Diet macronutrient composition reported before treatment predicts BMI change in obese children: the role of lipids.

In this study, we tested the hypothesis that diet composition reported by children before the beginning of an obesity treatment program could be a predicting factor of the clinical outcome. A sample of 138 obese 6-16-year-old children and adolescents were recruited. Anthropometry and dietary habits were recorded. Each patient participated in a multidimensional treatment program in an outpatient obesity public service clinic. Therapy was based on a 6-month educational program on nutrition, lifestyle and physical activity. Children with a lipid intake above 34.7% of total energy had a 2.5 times higher chance of reducing at least 1.5 units of BMI with treatment than children with lower lipid intake. These results suggest that the assessment of habitual diet, in particular diet composition before starting treatment, may help to identify obese children who are more sensitive to intervention and those who need more specific nutritional assistance.

A paraneoplastic retroperitoneal fibrosis resistant to corticosteroids treated with tamoxifen.

Retroperitoneal fibrosis (RPF) is a rare disease characterized by an inflammatory proliferative fibrosing process occurring in the retroperitoneum, often causing urinary tract obstruction. Medical therapy is not well-defined, but glucocorticoids have been the mainstay of therapy. Recently, positive response to tamoxifen, an antiestrogen drug, has been reported among patients with RPF. We report the case of a 65-year-old male with a renal cell carcinoma in the upper pole of the right kidney showing acute renal failure due to a biopsy-confirmed RPF determining bilateral hydronephrosis. After polar resection of the right kidney, a high-dose oral steroid therapy did not modify the hydronephrosis. At 6 months, therapy with tamoxifen determined the retroperitoneal fibrotic mass regression and resolved the ureteral obstruction, that persists at the 13th month of follow-up. Tamoxifen can be considered as an effective alternative to corticosteroids and immunosuppressors in treating RPF.

Hypokalemic nephropathy in an adult patient with partial empty sella: a classic Bartter's syndrome, a Gitelman's syndrome or both?

Bartter's syndrome belongs to a group of hypokalemic renal channel diseases. These channels are located in the lipid layer of cell membranes where they exist as water channels through which ion transport is performed. Based on the type of genetic disorder and clinical presentation, Bartter's syndrome is classified as neonatal, classical and Gitelman's syndrome. Most of the cases have been noted in pediatric age groups and adult-onset cases are very rare. Moreover, an association between Bartter's syndrome and empty sella has recently been reported in 3 children. We report here the second case of an adult patient affected by Bartter's syndrome with partial empty sella. The patient showed some clinical and histological characteristics of both classic Bartter's syndrome and Gitelman's syndrome, suggesting that genotype and phenotype of Bartter's syndrome are not so clear-cut and that phenotypic overlap may occur, according to a recent hypothesis. Magnetic resonance imaging disclosed a partial empty sella. A thorough endocrinological investigation showed normal hypophyseal, thyroidal, adrenal and gonadal function. Good therapeutic effects were achieved using spironolactone, ACE-inhibitor and potassium supplementation, with normalization of the kalemia. At present, the value of the association of Bartter's syndrome and empty sella remains unclear and future studies are needed to clarify the importance of this association, both in children and in adult patients affected by Bartter's syndrome.

Ring opening reactions: synthesis of AICAR analogs as potential antimetabolite agents.

In an attempt to improve the AzA selectivity of the 2-(aryl)alkylthio derivatives of adenosine, we planned the synthesis of the corresponding derivatives of the 5-N-ethylcarboxamidoadenosine (NECA). For this purpose, we designed the synthesis of 2-mercapto-NECA to be pursued by means of an opening-closure method We obtained the open AICAR analog; however, ring closure efforts failed to give the desired compound. The newly synthesized AICAR derivative could potentially be endowed with antiviral or antitumoral activity.

A2B adenosine receptor agonists: synthesis and biological evaluation of 2-phenylhydroxypropynyl adenosine and NECA derivatives.

In the search for agonists for the elusive A2B adenosine receptor subtypes, 2-phenylhydroxypropynyl-5'-N-methylcarboxamido adenosine (PHPMECA, 14), 2-phenylhydroxypropynyl-5'-N-propylcarboxamido adenosine (PHPPECA, 15), and N6-ethyl-2-phenylhydroxypropynyl-5'-N-ethylcarboxamidoadenosine (19) were synthesized on the basis that introduction of alkynyl chains in 2-position of adenosine derivatives resulted in reasonably good A2B potency compared to NECA [see N6-ethyl-2-phenylhydroxypropynyl adenosine (5) EC50 = 1,700 nM and 2-phenylhydroxypropynyl-5'-N-ethylcarboxamido adenosine (PHPNECA, 8) EC50 = 1,100 nM, respectively]. Radioligand binding studies and adenylyl cyclase assays, performed with recently cloned human A1, A2A, A2B, and A3 adenosine receptors, showed that these modifications produced a decrease in potency at A2B receptor, as well as a general reduction in affinity at the other receptor subtypes. On the other hand, the contemporary presence of an ethyl substituent in N6-position and of a 4'-ethylcarboxamido group in the same compounds led to (R,S)-N6-ethyl-2-phenylhydroxypropynyl-5'-N-ethylcarboxamidoadenosine and (S)-N6-ethyl-2-phenylhydroxypropynyl-5'-N-ethylcarboxamidoadenosine, which did not show the expected increase in potency at A2B subtype. Hence, (S)-2-phenylhydroxypropynyl-5'-N-ethylcarboxamidoadenosine [(S)-PHPNECA] with EC50 A2B = 220 nM remains the most potent agonist at A2B receptor reported so far.

2-Phenylhydroxypropynyladenosine derivatives as high potent agonists at A2B adenosine receptor subtype.

Adenosine derivatives bearing in 2-position the (R,S)-phenylhydroxypropynyl chain were evaluated for their potency at human A2B adenosine receptor, stably transfected on CHO cells, on the basis that (R,S)-2-phenylhydroxy-propynyl-5'-N-ethylcarboxyamidoadenosine [(R,S)-PHPNECA] was found to be a good agonist at the A2B receptor subtype. Biological studies demonstrated that the presence of small alkyl groups in N6-position of these molecules are well tolerated, whereas large groups abolished A2B potency. On the other hand, the presence of an ethyl group in the 4'-carboxamido function seems to be optimal, the (S)-PHPNECA resulting the most potent agonist at A2B receptor reported so far.

Human cytidine deaminase: understanding the catalytic mechanism.

In the absence of an experimentally elucidated three-dimensional structure of the human CDA, we built an homology model of this enzyme starting from the crystal structure of its E. coli homologous. Furthermore, we docked in the active site alternatively the substrate, the intermediate or the product. By means of molecular dynamics simulations, we determined the topology of the active site, identifying the amino acids involved in the catalytic mechanism, and outlining the central role played by E67.

Intersubunit interactions in human cytidine deaminase.

In order to design new efficient cytidine based drugs, an intersubunit interactions study related to the active site has been performed on the wild-type cytidine deaminase (CDA) and on the mutant enzyme F137W/W113F. F137 is the homologous to the Bacillus subtilis CDA F125 involved in the subunit interactions. In presence of the dissociating agent SDS, wild-type human CDA dissociate into enzymatically inactive monomers without intermediate forms via a non-cooperative transition. Extensive dialysis or dilution of the inactivated monomers restores completely the activity. The presence of the strong human CDA competitive inhibitor 5-fluorozebularine disfavour dissociation of the tetramer into subunits in the wild-type CDA but not in mutant enzyme F137W/W113F.

Plasmaperfusion on triptophan columns can improve the clinical outcome of patients affected with myasthenia gravis.

UNLABELLED: Myasthenia Gravis (MG) is a neuromuscular disease often associated with thymic pathology due to neuromuscular transmission impairment by circulating antibodies directed against the cholinergic postsynaptic receptor on the neuromuscular junction (Anti-AchR-Ab). The treatment of MG includes cholinesterase inhibitors, steroids and thymectomy. Plasmapheresis can remove Anti-AchR-Ab but more recently plasma-perfusion (PP), a more specific apheresis for selective removal of noxious plasma components, has been developed. AIM OF THE STUDY: To study the effect of PP treatment, performed by using specific immunocolumns for Anti-AchR-Ab, on the clinical outcome of MG patients non-responder to steroid therapy or thymectomy. MATERIALS AND METHODS: We treated 8 patients suffering from severe MG by a cycle of 6 sessions of PP. We used columns containing triptophan as a specific ligand for Anti-AchR-Ab. In order to evaluate the effectiveness of treatment we used functional tests (muscular tests, respiratory function, electromyography) and laboratory tests (Anti-AchR-Ab; immunoglobulins, complement fractions, immunocomplexes). RESULTS: After one to three PP sessions, early clinical improvement in bulbar and respiratory symptoms were found in all patients and EMG showed improvement of neuromuscular transmission. Serum concentration of immunological markers decreased progressively and significantly during the treatment. Clinical improvements were progressive despite the tendency for Anti-AchR-Ab to reach initial values between one session and another. We observed no side effects due to the type of immunocolumns used. CONCLUSIONS: Triptophan columns appear to be able to remove large quantities of Anti-AchR-Ab and immunological markers from plasma. Our experience shows that PP performed using triptophan columns in patients suffering from severe MG provides good clinical results, improving patients' outcome, without any risk linked to the procedure.

Resistive index in chronic nephropathies: predictive value of renal outcome.

UNLABELLED: The study of renovascular resistances by color Doppler ultrasound has become a useful diagnostic resource for nephrologists. In recent nephrological literature, many papers deal with the correlations between resistive index, anatomo-pathological patterns and renal function. AIMS: In our study, we have tried to discover if resistive index represents a prognostic index of progressive renal failure. MATERIAL AND METHODS: To this purpose we compared renal resistive index and blood creatinine obtained from 28 nephropathic patients at their first control, with blood creatinine values after a 3-year follow-up period. Using a linear regression test, we found a strong correlation between the initial value of resistive index and the value of creatinine variation (p = 0.006). RESULTS: All of the patients with normal resistive index at the beginning maintained a stable renal function. Conversely, the patients with high resistive index at their first control showed a progressive renal failure. CONCLUSION: Our study shows the reliability of resistive index in the prognostic evaluation of renal outcome.

Sodium and fluid modulation in dialysis: new approach.

Hypotension during hemodialysis is still an unsolved problem. The treatment of patients with cardiovascular instability is efficaciously carried out with the use of 'profiled dialysis' (PD) with computerized modulation of ultrafiltration and conductivity. We tested a new profile model which involves progressive decrease of ultrafiltration associated with variable conductivity ('bell pattern'). We observed 8 stable long-term patients receiving hemodialysis (4 men and 4 women, mean age 63.5 years) for 4 h three times a week. Before our test, sodium balance had reached a steady state in all patients and remained stable during the entire observation period. The sodium balance was established by means of a simple pattern suggested by Ursino and coworkers. The patients were observed for two periods of 1 month each (protocols A and B). The intradialytic mean arterial pressure was studied, checking every hour of dialysis. Statistical analysis was done by ANOVA for repeated measures. We compared standard dialysis with constant ultrafiltration rate and conductivity (protocol A) with sessions performed involving a progressive decrease of ultrafiltration together with a variable conductivity of -0.2, +0.2, +0.6, +0.6, 0, -0.4, -0.4, and -0.4 mS/s (protocol B). We found a lower incidence of hypotension (p < 0.01) with better cardiovascular stability during and after treatment in 'profiled dialysis'.

3'-deoxyribofuranose derivatives of 1-deaza and 3-deaza-adenosine and their activity as adenosine deaminase inhibitors.

2,6-Dichloro-1-deazapurine and 2,6-dichloro-3-deazapurine were coupled with 1,2-O-diacetyl-5-O-benzoyl-3-deoxy-beta-D-ribofuranose. Deprotection of the obtained compounds and reaction with liquid ammonia gave the desired 2-chloroadenine nucleosides, which were dechlorinated to afford the corresponding 1-deaza and 3-deazaadenosine derivatives. Biological studies performed on ADA from calf intestine showed that these new nucleosides are inhibitors of the enzyme.

Synthesis and adenosine receptor affinity and potency of 8-alkynyl derivatives of adenosine.

Adenosine derivatives bearing different (ar)alkynyl chains at the 8-position were synthesized and tested at human adenosine receptors. Binding studies showed that all compounds possess affinity for the A3 subtype in the high nM range. Moreover, guanosine 5'-O-(3-[35S]thio)triphosphate binding assay indicated that the 8-alkynyl adenosines behaved as antagonists of NECA at A3 receptors.

Coupling of 2,6-disubstituted purines to ribose-modified sugars.

1,2,3-Tri-O-acetyl-N-ethyl-beta-D-ribofuranuronamide was synthesized in three steps starting from 1-O-methyl-(2,3-O-isopropylidene)-beta-D-ribofuranuronic acid. Both the triacetyl and the 1-O-methyl-2,3-di-O-acetyl derivatives were coupled to the 2,6-dichloropurine to obtain the acetylated 1-(2,6-dichloro-9H- purin-9-yl)-1-deoxy-N-ethyl-beta-D-erythro-pentofuranuronamide. 1H NMR and n.O.e. data accounted for both anomeric and N-7/N-9 isomeric configuration.

Synthetic procedure for the preparation of novel potent and selective A3 adenosine receptor radioligands.

2-Phenylethynyladenosine and its N6-methyl derivative were synthesized and evaluated in binding assays at human adenosine receptors stably transfected on CHO cells. Results showed that the N6-methyl-2-phenylethynyladenosine is endowed with very high affinity and selectivity at A3 receptor subtype. Hence, an alternative procedure for the synthesis of tritiated N6-methyl-2-phenylethynyladenosine was set up to introduce tritiated methylamine in the final step.

Introduction of alkynyl chains on C-8 of adenosine led to very selective antagonists of the A(3) adenosine receptor.

Some 8-alkynyladenosines were synthesized and evaluated for their adenosine receptor activity, utilizing radioligand binding studies (A(1), A(2A), A(3)) or adenylyl cyclase activity assays (A(2B)). Furthermore, the maximal induction of guanosine 5'-(gamma-thio)triphosphate ([35S]GTPgammaS) binding to G proteins and the inhibition of NECA-stimulated binding, in membranes of CHO cells which express the human A(3) receptor, were used to determine the intrinsic activity of these nucleosides at the A(3) adenosine receptor. The results showed that these new adenosine derivatives are very selective ligands for the A(3) receptor subtype and behave as adenosine antagonists, since they do not stimulate basal [35S]GTPgammaS binding, but inhibit NECA-stimulated binding. This is the first report that adenosine derivatives, with unmodified ribose moiety, are adenosine receptor antagonists.

Adenosine deaminase: functional implications and different classes of inhibitors.

Adenosine deaminase (ADA) is an enzyme of the purine metabolism which catalyzes the irreversible deamination of adenosine and deoxyadenosine to inosine and deoxyinosine, respectively. This ubiquitous enzyme has been found in a wide variety of microorganisms, plants, and invertebrates. In addition, it is present in all mammalian cells that play a central role in the differentiation and maturation of the lymphoid system. However, despite a number of studies performed to date, the physiological role played by ADA in the different tissues is not clear. Inherited ADA deficiency causes severe combined immunodeficiency disease (ADA-SCID), in which both B-cell and T-cell development is impaired. ADA-SCID has been the first disorder to be treated by gene therapy, using polyethylene glycol-modified bovine ADA (PEG-ADA). Conversely, there are several diseases in which the level of ADA is above normal. A number of ADA inhibitors have been designed and synthesized, classified as ground-state and transition-state inhibitors. They may be used to mimic the genetic deficiency of the enzyme, in lymphoproliferative disorders or immunosuppressive therapy (i.e., in graft rejection), to potentiate the effect of antileukemic or antiviral nucleosides, and, together with adenosine kinase, to reduce breakdown of adenosine in inflammation, hypertension, and ischemic injury.

N-cycloalkyl derivatives of adenosine and 1-deazaadenosine as agonists and partial agonists of the A(1) adenosine receptor.

A number of cycloalkyl substituents (from C-3 to C-8) have been introduced on the 6-amino group of adenosine, 1-deazaadenosine, and 2'-deoxyadenosine, bearing or not a chlorine atom at the 2-position, to evaluate the influence on the A(1) and A(2A) affinity of steric hindrance and lipophilicity. Furthermore, the guanosine 5'-triphosphate (GTP) shift and the maximal induction of guanosine 5'-(gamma-thio)triphosphate ([(35)S]GTPgammaS) binding to G proteins in rat brain membranes were used to determine the intrinsic activity of these nucleosides at the A(1) adenosine receptor. All compounds of the ribose-bearing series proved to be full agonists, the 1-deaza derivatives showing affinities for the A(1) receptor about 10-fold lower than the corresponding adenosines. On the other hand, all the 2'-deoxyribose derivatives bind to the A(1) receptor with affinities in the high nanomolar range, with the 2-chloro substituted compounds showing slightly higher affinities than the 2-unsubstituted counterparts. In terms of the potencies, the most potent compounds proved to be those bearing four- and five-membered rings. Both GTP shifts and [(35)S]-GTPgammaS experiments showed that most of the 2'-deoxyadenosine derivatives are partial agonists. The 2'-deoxyadenosine derivatives which were identified as partial agonists consistently detected fewer A(1) receptors in the high-affinity state than full agonists. However, it is worthwhile noting that there was not a simple linear relationship between receptor occupancy and activation. These results indicate that a critical density of A(1) adenosine receptors in the high-affinity state is required for G protein activation.

Synthesis and receptor affinity of polysubstituted adenosines.

In a search for potent and selective adenosine agonists it has been found that 2-hexynyladenosine-5'-N-ethyluronamide (HENECA) displays high affinity at rat A2A receptor combined with a good A2A vs A1 selectivity. The finding that HENECA shows good affinity also for A3 receptors prompted us to investigate the effect of various substituents in different positions of this molecule.

Synthesis of di- and tri-substituted adenosine derivatives and their affinities at human adenosine receptor subtypes.

The synthesis of 2-(hex-1-ynyl)adenosine derivatives substituted at the N6- and/or 5'-position was carried out on the basis that 2-(hex-1-ynyl)adenosine-5'-N-ethyluronamide (HENECA, 2) showed good affinity and different degree of selectivity for rat adenosine receptors. All new compounds were tested in radioligand binding and adenylyl cyclase assays with recently cloned human A1, A2A, A2B, and A3 adenosine receptors.