurrence of the initial glomerular disease on the transplant. The plasma lipid profile showed hypercholesterolemia, hypertriglyceridemia, and elevated plasma ApoE levels with an abnormal Apo E phenotype. Our case report is a new typical case of lipoprotein glomerulopathy with recurrence of the initial disease on the renal allograft.

Superoxide (O2-) and nitric oxide (.NO) are free radicals which are known to react together leading to peroxynitrite anions that can decompose to form nitrogen dioxide (NO2) and hydroxyl radical (OH degrees). .NO has been reported to have a dual effect on LDL oxidation (pro or antioxidant). In the present study we have investigated in vitro the action of exogenous .NO on human LDL oxidation promoted by oxygen, copper or (2,2'-azobis (2-aminodinopropane hydrochloride)(ABAP). .NO was given as NO donnor (sodium nitroprussiate or S-nitroso-L-glutathione) from 10 to 500 microM. Oxidation of LDL was measured by monitoring continuously conjugated diene formation at 234 nm. Exogenous .NO inhibited in a dose dependent manner the progress of spontaneous LDL oxidation. Copper--or ABAP--induced oxidation were characterized by lag, propagation and decomposition phases. Exogenous .NO decreased the propagation rate of LDL oxidation and the level of maximal diene production. These effects are different of these of alpha-tocopherol, a chain-breaking antioxidant. In our experimental conditions, .NO exhibited antioxidant activities. In vivo, the continuous release of endogenous .NO could protect LDL from cell-induced oxidation.

Nitric oxide (NO) is a free radical produced enzymatically in biological systems from the guanidino group of L-arginine. Its large spectrum of biological effects is achieved through chemical interactions with different targets including oxygen (O2), superoxide (O2o-) and other oxygen reactive species (ROS), transition metals and thiols. Superoxide anions and other ROS have been reported to react with NO to produce peroxynitrite anions that can decompose to form nitrogen dioxide (NO2) and hydroxyl radial (OHo). Thus, NO has been reported to have a dual effect on lipid peroxidation (prooxidant via the peroxynitrite or antioxydant via the chelation of ROS). In the present study we have investigated in different models the in vitro and in vivo action of NO on lipid peroxidation. Copper-induced LDL oxidation were used as an in vitro model. Human LDL (100 micrograms ApoB/ml) were incubated in oxygene-saturated PBS buffer in presence or absence of Cu2+ (2.5 microM) with increasing concentrations of NO donnors (sodium nitroprussiate or nitroso-glutathione). LDL oxidation was monitored continuously for conjugated diene formation (234 nm) and 4-hydroxynonenal (HNE) accumulation. Exogenous NO prevents in a dose dependent manner the progress of copper-induced oxidation. Ischaemia-reperfusion injury (I/R), characterized by an overproduction of ROS, is used as an in vivo model. Anaesthetized rats were submitted to 1 hour renal ischaemia following by 2 hours of reperfusion. Sham-operated rats (SOP) were used as control. Lipid peroxidation was evaluated by measuring the HNE accumulated in rats kidneys in presence or absence of L-arginine or D-arginine infusion. L-arginine, but not D-arginine, enhances HNE accumulation in I/R but not in SOP (0.050 pmol/g tissue in SOP versus 0.6 nmol/g tissue in I/R), showing that, in this experimental conditions, NO produced from L-arginine, enhances the toxicity of ROS. This study shows that the pro- or antioxydant effects of NO are different in vivo and in vitro and could be driven by environmental conditions such as pH, relative concentrations of NO and ROS, ferryl species.

Interest in NO measurement strongly increased with the discovery that NO is endogenously produced by living tissues. This review describes the major techniques for quantification of NO and derivatives in biological models.

Monoclonal antibodies were proposed as an effective prophylactic immunosuppressive treatment in highly sensitized patients (HSP). In this study we compared the results obtained in HSP treated with OKT3 or antilymphocyte globulins (ALG). From January 1989 to January 1993, 38 transplantations were performed in patients with high panel reactive antibodies (PRA > 50%). The group comprised 22 women and 16 men, mean age 45 +/- 2 (23-67) years; ten were second grafts and two were third grafts. Peak PRA was > or = 80% in 24 sensitized patients and 50-80% in 14 sensitized patients. Patients were randomly assigned to either prophylactic OKT3 (n = 15) or ALG (n = 23). Oral cyclosporin A (10 mg/kg) was started at day 8 in the OKT3 group and when the serum creatinine level decreased to 200 micromol/l in the ALG group. OKT3 was systematically withdrawn on day 10 but ALG was stopped only when total blood cyclosporin A concentration reached 150-200 ng/ml. In both groups, azathioprine (150 mg/day) and prednisolone were given. During the first months, 6/15 grafts were lost in the OKT3 group (three hyperacute rejections, one renal vein thrombosis, one steroid-resistant rejection, one death); in the ALG group 4/23 grafts were lost (one hyperacute rejection, two steroid-resistant rejections, one death). Side effects were significantly more frequent in the OKT3 group than in the ALG group. After 12 months of follow up, the graft survival was 71% (27/38) and did not significantly differ (log-rank test, NS) between the OKT3 (60%, 9/15) and the ALG group (78%, 18/23). We conclude that the use of the monoclonal antibody OKT3 as a prophylactic agent in HSP does not improve the early graft survival when compared with prophylactic ALG. Polyclonal antibodies, which react with many epitopes and are much better tolerated seem to offer a good strategy for induction therapy in this population.

Leukocyte activation during haemodialysis (HD) was evaluated by reactive oxygen species (ROS) production and cell surface markers expression (CD11b: C3bi receptor and CD25: IL2 receptor). Eight end-stage renal disease patients were exposed to three dialysis phases according to a A/B/A protocol study. During phase A polysulphone (PS) membranes were used and during phase B cuprophane (CU) membranes were used. Each phase lasted 3 weeks. Timed samples were collected during the last session of each phase at 0, 15, and 30 min of HD. Flow cytometry analysis was performed both on monocytes (MO), polymorphonuclears (PMN), and lymphocytes (Ly). Hydroethydine was used as a marker of cell-ROS production. Specific monoclonal antibodies were used to analyse the cell surface markers. CU increased ROS production in PMN and MO by 10- and 2.4-fold respectively and had no significant effect on Ly. CU enhanced 16-fold CD11b expression on PMN, and increased also CD11b and CD25 expression on MO by 7- and 40-fold respectively. On the contrary, PS did not affect either ROS production or cell surface markers expression in MO, PMN, Ly. We conclude that oxydative metabolism and cell surface markers expression of PMN and MO were significantly increased with cuprophane membranes and not with polysulphone membranes, suggesting that complex cell-cell interactions were involved in membrane-related bioincompatibility phenomena.

BackgroundLipids appear to mediate depressive vulnerability in the elderly, however, sex differences and genetic vulnerability have not been taken into account in previous prospective studies.MethodsDepression was assessed in a population of 1040 women and 752 men aged 65 years and over at baseline and after 7-year follow-up. Clinical level of depression (DEP) was defined as having either a score of 16 and above on the Centre for Epidemiology Studies Depression scale or a diagnosis of current major depression on the Mini International Neuropsychiatric Interview. Lipid levels, apolipoprotein E and serotonin transporter linked promoter region (5-HTTLPR) genotypes were evaluated at baseline.ResultsMultivariate analyses adjusted by socio-demographic and behavioral variables, measures of physical health including ischemic pathologies, and genetic vulnerability indicated gender-specific associations between dyslipidemia and DEP, independent of the use of lipid lowering agents or apolipoprotein E status. Men with low LDL-cholesterol levels had twice the risk of prevalent and incident DEP whereas in women low HDL-cholesterol levels were found to be significantly associated with increased prevalent DEP (OR = 1.5) only. A significant interaction was observed between low LDL-cholesterol and 5-HTTLPR genotype, men with s/s or s/l genotype being at increased risk of DEP (OR = 6.0 and 2.7, respectively). No significant gene-environment interaction was observed for women.ConclusionsDEP is associated with higher atherogenic risk in women (low HDL-cholesterol), whereas the reverse is observed in men (low LDL-cholesterol). Late-life depression may have a complex gender-specific etiology involving genetic vulnerability in men.

The use of erythropoietin (Epo) in chronic renal failure patients improves anemia and avoids iterative transfusions. As a consequence, a significant decrease of anti-HLA antibodies might be observed. From 31.12.1985 to 30.07.1991, among the 61 highly sensitized patients (pts) waiting for renal transplantation at our institution, 23 (7 men, 16 women, mean age 43.3 +/- 2.3 years) were treated with Epo during 21.4 +/- 1.7 months. After introduction of Epo, the mean number of transfusions significantly decreased from 35 +/- 8 to 0.5 +/- 0.4 (p0.0001) and the Panel Reactive Antibodies decreased from 88.1 +/- 1.7 to 18.8 +/- 5.6% (p0.0001). HLA antibodies totally disappeared in 12 patients, decreased more than 30% in 9 patients and remained stable in the 3 others. Renal transplantation was performed in 9 patients; 4 with a negative cross-match in both historical and current sera and 5 with a negative current cross-match but with a positive historical cross-match. 6/9 patients are doing well with a good graft function; 2 patients returned to hemodialysis for rejection and 1 patient died with a functioning kidney. Since Epo permits the transfusion withdrawal, its introduction in chronic renal failure treatment suppresses both the main factor of immunisation and one of the most important mechanisms of persistence of antibodies.

Human adult Parvovirus B19 infection is a well known disease most frequently seen in haematology and cancerology. To our knowledge, however, renal involvement in this context has never been reported. A case of severe infection caused by this virus is reported, where multiple organ involvement is associated with acute renal failure. The outcome was favourable with temporary dialysis and symptomatic treatment. Mechanisms of renal failure, presumably multifactorial, are discussed in the light of this case.

Many elderly patients (aged over 60) are waiting for renal allograft but are considered a risk population for myocardial infarction or sudden cardiac death. Between 1987 and January 1st, 1990, 22 elderly patients (16 men and 6 women; mean age 63.4 +/- 0.7 years; range 60-69 years) underwent cadaveric kidney transplantation. Twenty patients had been on haemodialysis for 4.8 +/- 1 years (range: 1-16.3), one had been on peritoneal dialysis for 5 years and one had developed chronic rejection of a previous cadaveric allograft. Among these 22 patients, 6 (27 percent) developed antibodies directed against the random lymphocyte panel and one had a second transplant. Cardiovascular risk factors were systematically estimated, including medical history, cervical arterial Doppler, echocardiography and stress thallium testing (STT). Coronary angiography was performed if there was evidence of myocardial ischaemia. STT did not show any coronary disease in 16 patients and was not maximal (less than 60 percent of maximal theoretical stress) in 4 patients. In 2 patients, STT showed myocardial ischaemia: one, with a history of angina, underwent coronary angiography and angioplasty; the other had silent ischaemia and refused coronary angiography. Two patients died of myocardial infarction. One of them was the patient with silent ischaemia and positive STT test. The patient with angioplasty had a successful transplantation 20 months later, without any cardiovascular complication. After 9 to 32 months of follow-up, 19 patients have functioning allografts (mean serum creatinine level 151 +/- 6 mumol/l). One patient lost his kidney from urinary fistula. Eight patients (36 percent) developed acute rejection and all responded to intravenous corticosteroids. No immunological transplant loss was observed. After 18 months, the graft actuarial survival rate was 85 percent. We conclude that elderly patients free of clinical or silent myocardial ischaemia are good transplant recipients. The cardiovascular risk could be prevented by using STT during the pretransplant screening. If the STT is not maximal or shows coronary ischaemia, coronary angiography is mandatory.