Purification and characterization of a phospholipase A2 isoenzyme isolated from Lachesis muta snake venom

A new phospholipase A2 (PLA2) isoenzyme was isolated from Lachesis muta crude venom, and was named LM-PLA2-II. This enzyme was purified by gel filtration on a Sephacryl S-200 HR column followed by reverse-phase chromatography on a C2/C18 column. LM-PLA2-II consists of a single polypeptide chain with an apparent molecular mass of 18 kDa and an isoelectric point at pH 5.4. The amino terminal sequence of the enzyme revealed a high degree of homology with other PLA2s from several sources. LM-PLA2-II has a high indirect hemolytic activity and a potent inhibitory effect on platelet aggregation induced by ADP and collagen. It also produces a significant paw edema reaction in rats. The edematous response in rats was abolished by pretreatment with either indomethacin or dexamethasone, suggesting the involvement of cyclo-oxygenase. Pretreatment of LM-PLA2-II with p-bromophenacyl bromide abolished all of these actions, clearly indicating that the biological activities, including the edematogenic effect, are dependent entirely on its enzymatic activity.     

Modulation of dendritic release of dopamine by metabotropic glutamate receptors in rat substantia nigra

A superfusion system was used to study the effects of metabotropic glutamate receptor (mGluR) ligands upon the release of [(3)H]dopamine ([(3)H]DA) previously taken up by rat substantia nigra (SN) slices. trans-(+/-)-1-Amino-(1S,3R)-cyclopentane dicarboxylic acid (trans-ACPD; 100 and 600 microM), a group I and II mGluR agonist, evoked the release of [(3)H]DA from nigral slices. This last effect was reduced significantly by (2S,3S,4S)-2-methyl-2-(carboxycyclopropyl)-glycine (MCCG; 300 microM), an antagonist of group II mGluR, or by the addition of tetrodotoxin (D-APV; 1 microM) to the superfusion medium. D-(-)-2-Amino-5-phosphono-valeric acid (100 microM), an N-methyl-D-aspartate receptor antagonist, or the presence of Mg(2+) (1.2mM) in the superfusion medium did not modify trans-ACPD-induced [(3)H]DA release. In addition, a group II mGluR agonist such as (2S,1'R,2'R,3'R)-2-(2',3'-dicarboxycyclopropyl)-glycine (DCG-IV; 100 microM) significantly induced the release of [(3)H]DA from nigral slices, whereas a group I mGluR agonist such as (RS)-3,5-dihydroxyphenylglycine (DHPG; 50 and 100 microM) did not modify the release of the [(3)H]-amine. Further experiments showed that the NMDA (100 microM)-evoked release of [(3)H]DA was decreased significantly by prior exposure of SN slices to trans-ACPD. Finally, partial denervation of the DA nigro-striatal pathway with 6-hydroxydopamine (6-OH-DA) increased trans-ACPD-induced release of [(3)H]DA, whereas it decreased trans-ACPD inhibitory effects on NMDA-evoked release of [(3)H]DA from nigral slices. The present results suggest that the dendritic release of DA in the SN is regulated by mGluR activation. Such nigral mGluR activation may produce opposite effects upon basal and NMDA-evoked release of DA in the SN. In addition, such mGluR-induced effects in the SN are modified in response to partial denervation of the DA nigro-striatal pathway.     

Differential activation of protein kinase C isoforms by euxanthone,revealed by an in vivo yeast phenotypic assay

The protein kinase C (PKC) modulatory effects of euxanthone, isolated from the wood of Cratoxylum maingayi, on isoforms alpha, betaI, delta, eta and zeta were characterised using an alternative in vivo yeast phenotypic assay. The present study shows that euxanthone can activate isoforms alpha, betaI, delta, eta and zeta, being more effective on PKC-betaI, -delta, -eta and -zeta than the established PKC activators used (the phorbol ester PMA and arachidonic acid for PKC-zeta). Furthermore, euxanthone presents differences on its potency towards individual PKC isoforms, showing a remarkable selectivity for PKC-zeta. These results can help to clarify the molecular basis of the euxanthone-mediated effects.     

Amyloidosis and Alzheimer's disease

Alzheimer's disease (AD) is the most frequent type of amyloidosis in humans and the commonest form of dementia. Extracellular Abeta amyloid deposits in the form of amyloid plaques and cerebral amyloid angiopathy as well as intraneuronal neurofibrillary tangles co-exist in the brain parenchyma of AD patients, the cognitive areas being the most severely affected. This review focuses on the potential role of amyloid in the development of neurodegeneration and presents studies of AD and other unrelated inherited dementia syndromes associated with neuronal loss and amyloid deposition in the brain.     

Naltrexone improves outcome of a controlled drinking program

Naltrexone is widely used in therapeutic programs with abstinence as a goal. However, it has been used in only a few studies aimed at reducing alcohol consumption. The purpose of this study was to evaluate the efficacy of naltrexone as an adjunct in controlled drinking programs. This was an open randomized study of 12 weeks duration that compared two therapeutic strategies: use of naltrexone in a controlled drinking program (NTX+CD) and the controlled drinking program alone (CD), without NTX. Each group comprised 30 male patients with mild alcohol dependence. During treatment, there were no differences between groups in drinking behavior, though the NTX+CD group showed significantly less craving. In the 12-month follow-up period, the NTX+CD group showed significantly fewer drinking days and heavy drinking days and less craving than the CD group. The results of this study suggest a role for naltrexone in controlled drinking programs.     

Characterization of flagella produced by clinical strains of Stenotrophomonas maltophilia

Stenotrophomonas maltophilia is an emerging nosocomial pathogen associated with opportunistic infections in patients with cystic fibrosis, cancer, and HIV. Adherence of this organism to abiotic surfaces such as medical implants and catheters represents a major risk for hospitalized patients. The adhesive surface factors involved in adherence of these bacteria are largely unknown, and their flagella have not yet been characterized biochemically and antigenically. We purified and characterized the flagella produced by S. maltophilia clinical strains. The flagella filaments are composed of a 38-kDa subunit, SM(FliC), and analysis of its N-terminal amino acid sequence showed considerable sequence identity to the flagellins of Serratia marcescens (78.6%), Escherichia coli, Proteus mirabilis, Shigella sonnei (71.4%), and Pseudomonas aeruginosa (57.2%). Ultrastructural analysis by scanning electron microscopy of bacteria adhering to plastic showed flagellalike structures within the bacterial clusters, suggesting that flagella are produced as the bacteria spread on the abiotic surface.     

Nutritional aspects of liver transplantation

Most adult and pediatric liver transplantation candidates present several metabolic disturbances that lead to malnutrition. Because malnutrition may adversely affect morbidity and mortality of orthotopic liver transplantation, it is very important to carefully assess the nutritional status of the waiting list patients. Pretransplant nutritional therapy -- enteral or parenteral -- may positively influence liver metabolism, muscle function, and immune status. Nutrition therapy should continue in the short- and also in the long-term post-transplant periods. For malnourished patients, early post-transplant enteral or parenteral nutrition have been useful in improving nutritional status. Finally, the metabolic and nutritional care of the liver transplant donor must be considered to reduce allograft dysfunction indices.     

Enhancement of apomorphine-induced penile erection in the rat by a selective alpha(1D)-adrenoceptor antagonist

1. Effects of A-322312 (alpha(1B)-adrenoceptor (AR) antagonist), A-119637 (alpha(1D)-AR antagonist), prazosin (non-selective alpha(1)-AR antagonist), and yohimbine (alpha(2)-AR antagonist) were studied in rat corpus cavernosum (CC) and cavernous artery (Acc) preparations. Effects of intracavernous (i.c.) or intraperitoneal (i.p.) administration of alpha(1)-AR antagonists on apomorphine-induced erections were investigated. 2. A-119637 attenuated electrically induced contractions in isolated CC (-logIC(50); 8.12+/-0.15), and relaxed noradrenaline (NA)-contracted preparations by more than 90% at 10(-7) M. At the same concentration, the -logEC(50) value for NA in Acc was altered from 6.79+/-0.07 to 4.86+/-0.13. In the CC and Acc, prazosin similarly inhibited contractile responses. 3. Inhibitory effects of A-322312 (10(-7) M) in electrically activated CC were 32.3+/-5.1%, whereas no effect on concentration-response curves for NA was observed in the Acc. Yohimbine (10(-8) M and 10(-7) M), enhanced electrically-induced contractions in isolated CC by 20 to 50%. At 10(-6) M, inhibitory effects of yohimbine were obtained. 4. A-119637 (0.3 micromol kg(-1), i.p.) tripled the number of erections, and produced a 6 fold increase in the duration of apomorphine-induced erectile responses. A-322312, prazosin, or yohimbine did not enhance erections induced by apomorphine. None of the alpha(1)-AR antagonists significantly increased ICP upon i.c. administration. Decreases in blood pressure were seen with A-119637 and prazosin. 5. The present findings show that there is a functional predominance of the alpha(1D)-AR subtype in the rat erectile tissue, and that blockade of this receptor facilitates rat penile erection induced by a suboptimal dose of apomorphine.     

The role of migrating leukocytes in IL-1 beta-induced up-regulation of kinin B(1) receptors in rats

1. The present study examines the role of migrating leukocytes in the ability of IL-1 beta to induce the functional up-regulation of B(1) receptors, as assessed by kinin B(1) agonist-induced oedema in the rat paw. 2. Pre-treatment with the PAF receptor antagonist WEB 2086 inhibited des-Arg(9)-BK-induced oedema in IL-1 beta-treated paws, while the LTB(4) receptor antagonist CP105696 had no effect. Des-Arg(9)-BK-induced paw oedema was also inhibited by pre-treatment with the selectin blocker fucoidin or by an anti-CD-18 monoclonal antibody. 3. I.d. injection of IL-1 beta produced a 5 - 10-fold increase of myeloperoxidase (MPO) activity in the rat paw. The increase in MPO activity was significantly inhibited by WEB 2086 (46 +/- 9%), fucoidin (68 +/- 5%) or the CD-18 antibody (84 +/- 3%). In contrast, i.d. injection of TNF alpha a dose known to upregulate the B(1) receptor functionally did not induce any significant increase in MPO activity. 4. Des-Arg(9)-BK alone had no effect in MPO activity but enhanced (by about 40%) the response induced by IL-1 beta, an effect prevented by the B(1) receptor antagonist des-Arg(9)-[Leu(8)]-BK. 5. The concentration of TNF-alpha was increased in the paws after i.d. injection of IL-1 beta. Pre-treatment with fucoidin, WEB 2086, anti-CD-18 or CP 105695, significantly reversed the local increases in TNF-alpha concentrations (80 +/- 2; 75 +/- 4, 73 +/- 3 and 40 +/- 2%), respectively. 6. Finally, IL-1 beta induced an increase of B(1) receptor mRNA levels in the rat paw, an effect which was prevented by fucoidin treatment. 7. Taken together, these results indicate that up-regulation of B(1) receptors in the rat paw following IL-1 beta seems to involve the local recruitment of neutrophils and subsequent local TNF-alpha production. The cross-talk between kinins, cytokines and leukocytes implicate B(1) receptors in chronic inflammatory diseases.