Tonino Bucciarelli

Antileukotriene Drugs: Clinical Application, Effectiveness and Safety

Cysteinyl leukotrienes (Cys-LTs) are potent proinflammatory mediators derived from arachidonic acid through the 5-lypoxigenase (5-LO) pathway. They exert important pharmacological effects by interaction with at least two different receptors: Cys-LT(1) and Cys-LT(2). By competitive binding to the Cys-LT(1) receptor, leukotriene receptor antagonist drugs such as montelukast, zafirlukast, and pranlukast, block the effects of Cys-LTs and alleviate the symptoms of many chronic diseases, especially bronchial asthma and allergic rhinitis. Evidence obtained by randomized clinical trials as also by direct experience derived from patients suffering from asthma and allergic rhinitis justifies a broader role for leukotrienes receptor antagonists (LTRAs). Recently published studies and case reports have demonstrated beneficial effects of LTRAs on other diseases commonly associated with asthma (exercise induced asthma, rhinitis, chronic obstructive pulmonary disease, interstitial lung disease, chronic urticaria, atopic dermatitis, allergic fungal disease, nasal polyposis, and paranasal sinus disease) as well as other diseases not connected to asthma (migraine, respiratory syncytial virus postbronchiolitis, systemic mastocytosis, cystic fibrosis, pancreatitis, vulvovaginal candidiasis, cancer, atherosclerosis, eosinophils cystitis, otitis media, capsular contracture, and eosinophilic gastrointestinal disorders). The aim of this review is to show the most recent applications and effectiveness in clinical practice of the LTRAs.

DEVELOPMENTAL ASPECTS OF DETOXIFYING ENZYMES IN FISH (SALMO IRIDAEUS)

The activities of superoxide dismutase, catalase, glutathione reductase, glutathione peroxidase, glutathione transferase and glyoxalase I have been studied during the embryologic development of rainbow trout (Salmo iridaeus) and in several other trout tissues to investigate the protective development metabolism. A gradual increase of superoxide dismutase, catalase, glutathione reductase, glyoxalase I and glutathione transferase activities was noted throughout embryo development. In all trout tissues investigated glutathione peroxidase was found to be extremely low compared to catalase activity. The highest activity of superoxide dismutase, glyoxalase I and glutathione reductase was found in liver followed by kidney. No change in the number of GST subunits was noted with the transition from the embryonic to the adult stages of life according to the SDS/PAGE and HPLC analyses performed on the GSH-affinity purified fractions.

Determinants of platelet activation in Alzheimer's disease.

OBJECTIVES To investigate the rate of platelet thromboxane (TX) biosynthesis and its determinants in Alzheimers disease. METHODS AND RESULTS A cross-sectional comparison of urinary 11-dehydro-TXB(2) and 8-iso-prostaglandin (PG)F(2alpha) (markers of in vivo platelet activation and lipid peroxidation, respectively), plasma Vitamin E, C-reactive protein (CRP), tumor necrosis factor (TNF)-alpha and interleukin (IL)-6, was carried-out in 44 Alzheimer patients and 44 matched controls. To investigate the cyclooxygenase (COX)-isoform involved in TXA(2) biosynthesis, nine Alzheimer patients were treated with low-dose aspirin (100mg/d) or rofecoxib (25mg/d) for 4 days. Urinary 11-dehydro-TXB(2) and 8-iso-PGF(2alpha) were significantly higher in Alzheimer patients than in controls (Median: 1983.5 versus 517.5pg/mg creatinine and 938.5 versus 304.0pg/mg creatinine, p<0.0001, respectively), with a significant correlation between the two metabolites (rho=0.75, p<0.0001). An inverse correlation was observed between Vitamin E and both urinary metabolites (8-iso-PGF(2alpha): R(s)=-0.51, p=0.0004; 11-dehydro-TXB(2): R(s)=-0.44, p=0.0026) in Alzheimer patients. No difference was found in CRP, TNF-alpha and IL-6 levels between the two groups. Urinary 11-dehydro-TXB(2) was significantly reduced by aspirin, but not by rofecoxib, consistently with a COX-1-mediated TXA(2) biosynthesis. 8-iso-PGF(2alpha) excretion was not modified by either COX-inhibitor, consistently with its oxygen radical-catalyzed formation. CONCLUSIONS Platelet activation is persistently enhanced in Alzheimers disease. This is related, at least in part, to increased lipid peroxidation associated with inadequate levels of Vitamin E.

Characterization of toad liver glutathione transferase

The major form of glutathione transferase from the toad liver previously designed as Bufo bufo liver GST-7.6 (A. Aceto, B. Dragani, T. Bucciarelli, P. Sacchetta, F. Martini, S. Angelucci, F. Amicarelli, M. Miranda and C. Di Ilio, Biochem. J. 289 (1993) 417-422) has been characterized. According to its partial amino acid sequence, the toad enzyme may be included in the pi class GST and named bbGST P2-2. However, bbGST P2-2 appears to be immunologically, structurally and kinetically distinct from any other members of pi family, including bbGST P1-1, suggesting that it may constitute a subset of pi class GST. The data support the hypothesis that the transition from aquatic to terrestrial life causes a switch of the GST amphibian pattern promoting the expression of a GST form (bbGST P2-2) able to counteract, with higher efficiency, the toxic effects of reactive metabolites of oxidative metabolism and those of hydrophobic xenobiotics.

Developmental aspects of Bufo bufo embryo glutathione transferases

The expression of glutathione transferase isoenzymes has been studied during the development of Bufo bufo embryo. By analysing the GSH-affinity purified materials in terms of substrate specificities, SDS-PAGE pattern, HPLC elution profile, we conclude that, up to stage 22, no significant changes in the expression of glutathione transferases isoenzymes occurred during Bufo bufo embryo development. At stage 25 the distribution of glutathione transferases was found to be slightly different from those of all other foregoing stages. A marked decrease of embryonic glutathione transferases subunits with a parallel appearance of new structurally and immunologically different subunits was noted in toad liver and kidney. Toad ovary continued to express embryonic glutathione transferase subunits.

CIRCULATING PLASMA ANTIOXIDANTS, INFLAMMATORY MARKERS AND ASYMPTOMATIC CAROTID ATHEROSCLEROSIS IN END-STAGE RENAL DISEASE PATIENTS: A CASE CONTROL STUDY

Few studies have been conducted on the relationship between antioxidant plasma vitamin concentrations, inflammatory markers and carotid atherosclerosis with inconclusive results in end-stage renal disease (ESRD) patients. A case-control study was performed to investigate the relationship between plasma antioxidant concentrations, inflammatory markers, and carotid intima-media thickness (CIMT) in healthy subjects and in patients undergoing hemodialysis (HD). We enrolled 40 subjects (20 healthy, 20 with ESRD) asymptomatic for carotid atherosclerosis. After carotid ultrasound investigation (CUI), medical history data, physical examination, venous blood samples were collected. These were analyzed for concentrations of antioxidant vitamins (A, E), carotenoids (lycopene, β-carotene), inflammatory markers (C-reactive protein, fibrinogen), and lipid profile. Low concentrations of vitamin A, vitamin E, lycopene, and β-carotene were significantly associated with carotid atherosclerosis in patients with ESRD (p<0.001). In addition, high concentration of low density lipoprotein cholesterol and total cholesterol (p<0.01), C-reactive protein and fibrinogen (p<0.001) were also associated with carotid atherosclerosis, while other laboratory parameters considered (high density lipoprotein cholesterol and triglycerides) were not significantly associated with carotid atherosclerosis. A regular intake of foods rich in antioxidant vitamins with low fat concentrations may slow the progression of atherosclerotic process in this group of patients.

Calcium Binding Promotes Prion Protein Fragment 90–231 Conformational Change toward a Membrane Destabilizing and Cytotoxic Structure

The pathological form of prion protein (PrPSc), as other amyloidogenic proteins, causes a marked increase of membrane permeability. PrPSc extracted from infected Syrian hamster brains induces a considerable change in membrane ionic conductance, although the contribution of this interaction to the molecular mechanism of neurodegeneration process is still controversial. We previously showed that the human PrP fragment 90–231 (hPrP90–231) increases ionic conductance across artificial lipid bilayer, in a calcium-dependent manner, producing an alteration similar to that observed for PrPSc. In the present study we demonstrate that hPrP90–231, pre-incubated with 10 mM Ca++ and then re-suspended in physiological external solution increases not only membrane conductance but neurotoxicity as well. Furthermore we show the existence of a direct link between these two effects as demonstrated by a highly statistically significant correlation in several experimental conditions. A similar correlation between increased membrane conductance and cell degeneration has been observed assaying hPrP90–231 bearing pathogenic mutations (D202N and E200K). We also report that Ca++ binding to hPrP90–231 induces a conformational change based on an alteration of secondary structure characterized by loss of alpha-helix content causing hydrophobic amino acid exposure and proteinase K resistance. These features, either acquired after controlled thermal denaturation or induced by D202N and E200K mutations were previously identified as responsible for hPrP90–231 cytotoxicity. Finally, by in silico structural analysis, we propose that Ca++ binding to hPrP90–231 modifies amino acid orientation, in the same way induced by E200K mutation, thus suggesting a pathway for the structural alterations responsible of PrP neurotoxicity.

Analysis by limited proteolysis of domain organization and GSH-site arrangement of bacterial glutathione transferase B1-1

Limited proteolysis method has been used to study the structure-function relationship of bacterial glutathione transferase (GSTB1-1). In absence of three-dimensional structural data of prokaryote GST, the results represent the first information concerning the G-site and domains organization of GSTB1-1. The tryptic cleavages occur mainly at the peptide bonds Lys35-Lys36 and Phe43-Leu44, generating two major molecular species of 20-kDa, 3-kDa and traces of 10-kDa. 1-chloro-2,4-dinitrobenzene favoured the proteolysis of the 20-kDa fragment markedly enhancing the production of the 10-kDa peptide by cleaving the chemical bonds Lys87-Ala88 and Arg91-Tyr92. The tryptic cleavage sites of GSTB1-1 was found to be located close to those previously found for the mammalian GSTP1-1 isozyme. It was concluded that despite their low sequence homology (18%), GSTB1-1 and GSTP1-1 displayed similar structural features in their G-site regions and probably a common organization in structural domains.

Effect of 2-year treatment with low-dose rosuvastatin on intima-media thickness in hypercholesterolemic subjects with asymptomatic carotid artery disease

Objectives: Recent evidence indicates that rosuvastatin 40 mg may exert a beneficial effect in both carotid and coronary atherosclerosis progression. In particular, 2-year rosuvastatin treatment reduced the progression of carotid intima-media thickness (cIMT) in patients with low cardiovascular risk. However, despite the fact that in clinical practice lower doses of rosuvastatin are usually administered at this time, there are no clear data about its effect on cIMT. Thus, the aim of this study was to evaluate the effect of rosuvastatin 10 mg/day on cIMT over a 2-year follow-up. Methods: Forty-five patients with hypercholesterolemia and asymptomatic carotid atherosclerosis on baseline carotid ultrasound investigation were treated with rosuvastatin 10 mg/day for 24 months. cIMT and lipid profile were assessed after 12 months and at the end of the study (24 months). Results: After 24 months, the treatment showed a 35.67% reduction in low-density lipoprotein cholesterol concentration (171 vs 110 mg/dl; p < 0.001), a 32.27% reduction in total cholesterol (251 vs 170 mg/dl; p < 0.001), a 19.67% increase in high-density lipoprotein cholesterol concentration (49 vs 61 mg/dl; p < 0.001), and a 10% reduction in triglycerides (120 vs 108 mg/dl; p < 0.01). Rosuvastatin treatment was associated with a 26.6% reduction in left cIMT (1.20 vs 0.90 mm; p < 0.001) and a 22.2% reduction in right cIMT (1.22 vs 0.95 mm; p < 0.001). Conclusion: Two-year treatment with rosuvastatin 10 mg/day in hypercholesterolemic adults with evidence of subclinical atherosclerosis establishes a significant reduction in cIMT and improves lipid and lipoprotein levels, with a good tolerability profile.

Dilated cardiomyopathy following use of xenadrine EFX.

We describe a case of a 35-year-old man presented at the emergency room of our institution with acute onset of dyspnea and dizziness. He was a body builder and had been using Xenadrine EFX for weight loss reduction. The laboratory analyses were normal. A chest radiograph showed an enlarged cardiac silhouette with clear lung fields. Transtoracic two-dimensional color Doppler echocardiography revealed a diffuse hypokinesia with a marked decreased in systolic function and a high teledyastolic diameter. This case document the possible relation to use of Xenadrine EFX for weight loss and the recurrence of dilated cardiomyopathy.