Isabel Ramis

Markers of early renal changes induced by industrial pollutants. II. Application to workers exposed to lead.

The present study has been carried out in the framework of a collaborative research project on the development of new markers of nephrotoxicity. A battery of more than 20 potential indicators of renal changes has been applied to 50 workers exposed to lead (Pb) and 50 control subjects. After application of selection criteria 41 exposed and 41 control workers were eventually retained for the final statistical analysis. The average blood Pb concentration of exposed workers was 480 micrograms/l and their mean duration of exposure was 14 years. The battery of tests included parameters capable of detecting functional deficits (for example, urinary proteins of low or high molecular weight), biochemical alterations (for example, urinary eicosanoids, glycosaminoglycans, sialic acid) or cell damage (for example, urinary tubular antigens or enzymes) at different sites of the nephron or the kidney. The most outstanding effect found in workers exposed to Pb was an interference with the renal synthesis of eicosanoids, resulting in lower urinary excretion of 6-keto-PGF1 alpha and an enhanced excretion of thromboxane (TXB2). The health significance of these biochemical alterations, detectable at low exposure to Pb is unknown. As they were not associated with any sign of renal dysfunction, they may represent reversible biochemical effects or only contribute to the degradation of the renal function from the onset of clinical Pb nephropathy. The urinary excretion of some tubular antigens was also positively associated with duration of exposure to Pb. Another effect of Pb that might deserve further study is a significant increase in urinary sialic acid concentration.

Nephropathies and exposure to perchloroethylene in dry-cleaners

Even in specific risk groups, the relation between exposure to organic solvents and chronic renal diseases remains controversial. Thus, in a collaborative European study, we assessed the renal effects of occupational exposure to perchloroethylene (PCE) in dry-cleaners compared with matched controls who were simultaneously examined. Single high and low molecular weight proteins, kidney-derived antigens and enzymes, and prostanoids were measured in urine. beta 2-microglobulin, creatinine, laminin fragments, and anti-glomerular basement membrane antibodies were also measured in serum. A canonical function based on 23 such variables correctly classified 93% of individuals as either PCE-exposed or controls; with 13 markers, group membership was identified in 87% of subjects. Increased high molecular weight protein in urine was frequently (17/50 vs 1/50, p less than 0.0001) associated with tubular alterations. Changes were consistent with diffuse abnormalities along the nephron in workers exposed to low levels of PCE (median 15 parts per million). Generalised membrane disturbances might account for the increased release of laminin fragments, fibronectin, and glycosaminoglycans, for high molecular weight proteinuria, and for the increased shedding of epithelial membrane components from tubular cells with different location along the nephron (brush-border antigens and Tamm-Horsfall glycoprotein). These findings of early renal changes indicate that solvent-exposed subjects, especially dry-cleaners, need to be monitored for the possible development of chronic renal diseases.

Differential activity of nitric oxide synthase in human nasal mucosa and polyps

Nitric oxide (NO) plays an important regulatory role in airway function and seems to be implicated in the pathophysiology of several airway diseases. To better understand the involvement of NO in the upper airways, we examined the presence of nitric oxide synthase (NOS) activity in human nasal mucosa and nasal polyp tissues. Nasal mucosa was obtained from seven patients undergoing septoplasty, and nasal polyps came from nine patients following polypectomy. NOS activity was quantified in tissue homogenates using the citrulline release assay and localized in tissue sections using reduced nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase histochemistry. The results showed that nasal polyps (n = 9) contained higher levels of total NOS activity (mean +/- SD 5.94 +/- 5.71, range 1.29-18.0 pmol.min-1.mg protein) than nasal mucosa tissues (n = 7) (0.28 +/- 0.22, range 0.01-0.57 pmol.min-1.mg protein). In addition, nasal polyps mainly contained inducible NOS activity (4.67 +/- 4.57, range 1.23-15.5 pmol.min-1.mg protein) whereas in nasal mucosa all NOS activity detected was in constitutive form. In both cases, NOS activity was localized in the epithelial cells. Since NO synthase is induced in inflamed upper airways, we conclude that NO may be an important inflammatory mediator in the respiratory system and that the epithelium may be a source of NO production in the human upper airways.

Systemic prostacyclin in cirrhotic patients: Relationship with portal hypertension and changes after intestinal decontamination

The total body production of prostacyclin was shown to be increased in cirrhotic patients, suggesting that its synthesis by blood vessels of the systemic circulation is enhanced. However, the mechanism by which the synthesis of systemic prostacyclin is stimulated is not known. The present study investigated the urinary excretion of 2,3-dinor-6-keto-PGF1 alpha, an index of total body prostacyclin synthesis, first, in cirrhotics with portal hypertension (n = 19) as compared with cirrhotics with reduced portal pressure after portacaval shunt surgery (n = 18) and with control noncirrhotic subjects (n = 11), and; second, in cirrhotics before and after intestinal decontamination by oral nonabsorbable antibiotics (n = 9 antibiotic treated patients, n = 10 control nontreated cirrhotics). Control noncirrhotic subjects showed lower urinary excretion of 2,3-dinor-6-keto-PGF1 alpha than both groups of cirrhotics (P less than 0.001). Interestingly, urinary excretion of 2,3-dinor-6-keto-PGF1 alpha was significantly higher in cirrhotics with portacaval shunt than in those with portal hypertension (P less than 0.01). The urinary excretion of 2,3-dinor-6-keto-PGF1 alpha decreased significantly after intestinal decontamination in the antibiotic-treated group (580.1 +/- 232.4 vs. 431.2 +/- 219.2 pg/mg creatinine; P less than 0.05) but not in nontreated patients (543.9 +/- 214.4 vs. 581.2 +/- 281.4 pg/mg creatinine; P = NS). These data suggest that the increased urinary excretion of 2,3-dinor-6-keto-PGF1 alpha observed in cirrhotics is not directly related to portal hypertension itself but to portal blood factors that bypass the liver. Some such factors may be of intestinal bacterial origin.

Modern high-performance liquid chromatographic-radioimmunoassay strategies for the study of eicosanoids in biological samples

An evaluation of the most recent literature on the determination of eicosanoids by immunoassay methods confirms that owing to the inherent lack of specificity of many of the antibodies used for this purpose, immunological assays (radioimmunoassay or enzyme immunoassay) are often preceded by solid-phase extraction followed by further purification of the antigens of interest by routine reversed-phase high-performance liquid chromatographic methods. In this way the analytical potential of radioimmunoassay is remarkably enhanced and accuracy and precision of the assay are ensured.

Human exposure to hydrogen fluoride induces acute neutrophilic, eicosanoid, and antioxidant changes in nasal lavage fluid.

The development of asthmalike symptoms among aluminum potroom workers has been associated with exposure to fluorides. In the present study, the immediate nasal response in humans was examined subsequent to short-term hydrogen fluoride (HF) exposure. Ten healthy subjects were exposed to HF (3.3-3.9 mg/m 3) for 1 h. Nasal lavage (NAL) was performed before, immediately after, and 1.5 h after the end of exposure. Control lavages were performed on the same subjects at the same time points but without HF exposure. At the end of HF exposure, 7 of 10 individuals reported upper airway symptoms. A significant increase was observed in the number of neutrophils and total cells, while there was a decrease in cell viability. The changes in neutrophil numbers correlated significantly with the reported airway symptoms. HF also induced a significant increase in tumor necrosis factor-α and the total protein content of NAL fluid. Among the eicosanoids, prostaglandin E 2, leukotriene B 4, and peptide leukotrienes were elevated after exposure. Of the antioxidants measured, the concentration of uric acid increased after exposure. In conclusion, exposure to HF induced immediate nasal inflammatory and antioxidant responses in healthy human volunteers. These findings may contribute to a further understanding of the way HF exerts damage to the airways and show that HF could represent an occupational hazard.

Simultaneous reversed-phase extraction of lipoxygenase and cyclooxygenase metabolites of arachidonic acid in nasal secretions: Methodological aspects

An improved analytical method for the simultaneous solid-phase extraction of arachidonic acid metabolites in biological samples is described. The major aim of the work was to define the cause of the different recoveries reported in the literature for leukotrienes and hydroxyeicosatetraenoic acids. We used nasal lavages to carry out a comparative study of solid-phase extraction parameters of practical importance in securing good recoveries of leukotrienes and hydroxyeicosatetraenoic acids from biological samples. We evaluated the influence of the pH of the sample, the pH of the water used to wash the extraction cartridge before elution of the adsorbed analytes, the comparative behaviour of commercially available octadecyl adsorbents and the influence of the concentration-evaporation step on final recoveries. Data thus obtained show that there is no significant difference in results when the samples and the water used to wash the cartridge before analyte elution are adjusted to pH values ranging between 4.0 and 7.4. Below pH 4.0, losses may be significant. Furthermore, recoveries can be very much dependent on the type of solid-phase cartridge material and on the eluent evaporation method, especially with regard to aqueous phase removal.

Pyrazine-based Syk kinase inhibitors.

A series of aminopyrazines as inhibitors of Syk kinase activity and showing inhibition of LAD2 cells degranulation is described. Optimization of the carboxamide motif with aminomethylpiperidines provided high potency inhibiting Syk but low cellular activity. Amides of cis and trans adamantanol showed good inhibitory activity against Syk as well as remarkable activity in LAD2 cells degranulation assay.

Highly potent aminopyridines as Syk kinase inhibitors.

A novel class of potent Syk inhibitors has been developed from rational design. Highly potent aminopyridine derivatives bearing a 4-trifluoromethyl-2-pyridyl motif and represented by compound 13b IC(50): 0.6 nM were identified. Substitution by a 2-pyrazinyl motif and SAR expansion in position 4 of the central core provided diverse potent non-cytotoxic Syk inhibitors showing nanomolar activity inhibiting human mast cell line LAD2 degranulation.

Release of chemical mediators and inflammatory cell influx during early allergic reaction in the nose: Effect of furosemide

BACKGROUND We evaluated the effect of furosemide on allergen-induced rhinitis in a double-blind, crossover, placebo-controlled experiment. METHODS Fourteen patients with rhinitis who were allergic to house dust were nebulized with an intranasal dose of 20 mg of furosemide or placebo before allergen challenge with an extract of Dermatophagoides pteronyssinus (100 BU). Clinical evaluation and nasal lavages with normal saline solution were performed at baseline; after placebo-furosemide nebulization, and at 10, 30, and 60 minutes after allergen challenge. Number of sneezes and a composite symptom score were recorded to evaluate clinical response. Prostaglandin E2 (PGE2), PGD2 peptide leukotrienes and 15-hydroxy, 5,8,11,13-eicosatetraenoic acid (15-HETE) were measured by radioimmunoassay in nasal lavages. Cells were counted and classified as epithelial cells, neutrophils, eosinophils, and others. RESULTS No differences in either clinical symptoms or cell influx after allergen challenge were found between furosemide and placebo groups. PGE2 levels did not change after provocation, and furosemide had no effect on its production. Ten minutes after antigen challenge there was a marked increase of PGD2 (p < 0.01), peptide leukotrienes (p < 0.01), and 15-HETE (not significant) on both study days. However, no significant differences in the release of eicosanoids were found between furosemide and placebo groups. CONCLUSIONS Our observations in the nasal mucosa suggest that furosemide has no effect on the release of proinflammatory and bronchoconstrictor metabolites (PGD2, peptide leukotrienes, and 15-HETE). In contrast to bronchial asthma, allergen-induced rhinitis was not effectively prevented by furosemide.