Effects of a novel fermented soy product on the serum lipids of hypercholesterolemic rabbits

OBJECTIVE: To assess the effect of a new feed soy product fermented by Enterococcus faecium and Lactobacillus jugurti on the serum lipid levels of rabbits with induced hypercholesterolemia. METHODS: Thirty-two rabbits were divided into 4 groups as follows: 1) control (C); 2) hypercholesterolemic (H); 3) hypercholesterolemic + fermented product (HPF); and 4) control + fermented product (CPF). The H and HPF groups were fed with a diet with 0.15% (p/p) cholesterol in the first 15 days. C and CPF groups received regular food preparation. The HPF and CPF groups received 10 mL daily of the fermented 30 days. Blood samples were drawn at the beginning of the study and at the 15th and 30th days. Concentrations of total cholesterol, HDL-cholesterol, and triglycerides were analyzed. RESULTS: After 15 days, the HPF group showed a total cholesterol concentration lower (18.4%) than that of the H group (p=0.05), but this difference disappeared after 30 days. No change was observed in total cholesterol levels of C and CPF groups. After 15 days, the HDL-cholesterol was higher (17.8%) in the HPF group, but the triglyceride levels remained unchanged in all groups during the same period of time. CONCLUSION: The soy fermented product caused an 18.4% reduction in total cholesterol and a 17.8% increase in the HDL-fraction. It may, therefore, be a possible coadjutor in the treatment of hypercholesterolemia.

QRS Voltage-Duration Product in the Identification of Left Ventricular Hypertrophy in Spontaneously Hypertensive Rats

OBJECTIVE - Evaluation of the performance of the QRS voltage-duration product (VDP) for detection of left ventricular hypertrophy (LVH) in spontaneously hypertensive rats (SHR). METHODS - Orthogonal electrocardiograms (ECG) were recorded in male SHR at the age of 12 and 20 weeks, when systolic blood pressure (sBP) reached the average values of 165±3 mmHg and 195±12 mmHg, respectively. Age- and sex- matched normotensive Wistar Kyoto (WKY) rats were used as controls. VDP was calculated as a product of maximum QRS spatial vector magnitude and QRS duration. Left ventricular mass (LVM) was weighed after rats were sacrificed. RESULTS - LVM in SHR at 12 and 20 weeks of age (0.86±0.05 g and 1.05±0.07 g, respectively) was significantly higher as compared with that in WKY (0.65±0.07 g and 0.70±0.02 g). The increase in LVM closely correlated with the sBP increase. VDP did not reflect the increase in LVM in SHR. VDP was lower in SHR as compared with that in WKY, and the difference was significant at the age of 20 weeks (18.2mVms compared with 10.7mVms, p<0.01). On the contrary, a significant increase in the VDP was observed in the control WKY at the age of 20 weeks without changes in LVM. The changes in VDP were influenced mainly by the changes in QRSmax. CONCLUSION - LVM was not the major determinant of QRS voltage changes and consequently of the VDP. These data point to the importance of the nonspatial determinants of the recorded QRS voltage in terms of the solid angle theory.

Inhibition of growth of Leishmania mexicana mexicana by Leishmania mexicana amazonensis during "in vitro" co-cultivation

Inhibition of one Leishmania subspecies by exometabolites of another subspecies, a phenomenon not previously reported, is suggested by our recent observations in cell cloning experiments with Leishmania mexicana mexicana and Leishmania mexicana amazonensis. Clones were identified using the technique of schizodeme analysis. The phenomenon observed is clearly relevant to studies of parasite isolation, leishmanial metabolism, cross-immunity and chemotherapy.

Serological diagnosis of dengue by an Elisa inhibition method (EIM)

An ELISA Inhibition Method (EIM) was proposed for the serologic diagnosis of dengue, comparing its results with the Hemagglutination Inhibition (HI) and the IgM capture-ELISA (MAC-ELISA). Advantages and disadvantages of both methods are discussed according to sensitivity, specificity, performance and usefulness. As a conclusion we recommend the complementary inclusion of the EIM and MAC-ELISA substituting the HI for laboratories engaged in the diagnosis and surveillance of dengue.

Inhibition of gastric secretion by a water extract from Baccharis triptera, Mart

Baccharus triptera Mart, is a widespread Compositae used in Brazilian folk medicine to treat gastrointestinal disturbances, rheumatic disease, mild fever, diabetes and as an anti-helminthic. Water extract of small branches of the plant (WE) administered to mice and rats (0.1 to 2 g/Kg, p.o) did not alter spontaneous motor activity, sleeping time induced by barbiturates or the tailflick response in mice. The extract decreased by 40 por cento the number of writhings induced by 0.8 por cento scetic acid, i.p., but did not influence paw edema induced by carrageenan or dextran in rats WE (2g/Kg, p.o.) decreased the intestinal transit of charcoal in mice by 20//. Gastric secretion in pylorus ligated rats was reduced after treatment with WE (1 and 2 g/Kg. i.p. or intraduodenal and the gastric pH was raised. The extract (1 g/Kg, p.o.) prevented gastric ulcers induced in rats by immobilization at 4ºC, but not those induced by indomethacin (10 mg/Kg, s.c.). The results indicate that WE may relieve gastrointestinal disorders by reducing acid secretion and gastrointestinal hiperactivity. Neither analgesic nor anti-inflammatory activities were detectable.

Basic biochemical investigations as rationale for the design of original antimalarial drugs. An example of phospholipid metabolism

The future of antimalarial chemotherapy is particulary alarming in view of the spread of parasite cross-resistances to drugs that are not even structurally related. Only the availability of new pharmacological models will make it possible to select molecules with novel mechanisms of action, thus delaving resistance and allowing the development of new chemotherapeutic strategies. We reached this objective in mice. Our approach is hunged on fundamental and applied research begun in 1980 to investigate to phospholipid (PL) metabolism of intraerythrocytic Plasmodium. This metabolism is abundant, specific and indispensable for the production of Plasmodium membranes. Any drug to interfere with this metabolism blocks parasitic development. The most effective interference yet found involves blockage of the choline transporter, which supplies Plasmodium with choline for the synthesis of phosphatidylcholine, its major PL, this is a limiting step in the pathway. The drug sensitivity thereshold is much lower for the parasite, which is more dependent on this metabolism than host cells. The compounds show in vitro activity against P. falciparum at 1 to 10 nM. They show a very low toxicity against a lymphblastoid cell line, demonstrating a total abscence of correlation between growth inhibition of parasites and lymphoblastoid cells. They show antimalarial activity in vivo, in the P. berghei or P. chabaudi/mouse system, at doses 20-to 100-fold lower than their in acute toxicity limit. The bioavailability of a radiolabeled form of the product seemed to be advantageous (slow blood clearance and no significant concentration in tissues). Lastly, the compounds are inexpensive to produce. They are stable and water-soluble.

Optimization and inhibition of the adherent ability of Plasmodium falciparum-infected erythrocytes

The vast majority of the 1-2 million malaria associated deaths that occur each year are due to anemia and cerebral malaria (the attachment of erythrocytes containing mature forms of Plasmodium falciparum to the endothelial cells that line the vascular beds of the brain). A "model" system"for the study of cerebral malaria employs amelanotic melanoma cells as the "target"cells in an vitro cytoadherence assay. Using this model system we determined that the optimum pH for adherence is 6.6 to 6.8, that high concentrations of Ca²* (50mM) result in increased levels of binding, and that the type of buffer used influences adherence (Bis Tris > MOPS > HEPES > PIPES). We also observed that the ability of infected erythrocytes to cytoadhere varied from (erythrocyte) donor to donor. We have produced murine monoclonal antibodies against P. falciparum-infected red cells which recognized modified forms of human band 3; these inhibit the adherence of infected erythrocytes to melanoma cells in a doso responsive fashion. Antimalarials (chloroquine, quinacrine, mefloquine, artemisinin), on the other hand, affected adherence in an indirect fashion i.e. since cytoadherence is due, in part to the presence of knobs on the surface of the infected erythrocyte, and knob formation is dependent on intracellular parasite growth, when plasmodial development is inhibited so is knob production, and consequently adherence is ablated.

An inhibition ELISA to determine alpha macroglobulin levels in mouse plasma

A sensitive method for quantifying mouse plasma alpha-macroglobulins (AM) using an inhibition ELISA is described. AM are important plasmaproteinase inhibitors that possibly act also as immunomodulatory molecules. The standard protocol develope in our experiments involves coating well with 10 µg/ml A2M in carbonate buffer, followed by incubation with a 1:1 (v/v) mixture of the plasma to be tested (diluted 1/1000) and goat anti-AM (diluted 1/1250). This is followed by further incubation, first with the enzyme-conjugated antibody and with the substrate prior to the reading of absorbance levels of the reaction products. Standard curve samples must be included in each plate, employing known amounts of the purified Murine Alpha-2-Macroglobulin (MuA2M) used for coating, with concentrations ranging from 0.001 to 10 µg/ml. Using test samples in triplicates and a 6-point standard curve in a single ELISA plate, 25 plasma samples can be tested accurately. The method offers an useful tool for establishing AM levelsin small samples of mouse plasma.

Partial inhibition of hemocyte agglutination by Lathyrus odoratus lectin in Crassotrea virginica infected with Perkinsus marinus

Quantitative determinations of agglutination of hemocytes from oysters, Crassostrea virginica, by the Lathyrus odoratus lectin at five concentrations revealed that clumping of hemocytes from oysters infected with Perkinsus marinus is partially inhibited. Although the nature of the hemocyte surface saccharide, which is not D(+)-glucose, D(+)mannose, or alpha-methyl-D-mannoside, remains to be determined, it may be concluded that this molecule also occurs on the surface of P. marinus. It has been demonstrated that the panning technique (Ford et al. 1990) is qualitatively as effective for determining the presence of P. marinus in C. virginica as the hemolymph assay method (Gauthier & Fisher 1990).

Interleukin-4 and interleukin-5 as targets for the inhibition of eosinophilic inflammation and allergic airways hyperreactivity

Clinical and experimental investigations suggest that allergen-specific CD4+ T-cells, IgE and the cytokines IL-4 and IL-5 play central roles in initiating and sustaining an asthmatic response by regulating the recruitment and/or activation of airways mast cells and eosinophils. IL-5 plays a unique role in eosinophil development and activation and has been strongly implicated in the aetiology of asthma. The present paper summarizes our recent investigations on the role of these cytokines using cytokine knockout mice and a mouse aeroallergen model. Investigations in IL-5-/- mice indicate that this cytokine is critical for regulating aeroallergen-induced eosinophilia, the onset of lung damage and airways hyperreactivity during allergic airways inflammation. While IL-4 and allergen-specific IgE play important roles in the regulation of allergic disease, recent investigations in IL4-/- mice suggest that allergic airways inflammation can occur via pathways which operate independently of these molecules. Activation of these IL-4 independent pathways are also intimately associated with CD4+ T-cells, IL-5 signal transduction and eosinophilic inflammation. Such IL-5 regulated pathways may also play a substantive role in the aetiology of asthma. Thus, evidence is now emerging that allergic airways disease is regulated by humoral and cell mediated processes. The central role of IL-5 in both components of allergic disease highlights the requirements for highly specific therapeutic agents which inhibit the production or action of this cytokine.

Inhibition of the polymerase chain reaction by sputum samples from tuberculosis patients after processing using a silica-guanidiniumthiocyanate DNA isolation procedure

With the objective to evaluate PCR-mediated detection of Mycobacterium tuberculosis DNA as a diagnostic procedure for diagnosis of tuberculosis in individuals attending ambulatory services in Primary Health Units of the City Tuberculosis Program in Rio de Janeiro, Brazil, their sputum samples were collected and treated with a DNA extraction procedure using silica-guanidiniumthiocyanate. This procedure has been described to be highly efficient for extraction of different kind of nucleic acids from bacteria and clinical samples. Upon comparing PCR results with the number of acid-fast bacilli, no direct relation was observed between the number of bacilli present in the sample and PCR positivity. Part of the processed samples was therefore spiked with pure DNA of M. tuberculosis and inhibition of the PCR reaction was verified in 22 out of 36 (61%) of the samples, demonstrating that the extraction procedure as originally described should not be used for PCR analysis of sputum samples.

Immunogenicity test of tetanus component in adsorbed vaccines by toxin binding inhibition test

Samples from 20 lots of diphtheria-tetanus (adult use dT) vaccine and from 20 lots of diphtheria-tetanus-pertussis (DTP) vaccine were used to standardize and validate the in vitro toxin binding inhibition (ToBI) test for the immunogenicity test of the tetanus component. The levels of tetanus antitoxin obtained by ToBI test were compared to those obtained using the toxin neutralization (TN) test in mice routinely employed to perform the quality control of the tetanus component in adsorbed vaccines. The results ranged from 1.8 to 3.5 IU/ml for dT and 2 to 4 IU/ml for DTP by ToBI test and 1.4 to 3 IU/ml for dT and 1.8 to 3.5 IU/ml for DTP by TN in mice. These results were significantly correlated. From this study, it is concluded that the ToBI test is an alternative to the in vivo neutralization procedure in the immunogenicity test of the tetanus component in adsorbed vaccines. A substantial refinement and a reduction in use of animals can be achieved.

Trypanosoma cruzi: inhibition of alpha-hydroxyacid dehydrogenase isozyme II by N-allyl and N-propyl oxamates and their effects on intact epimastigotes

N-allyl (NAOx) and N-propyl (NPOx) oxamates were designed as inhibitors of alpha-hydroxyacid dehydrogenase (HADH) isozyme II from Trypanosoma cruzi. The kinetic studies showed that NAOx and NPOx were competitive inhibitors of HADH-isozyme II (Ki = 72 µM, IC50 = 0.33 mM and 70 µM, IC50 = 0.32 mM, respectively). The attachment of the allylic and propylic chains to nitrogen of the competitive inhibitor oxamate (Ki = 0.91 mM, IC50 = 4.25 mM), increased 12.6 and 13-folds respectively, the affinity for T. cruzi HADH-isozyme II. NAOx and NPOx were selective inhibitors of HADH-isozyme II, because other T. cruzi dehydrogenases were not inhibited by these substances. Since HADH-isozyme II participates in the energy metabolism of T. cruzi, a trypanocidal effect can be expected with these inhibitors. However, we were not able to detect any trypanocidal activity with these oxamates. When the corresponding ethyl esters of N-allyl (Et-NAOx) and N-propyl (Et-NPOx) oxamates were tested as a possible trypanocidal prodrugs, in comparison with nifurtimox and benznidazole, the expected trypanocidal effects were obtained.

The pathogenesis of diabetic complications: the role of DNA injury and poly(ADP-ribose) polymerase activation in peroxynitrite-mediated cytotoxicity

Recent work has demonstrated that hyperglycemia-induced overproduction of superoxide by the mitochondrial electron-transport chain triggers several pathways of injury [(protein kinase C (PKC), hexosamine and polyol pathway fluxes, advanced glycation end product formation (AGE)] involved in the pathogenesis of diabetic complications by inhibiting glyceraldehyde-3-phosphate dehydrogenase (GAPDH) activity. Increased oxidative and nitrosative stress activates the nuclear enzyme, poly(ADP-ribose) polymerase-1 (PARP). PARP activation, on one hand, depletes its substrate, NAD+, slowing the rate of glycolysis, electron transport and ATP formation. On the other hand, PARP activation results in inhibition of GAPDH by poly-ADP-ribosylation. These processes result in acute endothelial dysfunction in diabetic blood vessels, which importantly contributes to the development of various diabetic complications. Accordingly, hyperglycemia-induced activation of PKC and AGE formation are prevented by inhibition of PARP activity. Furthermore, inhibition of PARP protects against diabetic cardiovascular dysfunction in rodent models of cardiomyopathy, nephropathy, neuropathy, and retinopathy. PARP activation is also present in microvasculature of human diabetic subjects. The present review focuses on the role of PARP in diabetic complications and emphasizes the therapeutic potential of PARP inhibition in the prevention or reversal of diabetic complications.