The use of multiple antigenic peptide (MAP) in the immunodiagnosis of human immunodeficiency virus infection

In order to develop an ELISA system for the antibody detection of HIV-1 or HIV-2 infections, MAPs for HIV-1 gp41(584-618) and HIV-2 gp36(574-602) corresponding to the immunodominant regions of HIV-1 gp41 and HIV-2 gp36 were used as coating antigens in the ELISA. The MAPs were synthesized by the solid phase method using Fmoc-Lys(Fmoc)-OH and their molecular weights were confirmed by tricine gel electrophoresis. The MAPs reacted with all HIV positive sera (64 samples), but did not react with HIV negative sera (48 samples) obtained from healthy blood donors. The MAPs showed high sensitivity and specificity in anti-HIV 1/2 combo panel and anti-HIV-1 seroconversion panels. The results indicated that the ELISA system using synthetic MAPs of gp41(584-618) and gp36(574-602) as coating antigens can be used as an analytical system for the immunodiagnosis of HIV-1 or HIV-2 infections.     

Leptin is present in human milk and is related to maternal plasma leptin concentration and adiposity

Leptin is elevated during pregnancy and may be involved in the regulation of milk production in women. Immunoreactive leptin was quantified in human milk by modified radioimmunoassay. Leptin concentration was higher in whole vs. skim milk fractions; however, leptin concentration was not correlated with percentage milk fat. Leptin concentrations in whole and skim milk were correlated with maternal plasma leptin concentrations, maternal body weight, body mass index, and tricep skinfold thickness, but not with plasma insulin concentration. These data provide the first evidence for the presence of leptin in human milk in the range of concentrations found in human plasma and indicate that the concentration of leptin in milk reflects maternal adiposity. Determining the biological role(s) of milk-borne leptin could add to our understanding of neonatal metabolism and the mechanisms underlying the development of body fat and obesity in humans.     

Deterioration of PCPV concrete

This paper presents results obtained from an experimental investigation into the deterioration of concretes used in prestressed concrete pressure vessels (PCPVs) in the UK. The experimental works were focused on the dependence of the residual properties of the thermally loaded concretes on the mix design (3 representative PCPVs), levels of exposed temperature (from 20 to 450°C), test age (3 months and 1 year) and exposure condition (sealed and unsealed). The residual properties investigated in this project were conducted by fully using both destructive (compressive, tensile splitting and static modulus) and non-destructive (dynamic modulus and ultrasonic pulse velocity) test techniques with the aim of better understanding and characterising the deterioration of PCPV concretes when subjected to both ambient and elevated temperatures under the environmental conditions likely to be experienced in real structures. Detailed statistical analyses and discussions on the experimental results were also performed and presented.     

Possible involvement of plasmids in degradation of malathion and chlorpyriphos by Micrococcus sp

Culture is the basic method in bacteriology. It allowed the discovery of Helicobacter pylori. Problems in the culture of this fragile, slow-growing bacterium concern transport and processing in the laboratory, but they can be solved. Culture has a 100% specificity. When performed properly, it has a sensitivity in the range of the other best diagnostic methods for Helicobacter pylori. It allows strain typing and, most importantly, susceptibility testing to antibiotics, because an increased rate of acquired resistance of Helicobacter pylori is currently observed. Culture must be performed in clinical trials, at least when antibiotics, to which Helicobacter pylori may be resistant, are used. In clinical practice, culture and susceptibility testing can generally be restricted to treatment failures. However, it is important to monitor Helicobacter pylori susceptibility to antibiotics at a national or regional level in order to give recommendations for primary treatment.     

CD4-CD8-C.B-17 SCID thymocytes enter the CD4+CD8+ stage in the presence of neonatally grafted T cells

Our previous studies in iron-loaded rat heart cells showed that in vitro iron loading results in peroxidative injury, manifested in a marked decrease in rate and amplitude of heart cell contractility and rhythmicity, which is correctable by treatment with deferoxamine (DF). In the present studies we explored the role of mitochondrial damage in myocardial iron toxicity. Iron loading by 24-hour incubation with 0.36 mmol/L ferric ammonium citrate resulted in a decrease in the activity of nicotinamide adenine dinucleotide (NADH)-cytochrome c oxidoreductase (complex I+III) to 35.3%+/-11.2% of the value in untreated controls; of succinate-cytochrome c oxidoreductase (complex II+III) to 57.4%+/-3.1%; and of succinate dehydrogenase to 63.5%+/-12.6% (p < 0.001 in all cases). The decrease in activity of other mitochondrial enzymes, including NADH-ferricyanide reductase, succinate ubiquinone oxidoreductase (complex II), cytochrome c oxidase (complex IV), and ubiquinol cytochrome c oxidoreductase (complex III), was less impressive and ranged from 71.5%+/-15.8% to 91.5%+/-14.6% of controls. That the observed loss of respiratory enzyme activity was a specific effect of iron toxicity was clearly demonstrated by the complete restoration of enzyme activities by in vitro iron chelation therapy. Sequential treatment with iron and doxorubicin caused a loss of complex I+III and complex II+III activity that was greater than that seen with either agent alone but was only partially correctable by DF treatment. Alterations in cellular adenosine triphosphate measurements paralleled very closely the changes observed in respiratory complex activity. These findings demonstrate for the first time the impairment of cardiac mitochondrial respiratory enzyme activity caused by iron loading at conditions formerly shown to produce severe abnormalities in contractility and rhythmicity.     

Beneficial effects of S-adenosyl-L-methionine on blood-brain barrier breakdown and neuronal survival after transient cerebral ischemia in gerbils

In the present study, the effect of bradykinin on basal and precontracted mouse-isolated trachea was investigated. In basal conditions mouse-isolated tracheal rings do not respond to bradykinin. However, when the tracheal rings were precontracted with carbachol (10(-7) M) a relaxation with bradykinin (3 x 10(-9)-3 x 10(-7)) was found. The maximal response amounted 69.7+/-4.1% (n=15) with a pD2 value of 7.2+/-0.21. The selective bradykinin B2 receptor antagonist HOE 140 (10(-10)-10(-8) M) antagonized the bradykinin-induced relaxation, while the bradykinin B1 receptor antagonist des-Arg9-Leu8-bradykinin (10(-6) M) had no influence. The selective bradykinin B1 receptor agonist des-Arg9-bradykinin (10(-6) M) caused a small relaxation (8.4+/-2.5%, n=6), which could be antagonized completely by the selective bradykinin B1 receptor antagonist des-Arg9-Leu8-bradykinin (10(-6) M) while addition of the selective bradykinin B2 receptor antagonist HOE 140 (10(-8) M) was without effect. In the presence of indomethacin (10(-6) M) the relaxation of bradykinin was completely abolished. Pretreatment of the tracheal rings with capsaicin, or the presence of the selective NK1 receptor antagonist RP 67851 (10(-6) M) or the presence of the nitric oxide synthase inhibitor L-NAME (3 x 10(-4) M) had no effect on the bradykinin-induced relaxation. In conclusion, these results demonstrate that the mouse-isolated tracheal is a preparation in which bradykinin exerts a relaxant response via stimulation of bradykinin B2 receptors. This response is probably mediated by prostaglandins.     

Ascorbic acid inhibits lipid peroxidation but enhances DNA damage in rat liver nuclei incubated with iron ions

In this report we studied DNA damage and lipid peroxidation in rat liver nuclei incubated with iron ions for up to 2 hrs in order to examine whether nuclear DNA damage was dependent on membrane lipid peroxidation. Lipid peroxidation was measured as thiobarbituric acid-reactive substances (TBARS) and DNA damage was measured as 8-OH-deoxyguanosine (8-OH-dG). We showed that Fe(II) induced nuclear lipid peroxidation dose-dependently but only the highest concentration (1.0 mM) used induced appreciable 8-OH-dG. Fe(III) up to 1 mM induced minimal lipid peroxidation and negligible amounts of 8-OH-dG. Ascorbic acid enhanced Fe(II)-induced lipid peroxidation at a ratio to Fe(II) of 1:1 but strongly inhibited peroxidation at ratios of 2.5:1 and 5:1. By contrast, ascorbate markedly enhanced DNA damage at all ratios tested and in a concentration-dependent manner. The nuclear DNA damage induced by 1 mM FeSO4/5 mM ascorbic acid was largely inhibited by iron chelators and by dimethylsulphoxide and mannitol, indicating the involvement of OH. Hydrogen peroxide and superoxide anions were also involved, as DNA damage was partially inhibited by catalase and, to a lesser extent, by superoxide dismutase. The chain-breaking antioxidants butylated hydroxytoluene and diphenylamine (an alkoxyl radical scavenger) did not inhibit DNA damage. Hence, this study demonstrated that ascorbic acid enhanced Fe(II)-induced DNA base modification which was not dependent on lipid peroxidation in rat liver nuclei.     

Importance of the Arg-Gly-Asp triplet in human thrombin for maintenance of structure and function

Site-directed mutagenesis was employed to assess the importance of the Arg-Gly-Asp triplet that comprises residues 197 to 199 in the B-chain of thrombin. Properties of the R197E and the D199E variants were compared with those of zeta-thrombin and the inactive S205A variant wherein the active site Ser is replaced by Ala. Relative to zeta-thrombin, the R197E thrombin variant under the assay conditions used exhibits 26% activity toward a small chromogenic substrate, 13% activity in the activation of protein C in the presence of thrombomodulin, < 3% activity in processing fibrinogen, and 1% activity in inducing platelet activation. Thus, the substrate specificity of thrombin was altered by the R197-->E replacement. The D199E variant was essentially inactive. It exhibited only 0.02% of the activity of thrombin toward the chromogenic substrate and its reactivity toward the active site-directed alkylating agent D-Phe-Pro-Arg-CH2Cl was 10,000-fold lower than that of thrombin. Like the inactive S205A thrombin variant, the D199E variant antagonized the interactions of thrombin with hirudin and thrombomodulin, but was a less effective antagonist. The dependence of the antagonism of the thrombin-thrombomodulin interaction on the concentration of D199E thrombin variant provided evidence suggesting the presence of two or more domains in thrombin that independently interact with their counterparts in thrombomodulin. Although the S205A thrombin variant antagonized the action of thrombin on platelets no such activity could be demonstrated for the D199E variant in the concentration range studied (< 800 nm). Comparison of the circular dichroism spectra of zeta-thrombin, the D199E, R197E, and S205A variants indicated that subtle differences in conformation exist between the D199E variant and the other thrombins. These differences in conformation might well account for the altered behavior of the D199E variant with respect to its interactions toward thrombomodulin, hirudin, and platelets.