Heterogeneity of metallo and serine extracellular proteinases in oral clinical isolates of Candida albicans in HIV-positive and healthy children from Rio de Janeiro, Brazil

Candida yeasts frequently cause life-threatening systemic infections in immunocompromised hosts. In the present study, gelatin-SDS-PAGE analysis was used to characterize extracellular proteinases in 44 oral clinical isolates of Candida albicans from HIV-positive (29/50) and healthy children (15/50). Our survey indicates that these oral clinical isolates of C. albicans have complex extracellular proteolytic activity profiles, which illustrates the heterogeneity of this species. We showed four distinct proteolytic patterns composed of distinct serine (30-58 kDa) and metalloproteinase (64-95 kDa) activities, based on the inhibition profile with phenylmethylsulfonyl fluoride and 1,10-phenanthroline, respectively. This is the first report on secreted serine and metalloproteinases present in the culture supernatant fluids of C. albicans; however, we did not observe a significant correlation between proteolytic profile expressed by the C. albicans isolates from HIV-positive children and CD4(+) T cell count and plasma viral load.     

Effects of zinc on cell-mediated immunity in chronic hemodialysis patients

Thirteen healthy subjects and 20 hemodialysis patients were studied to observe the delayed hypersensitivity skin tests (DHSTs) and phytohemagglutinin (PHA)-stimulating lymphocyte blastogenesis. Significant differences were observed between the groups. Controls had a higher proportion of positive skin reaction than hemodialysis patients in relation to Escherichia coli (p<0.01) and tuberculin (PPD) (p<0.05). Regarding lymphocyte blastogenesis stimulated by phytohemagglutinin (PHA), cell proliferation was more accentuated in controls than hemodialysis patients (p<0.05). On the other hand, serum zinc was elevated in controls (78 +/- 8 microg/dL) in comparison to hemodialysis patients (71 +/- 33 microg/dL) (p<0.05). Of the 20 hemodialysis patients, 8 patients were maintained on long-term hemodialysis before and after zinc therapy, with the aim of studying DHST and PHA-stimulating lymphocyte blastogenesis. There was a significant improvement of DHST response to E. coli antigen after 100 d of zinc treatment (p<0.01), and with the discontinuation of therapy, the DHST responses decreased back to the initial values (p<0.05). Zinc administration also increased the lymphocyte proliferation induced by PHA from 31386 +/- 3974 to 42480 +/- 5242 cpm (mean +/- SD) (p<0.05). These results indicated that zinc therapy improved in vivo and in vitro DHST and lymphocyte function of hemodialysis patients and that its discontinuation suppressed all of the benefits observed.     

Cyclooxygenase enzymes and prostaglandins in reproductive tract physiology and pathology

Prostaglandins, thromboxanes (TX) and leukotrienes, collectively referred to as eicosanoids, are cyclooxygenase (COX) metabolites of arachidonic acid (AA). Prostaglandins, have been recognised for many years as key molecules in regulating reproductive tract physiology and pathology. Numerous recent studies in in vitro model systems and knockout mouse models have demonstrated specific functional roles for the respective cyclooxygenase enzymes, prostaglandins and prostanoid receptors. Here we review the findings obtained in several of these studies with emphasis on the roles played by cyclooxygenase enzymes and prostaglandins, specifically prostaglandin E2 (PGE2) and F2alpha in reproductive tract physiology and pathology.     

Combination ecotoxicity and testing of common chemical discharges to sewer using the<Emphasis Type="Italic"> Vibrio fischeri</Emphasis> luminescence bioassay

In order to investigate possible synergistic or antagonistic (more or less than additive) toxicity effects, mixtures of chemicals were tested in water using a microbial bioassay. Ten toxicants (3,4-dichloroaniline, 3,5-dichlorophenol, cadmium, chromium, copper, Lindane, linear alkylbenzene sulphonate, pentachlorophenol, toluene, zinc) were chosen on the basis of their common occurrence in industrial effluents within local waste water treatment plants. These toxicants also cover a wide range of modes of toxic action, namely, polar and non-polar narcosis, membrane disruption, respiratory disruption, uncouplers of oxidative phosphorylation, biochemical disruption and enzyme inhibition. Efficient screening for possible combination toxicity between toxicants involved testing the chemicals both singly and in triplet combinations. The triplets were based on four replicates of a balanced incomplete block design (BIB). A standardised Vibrio fischeri rapid toxicity bioluminescence assay was used. The combinations tested showed that only one mixture was found to be significantly more toxic than expected from the pure single-toxicant results. Two triplets were significantly less toxic. Further tests on the more toxic triplet showed that the effect was due to only one of the 45 pairs originally screened. It is concluded that synergistic effects in combinations of toxicants are rather rare in bioluminescence systems utilising common effluents discharged to sewer. Electronic Publication     

Isolation of fungi in Musca domestica Linnaeus, 1758 (Diptera: Muscidae) captured at two natural breeding grounds in the municipality of Serop dica,Rio de Janeiro,Brazil

The objective of this study was to isolate and identify fungal species found in natural association with adults of Musca domestica. The adult insects were collected from two natural breeding grounds: hog pens and an urban sanitary landfill. The isolated fungi were identified as: Aspergillus flavus (23.8%), A. niger var. niger (14.4%), Penicillium corylophilum (21.4%), P. fellutanum (11.9%), Cladosporium cladosporoides (4.7%), Fusarium sp. (4.7%), Alternaria alternata (11.9%), Curvularia brachyspora (2.4%), Mycelia sterilia (2.4%) and the Mucorales order (2.4%).     

The effect of ambient temperature on body temperature and on ultrasonic behaviour in litters of albino laboratory mice deprived of their mothers

Litters of mouse pups of various ages when deprived of their mother and kept for one hour at ambient temperatures of 3°C, 12°C and 22°C lose heat less rapidly than pups isolated individually at similar temperatures. The litters are able to maintain constant body temperatures of 35–37°C by day 8 at 22°C, by day 19 at 12°C and by day 21 at 3°C. Very little ultrasound is detected from litters of any age up to 21 days at 22°C while at 12°C ultrasound is mainly detected from pups of 4 and 6 days; very few calls are produced by older pups at this temperature. Considerably more calls are emitted at 3°C than at either of the two higher temperatures. At 3°C the total number of calls emitted during the hour-long exposure period varied markedly with age. There were two peaks of calling, one on day 6 and the other on day 16. The temporal pattern of calling throughout the hour-long exposure period at 3°C was similar in pups of 4 and 6 days and in pups of 8, 10, 12, 14 and 16 days, but differed between these two groups. In the younger group of pups the rate of calling rose to a peak and then decreased to zero as the pups became comatose; in the older pups the rate of calling was low initially then either rose throughout the rest of the hour or rose to a high level that was maintained. It is suggested that the various patterns of ultrasonic calling can be associated with the physiological development of mouse litters.     

Analysis of the metabolites of ethyl loflazepate by gas chromatography with electron-capture detection

A gas chromatographic assay with electron-capture detection (GC—EC) is described for the metabolites of ethyl loflazepate (Victan), a new benzodiazepine with a potent anti-anxiety activity, in biological fluids. Since the parent drug undergoes a first-pass effect, pharmacokinetic data may only be obtained by measuring the total levels of two of the major metabolites. Accurate data can not be obtained for the metabolites separately since one of them (M1) is chemically transformed to the other (M2) during plasma sampling, storage and extraction.A sensitive, specific and accurate GC—EC assay is developed using a synthetic analogue of M2 as an internal standard. The limit of detection in plasma is approximately 2 ng/ml and the precision about 3% (within-run and between-run).The method is applied to plasma samples collected after oral administration of 2 mg and 4 mg of the drug in tablet form to human volunteers. The results obtained are correlated with those from an existing gas chromatographic—mass spectrometric assay. A very good correlation between the results (inter-laboratory comparison) is obtained, validating both techniques.     

Block of outward current in cardiac purkinje fibers by injection of quaternary ammonium ions

We have studied the effects of iontophoretic injection of the quaternary ammonium compounds tetraethylammonium (TEA) and tetrabutylammonium (TBA) in cardiac purkinje fibers. We find that TBA(+) is a more effective blocker than TEA(+), but injection of either compound reduces the time-dependent outward plateau currents, transient outward current (I(to)), and the delayed rectifier (I(x)). Our findings provide evidence that these outward cardiac currents are carried by channels that in some respects are pharmacologically similar to squid axon potassium channels. We demonstrate that this procedure is a new tool that can be useful in the analysis of membrane currents in the heart.     

Active immunization combined with cisplatin confers enhanced therapeutic protection and prevents relapses of HPV-induced tumors at different anatomical sites

The active immunotherapy concept relies on the use of vaccines that are capable of inducing antitumor immunity, reversion of the suppressive immunological environment, and long-term memory responses. Previously, antitumor vaccines based on a recombinant plasmid (pgDE7h) or a purified protein (gDE7) led to regression of early-established human papillomavirus (HPV)-associated tumors in a preclinical model. In this work, the anticancer vaccines were combined with cisplatin to treat HPV-induced tumors at advanced growth stages. The antitumor effects were evaluated in terms of tumor regression, induction of specific CD8⁺ T cells, and immune modulation of the tumor microenvironment. Acute toxicity induced by the treatment was measured by weight loss and histological alterations in the liver and kidneys. Our results revealed that the combination of cisplatin with either one of the tested immunotherapies (pgDE7h or gDE7) led to complete tumor regression in mice. Also, the combined treatment resulted in synergistic effects, particularly among mice immunized with gDE7, including activation of systemic and tumor-infiltrating E7-specific CD8⁺ T cells, tumor infiltration of macrophages and dendritic cells, and prevention of tumor relapses at different anatomical sites. Furthermore, the protocol allowed the reduction of cisplatin dosage and its intrinsic toxic effects, without reducing antitumor outcomes. These results expand our knowledge of active immunotherapy protocols and open perspectives for alternative treatments of HPV-associated tumors.     

Autoantibodies against Muscarinic Receptors in Breast Cancer: Their Role in Tumor Angiogenesis

The presence of autoantibodies in cancer has become relevant in recent years. We demonstrated that autoantibodies purified from the sera of breast cancer patients activate muscarinic acetylcholine receptors in tumor cells. Immunoglobulin G (IgG) from breast cancer patients in T1N0Mx stage (tumor size≤2 cm, without lymph node metastasis) mimics the action of the muscarinic agonist carbachol stimulating MCF-7 cell proliferation, migration and invasion. Angiogenesis is a central step in tumor progression because it promotes tumor invasion and metastatic spread. Vascular endothelial growth factor-A (VEGF-A) is the main angiogenic mediator, and its levels have been correlated with poor prognosis in cancer. The aim of the present work was to investigate the effect of T1N0Mx-IgG on the expression of VEGF-A, and the in vivo neovascular response triggered by MCF-7 cells, via muscarinic receptor activation. We demonstrated that T1N0Mx-IgG (10⁻⁸ M) and carbachol (10⁻⁹ M) increased the constitutive expression of VEGF-A in tumor cells, effect that was reverted by the muscarinic antagonist atropine. We also observed that T1N0Mx-IgG and carbachol enhanced the neovascular response produced by MCF-7 cells in the skin of NUDE mice. The action of IgG or carbachol was reduced in the presence of atropine. In conclusion, T1N0Mx-IgG and carbachol may promote VEGF-A production and neovascularization induced by breast tumor cells via muscarinic receptors activation. These effects may be accelerating breast tumor progression.