Presence, enzymatic activity, and subcellular localization of paraoxonases 1, 2, and 3 in human lung tissues.

AIMS: The human paraoxonases family (PONs) includes three calcium-dependent esterases: PON1, PON2, and PON3. The presence of PONs mRNA in human lungs is known, however, their enzymatic activity and subcellular localization have not been sufficiently explored. MAIN METHODS: In this work, the presence of PONs in human lung tissues, at both mRNA and protein levels, was confirmed by Real-Time RT-PCR and Western blot analysis. Moreover, the activities of PONs were determined in cytosol and microsomes of 30 subjects and in mitochondria of 8 representative lung tissues using selective and non-selective substrates. Besides, to exclude the possible contribution of other esterases on PON1 organophosphate activity, the effect of bis-p-nitrophenyl phosphate (BNPP) and phenylmethylsulfonyl fluoride (PMSF), esterase inhibitors, and ethylenediaminetetraacetic acid (EDTA), a general paraoxonase inhibitor, was tested. Finally, the presence and activities of PONs in the A549 pulmonary cell line were also evaluated in order to be used as a model for studies on paraoxonases' metabolism. KEY FINDINGS: Our results demonstrated high interindividual variability in both PONs mRNA/protein levels and enzymatic activities and pointed out the presence of all PONs in human lungs and their subcellular distribution in the cytosol, microsomes, and mitochondria. SIGNIFICANCE: These findings add further information to our knowledge of pulmonary metabolism and, given that PON1 can metabolize some drugs used for respiratory diseases, the presence of PON1 activity in the lung tissue should no longer be ignored in the development of treatment plans and the design of new drugs.

Medical and surgical treatments for tinnitus: the efficacy of combined treatment with sulodexide and melatonin.

AIM: Tinnitus is the perception of sound in the absence of an apparent acoustic stimulus. A widespread and highly debilitating disease difficult to cure. Several treatments have been advocated for tinnitus in the last years, including surgery, pharmacotherapy, counseling, cognitive behavioral therapy, sound therapy, but unfortunately without definitive conclusions. The surgery treatments could represent an important therapeutic choice on specific subgroups of tinnitus with defined causes but obviously this approach represent an invasive treatment and it should be considered with extreme caution and then, alternative pharmacological options should be investigated. METHODS: In this retrospective study 30 patients with tinnitus were treated with sulodexide (250 LSU BID, in the morning and in the evening) and melatonin (3 mg in the evening before going to sleep) for 80 days. The evaluations were performed comparing different parameters at basal (T0) and after 40 days (T1) and 80 days (T2) of treatment. RESULTS: The results of Tinnitus Handicap Inventory (THI) and acufenometry showed a significative improvement of tinnitus after treatment with sulodexide and melatonin. In particular, THI total score was reduced from 37±20 to 27±18 (P<0.001) and 21±19 (P<0.001) at T1 and T2, respectively. The percentage of patients with improved symptoms (i.e. reduced score at THI) was 76.7% at T1 and 90.0% at T2. Finally a significant improvement was also detected in the tone audiometry test. No side effects were observed during the treatment period. CONCLUSION: In conclusion, the combined use of sulodexide, a natural glycosaminoglycan with antithrombotic, profibrinolytic and vascular anti-inflammatory properties used in the treatment of many vascular diseases, included the vertigo of vascular origin and melatonin, a neurohormone produced by the pineal gland and related to multiple physiological functions, confirms to an important and promising therapeutically option in the tinnitus management.

GEBR-7b, a novel PDE4D selective inhibitor that improves memory in rodents at non-emetic doses.

BACKGROUND AND PURPOSE: Strategies designed to enhance cerebral cAMP have been proposed as symptomatic treatments to counteract cognitive deficits. However, pharmacological therapies aimed at reducing PDE4, the main class of cAMP catabolizing enzymes in the brain, produce severe emetic side effects. We have recently synthesized a 3-cyclopentyloxy-4-methoxybenzaldehyde derivative, structurally related to rolipram, and endowed with selective PDE4D inhibitory activity. The aim of the present study was to investigate the effect of the new drug, namely GEBR-7b, on memory performance, nausea, hippocampal cAMP and amyloid-ß (Aß) levels. EXPERIMENTAL APPROACH: To measure memory performance, we performed object recognition tests on rats and mice treated with GEBR-7b or rolipram. The emetic potential of the drug, again compared with rolipram, was evaluated in rats using the taste reactivity test and in mice using the xylazine/ketamine anaesthesia test. Extracellular hippocampal cAMP was evaluated by intracerebral microdialysis in freely moving rats. Levels of soluble Aß peptides were measured in hippocampal tissues and cultured N2a cells by elisa. KEY RESULTS: GEBR-7b increased hippocampal cAMP, did not influence Aß levels and improved spatial, as well as object memory performance in the object recognition tests. The effect of GEBR-7b on memory was 3 to 10 times more potent than that of rolipram, and its effective doses had no effect on surrogate measures of emesis in rodents. CONCLUSION AND IMPLICATIONS: Our results demonstrate that GEBR-7b enhances memory functions at doses that do not cause emesis-like behaviour in rodents, thus offering a promising pharmacological perspective for the treatment of memory impairment.

Antifungal resistance in Candida spp. isolated in Italy between 2002 and 2005 from children and adults.

The activity of amphotericin B, fluconazole, flucytosine, itraconazole and voriconazole was tested in vitro against 618 clinical Candida spp. isolates, using the broth microdilution or the disk diffusion method (voriconazole). Amphotericin B and voriconazole were the most potent antifungal agents assayed (100% of susceptible strains). Resistance to fluconazole and itraconazole was detected in three (0.7%) and 11 (2.7%) isolates of Candida albicans and in four (3.7%) isolates of Candida glabrata. Flucytosine intermediate, resistant strains, or both, were observed in C. albicans (0.3% and 0.7%), C. glabrata (2.8% intermediate) and C. tropicalis (15.2% and 15.2%). C. krusei was the least susceptible species to azoles. No statistically significant differences in the rates of resistant isolates depending on site of infection and age of the patient were observed, with the exception of C. albicans and itraconazole (higher percentage of resistance in children). At present, acquired antifungal resistance represents an uncommon finding in most Candida spp. circulating in Northern Italy.

Nocardia keratitis: a case report.

PURPOSE: To describe a case of Nocardia keratitis resistant to 2% amikacin, with a toxic-allergic reaction to fortified topical 5% amikacin, and recurrence of the infection with topical corticosteroids. METHODS: Nocardia was diagnosed from a smear and positive culture and identified as Nocardia asteroides by gas chromatography and quantitative fatty acid analysis using the Microbial Identification System. Treatment was started with topical 2% amikacin, which was subsequently raised to 5% because of clinical resistance. RESULTS: A toxic-allergic reaction was observed after 5% amikacin so the drug was discontinued and commercially available drugs combining 1% chloramphenicol, 0.5% tetracycline, and 18 mil IU colistin with 0.3% ofloxacin were given. These were well tolerated and the infection improved quickly. After 1 month the antibiotics were discontinued and topical 0.1% clobetasone was given to reduce scar formation. The infection recurred after 1 week but responded to 3 months of the previous antibiotic combination and its sensitivity was checked with the Epsilometer test. CONCLUSIONS: Nocardia keratitis may not respond to 2% topical amikacin and fortified topical 5% amikacin may cause a strong toxic-allergic reaction. A commercially available combination of chloramphenicol, tetracycline, and colistin, with ofloxacin, may be effective but the treatment must be continued for several months. Topical steroids should only be used with considerable caution since they can lead to relapse of the infection.

Nocardia keratitis: A case report.

PURPOSE: To describe a case of Nocardia keratitis resistant to 2% amikacin, with a toxic-allergic reaction to fortified topical 5% amikacin, and recurrence of the infection with topical corticosteroids. METHODS: Nocardia was diagnosed from a smear and positive culture and identified as Nocardia asteroides by gas chromatography and quantitative fatty acid analysis using the Microbial Identification System. Treatment was started with topical 2% amikacin, which was subsequently raised to 5% because of clinical resistance. RESULTS: A toxic-allergic reaction was observed after 5% amikacin so the drug was discontinued and commercially available drugs combining 1% chloramphenicol, 0.5% tetracycline, and 18 mil IU colistin with 0.3% ofloxacin were given. These were well tolerated and the infection improved quickly. After 1 month the antibiotics were discontinued and topical 0.1% clobetasone was given to reduce scar formation. The infection recurred after 1 week but responded to 3 months of the previous antibiotic combination and its sensitivity was checked with the Epsilometer test. CONCLUSIONS: Nocardia keratitis may not respond to 2% topical amikacin and fortified topical 5% amikacin may cause a strong toxic-allergic reaction. A commercially available combination of chloramphenicol, tetracycline, and colistin, with ofloxacin, may be effective but the treatment must be continued for several months. Topical steroids should only be used with considerable caution since they can lead to relapse of the infection. (Eur J Ophthalmol 2006; 16: 164-7).

A new simple immunoassay for detecting Helicobacter pylori infection: antigen in stool specimens.

BACKGROUND/AIM: Several diagnostic tests are available for evaluating Helicobacter pylori (Hp) infection: histological examination, culture of gastric biopsy specimens, rapid urease test, urea breath test and serology. A recently marketed direct enzyme immunoassay (HpSA) detects Hp antigen in stool samples. The aim of our study was to evaluate overall diagnostic sensitivity, specificity and positive and negative predictive values of this new diagnostic test. METHODS: We included in the study 84 patients (39 males and 45 females; mean age 49.57 years) with dyspeptic symptoms who were examined by upper gastrointestinal endoscopy. Exclusion criteria were previous treatment with proton pump inhibitors, bismuth compounds or antibiotics. During the endoscopic examination biopsies were taken from antrum and corpus for Hp culture and histological examination, and stool specimens were submitted to the laboratory to be stored until the HpSA test. Hp was judged to be present when culture or histology and culture were positive. The (13)C-urea breath test was done only in culture-negative patients in whom either histology or immunoassay or both were positive. RESULTS: Hp was found in 55 patients by both culture and histology. Stool antigen has been detected in 54 of the 55 Hp-positive patients, giving a sensitivity of 98.2% and a negative predictive value of 96.4%. In 2 out of 29 patients HpSA gave a positive result, but the biopsy-based methods were negative, resulting in a low rate of false-positives, with 93.1% specificity and 96.4% positive predictive value; the (13)C-urea breath test confirmed these results as negative. CONCLUSION: Our results show that this new test is highly sensitive and specific for the detection of Hp infection, and it is satisfactorily reproducible.

Mycobacterium chelonae keratitis after excimer laser photorefractive keratectomy.

This is the first report of a severe case of Mycobacterium chelonae keratitis; it occurred in a 26-year-old man after he had undergone excimer laser photorefractive keratectomy for the correction of severe myopia, once the epithelium was already healed. The diagnosis was made by culture results and acid-fast staining of corneal scrapings. Topical ciprofloxacin sodium, 0.3 mg/mL, plus amikacin sodium, 10 mg/mL, and oral clarithromycin sodium led to remission of the ulceration after 3 months of therapy. Subsequent topical corticosteroid therapy led to complete visual recovery during 1 year of follow-up. There may be an increased risk of severe keratitis during the first postoperative months in eyes that have already undergone photorefractive keratectomy, due to the presence of some microepithelial defects symptomatically negative and not easily detectable by slit-lamp examination.

HTLV-II infection in Florida Indians.

A significantly increased prevalence of antibodies to human T-cell leukemia virus (HTLV) has been described in several native American populations in the United States and Latin America. Initial virologic studies indicate that HTLV-II is the predominant virus responsible for this antibody pattern. We obtained blood samples from 106 Seminole Indians living on four reservations in Southern Florida. Seropositivity to HTLV-I/II was found in 14 (13.2%) of these individuals. Polymerase chain reaction (PCR) documented HTLV-II and the absence of HTLV-I in 7 of the 9 donors available for follow-up testing of white blood cells. Evaluation of various risk factors excluded blood transfusion or intravenous drug use as an important contributing factor to the HTLV-II seroprevalence rate. These studies support the hypothesis that HTLV-II is endemic in many native American tribes in the Western hemisphere.

Intracellular pH regulates the production of different oxygen metabolites in neutrophils: effects of organic acids produced by anaerobic bacteria.

The effects of short chain carboxylic acids (SCCA), namely succinic, butyric, and iso-butyric, on neutrophil metabolic activation were assessed. SCCA induced a significant decrease in O2.- recovery and chemiluminescent response in neutrophils activated with the diacylglycerol analog tetradecanoyl-phorbol-acetate (TPA). SCCA did not alter O2 consumption, H2O2 production, or the calorimetrically determined energy expenditure occurring during the metabolic burst. SCCA also induced a significant acidification of intracellular pH (pHi). These results are consistent with an increased divalent versus univalent O2 reduction performed by the NADPH oxidase at a more acidic intracellular pH.

Suppression of tricarboxylic acid cycle in Escherichia coli exposed to sub-MICs of aminoglycosides.

The metabolic activity of Escherichia coli ATCC 25922 challenged with sub-MICs of aminoglycosides was analyzed with a batch calorimeter. High-performance and gas-liquid chromatographic techniques were utilized to evaluate the concentrations of metabolic reactants, intermediates, and end products. The data reported indicate that aminoglycosides inhibit or delay bacterial catabolism of carboxylic acids, with the following relative degrees of activity: amikacin greater than gentamicin greater than sisomicin greater than netilmicin greater than kanamycin. The decrease in total biomass production was proportional to the degree of tricarboxylic acid cycle inhibition.

Short-chain fatty acids produced by anaerobic bacteria inhibit phagocytosis by human lung phagocytes.

The effect of short-chain fatty acids on the phagocytic activity of human alveolar macrophages and neutrophils was investigated. These acids, butyric, propionic, and succinic, are produced by anaerobic bacteria. The results indicate that phagocytosis of Staphylococcus aureus by human lung phagocytes is strongly inhibited by the end products of anaerobic catabolism and support the hypothesis that the antiphagocytic activity present in the supernatants of anaerobic cultures may be dependent on the presence of short-chain fatty acids.

[Action of clindamycin on bacteria adhering to plastic materials]. / Attività della clindamicina su batteri adesi alle materie plastiche.

The coagulase negative staphylococci capable of adhering to solid surfaces have been recognised as aetiological agents of infections associated with the presence of plastic biomedical prostheses. The study of antibiotic activity by traditional in vitro methods has proved unpredictive when applied to this type of infection: we have therefore used a calorimetric technique. Like other antibiotics, clindamycin was not effective in eradicating colonisation of biomedical prostheses. Studies on the activity of the antibiotic used in the prophylaxis of these infections in likely to be more promising.

Post-antibiotic effect induced by ofloxacin in both gram-positive and gram-negative bacteria.

We estimated the relationship of Post Antibiotic Effect (PAE) induced by ofloxacin in both gram-positive and gram-negative bacteria, with the Percent Growth Rate Average (PGRA) considered on an increase of 1 log (CFUs/ml). The results showed a good correlation between parameters (0.84, p less than 0.001), and enabled us to find out that the drug-induced effect persisted after the period of time considered in the standard procedure. Where the values of PAE are much greater, the growth curve rates were faster after the apparent termination of the phenomenon.