Targeting NADPH Oxidase and Phospholipases A2 in Alzheimer’s Disease

Alzheimer’s disease (AD) is marked by an increase in the production of extracellular beta amyloid plaques and intracellular neurofibrillary tangles associated with a decline in brain function. Increases in oxidative stress are regarded as an early sign of AD pathophysiology, although the source of reactive oxygen species (ROS) and the mechanism(s) whereby beta amyloid peptides (Aβ) impact oxidative stress have not been adequately investigated. Recent studies provide strong evidence for the involvement of NADPH oxidase and its downstream oxidative signaling pathways in the toxic effects elicited by Aβ. ROS produced by NADPH oxidase activate multiple signaling pathways leading to neuronal excitotoxicity and glial cell-mediated inflammation. This review describes recent studies demonstrating the neurotoxic effects of Aβ in conjunction with ROS produced by NADPH oxidase and the downstream pathways leading to activation of cytosolic phospholipase A₂ (PLA₂) and secretory PLA₂. In addition, this review also describes recent studies using botanical antioxidants to protect against oxidative damage associated with AD. Investigating the metabolic and signaling pathways involving Aβ NADPH oxidase and PLA₂ can help understand the mechanisms underlying the neurodegenerative effects of oxidative stress in AD. This information should provide new therapeutic approaches for prevention of this debilitating disease.     

Evaluation of the Qualitative and Automated Quantitative Microhemagglutination Assay for Antibodies to Treponema pallidum

Qualitative and quantitative microhemagglutination assays for antibodies to Treponema pallidum (MHA-TP) were performed on 314 syphilitic and 597 presumably nonsyphilitic sera, and the results were compared with those of the fluorescent treponemal antibody-absorbed (FTA-ABS), the Treponema pallidum immobilization (TPI), and the Veneral Disease Research Laboratory (VDRL) tests. MHA-TP sensitivity was similar to that of the other tests in all stages of syphilis except primary syphilis, in which MHA-TP reactivity was only 64% compared with 82% in the FTA-ABS test, 73% in the VDRL test, and 67% in the TPI test. MHA-TP specificity was satisfactory and comparable to that of the other treponemal tests. Quantitation of the MHA-TP test was automated by use of Autotiter II equipment. Titers tended to become elevated later in the course of syphilis and to remain elevated longer than did VDRL titers. Reproducibility of the quantitative MHA-TP test was satisfactory, with duplicate tests agreeing within one doubling dilution on 97.5% of 351 reactive sera. Poor reproducibility was obtained with sera giving minimal reactions in the qualitative test, and such sera should be routinely retested. The MHA-TP is less time-consuming and costly than the FTA-ABS test and could be used in conjunction with the VDRL or another reagin test for syphilis to eliminate a large number of the FTA-ABS tests now required.     

CGS 10746B: an atypical antipsychotic candidate that selectively decreases dopamine release at behaviorally effective doses

CGS 10746B, a benzothiadiazepine, has a behavioral profile in mice and monkeys similar to the atypical antipsychotic clozapine. Unlike clozapine, CGS 10746B suppresses dopamine neuron firing rates and, when administered at behaviorally effective doses by the oral or intraperitoneal route, decreases neostriatal dopamine release without changing dopamine metabolism or occupying D2 receptors. CGS 10746B is the first atypical antipsychotic candidate that selectively decreases dopamine release.     

Schistosoma mansoni: serotonin uptake and its drug inhibition

5-Hydroxytryptamine (serotonin) uptake by male and female Schistosoma mansoni was studied in vitro. Initial uptake was determined during the first 2 min of incubation with the indoleamine. There were two types of uptake: a saturable component that is apparent at low concentrations of serotonin, and a passive component that predominates at high concentrations. Female S. mansoni take up more 5-hydroxytryptamine per milligram protein than do the males. Studies on the distribution of 5-hydroxytryptamine taken up by the parasites show that a large proportion of the indoleamine is in the tegument. The female tegument had a greater proportion of the 5-hydroxytryptamine taken up than did the male tegument. The evidence indicated that 5-hydroxytryptamine associated with tegument represented the passive uptake while uptake in the main parasite body was both active and passive. Compounds that were shown to inhibit 5-hydroxytryptamine uptake include methylclonazepam, metergoline, imipramine, and ouabain. Maximal inhibition by these compounds ranged from 50 to 60%. Inhibition appeared to be confined to the saturable uptake component of the body fraction.     

Preparation of standards and determination of sizes of long-chain polyphosphates by gel electrophoresis

Procedures are presented for isolating fractions of long-chain polyphosphates which have a narrow range of sizes and for determining their chain lengths. The polyphosphates are isolated by elution from preparative polyacrylamide gels. Then, the lengths of these polymers are determined by a method of successive approximations of length from data obtained by electrophoresis on several different gels of varying polyacrylamide concentrations. Once sized, these isolated polyphosphates may be used as electrophoresis standards, making it possible to rapidly and accurately ascertain the size of other samples having unknown chain lengths. By comparison with two other procedures for sizing polyphosphates, it is shown that the method is definitely valid to a length of 450 and most likely to a length of at least 900. This electrophoresis procedure allows, for the first time, the determination of the range of sizes present and the average chain length with only 2-20 micrograms of polyphosphate.     

Pathogenicity of mouse hepatitis virus for preimplantation mouse embryos

Mouse embryos which were hatched from the zona pellucida in vitro in the presence of mouse hepatitis virus (MHV) or outgrown on coverslips and then exposed to MHV were shown by immunohistochemical staining to have virally infected trophoblast cells. Zona-intact embryos incubated with MHV for 48 h (2-cell embryos) or 1.5 h (blastocysts) were resistant to infection. Morulae and early blastocysts collected from donor mice experimentally infected with MHV were not infected, but the medium in which they were flushed from the uterine horns was contaminated with virus. No virus was detected after embryos were washed through three changes of uncontaminated medium. MHV was transmitted to foster mothers when embryos were transferred in medium contaminated with the virus. Fetal and decidual tissues were not infected. We suggest that embryo transfer is an effective and simple alternative to Caesarian rederivation of MHV-contaminated mice.     

HIV Evolution in Early Infection: Selection Pressures,Patterns of Insertion and Deletion,and the Impact of APOBEC

The pattern of viral diversification in newly infected individuals provides information about the host environment and immune responses typically experienced by the newly transmitted virus. For example, sites that tend to evolve rapidly across multiple early-infection patients could be involved in enabling escape from common early immune responses, could represent adaptation for rapid growth in a newly infected host, or could represent reversion from less fit forms of the virus that were selected for immune escape in previous hosts. Here we investigated the diversification of HIV-1 env coding sequences in 81 very early B subtype infections previously shown to have resulted from transmission or expansion of single viruses (n = 78) or two closely related viruses (n = 3). In these cases, the sequence of the infecting virus can be estimated accurately, enabling inference of both the direction of substitutions as well as distinction between insertion and deletion events. By integrating information across multiple acutely infected hosts, we find evidence of adaptive evolution of HIV-1 env and identify a subset of codon sites that diversified more rapidly than can be explained by a model of neutral evolution. Of 24 such rapidly diversifying sites, 14 were either i) clustered and embedded in CTL epitopes that were verified experimentally or predicted based on the individual''s HLA or ii) in a nucleotide context indicative of APOBEC-mediated G-to-A substitutions, despite having excluded heavily hypermutated sequences prior to the analysis. In several cases, a rapidly evolving site was embedded both in an APOBEC motif and in a CTL epitope, suggesting that APOBEC may facilitate early immune escape. Ten rapidly diversifying sites could not be explained by CTL escape or APOBEC hypermutation, including the most frequently mutated site, in the fusion peptide of gp41. We also examined the distribution, extent, and sequence context of insertions and deletions, and we provide evidence that the length variation seen in hypervariable loop regions of the envelope glycoprotein is a consequence of selection and not of mutational hotspots. Our results provide a detailed view of the process of diversification of HIV-1 following transmission, highlighting the role of CTL escape and hypermutation in shaping viral evolution during the establishment of new infections.     

Gene expression changes reflect clinical response in a placebo-controlled randomized trial of abatacept in patients with diffuse cutaneous systemic sclerosis

IntroductionSystemic sclerosis is an autoimmune disease characterized by inflammation and fibrosis of the skin and internal organs. We sought to assess the clinical and molecular effects associated with response to intravenous abatacept in patients with diffuse cutaneous systemic.MethodsAdult diffuse cutaneous systemic sclerosis patients were randomized in a 2:1 double-blinded fashion to receive abatacept or placebo over 24 weeks. Primary outcomes were safety and the change in modified Rodnan Skin Score (mRSS) at week 24 compared with baseline. Improvers were defined as patients with a decrease in mRSS of ≥30 % post-treatment compared to baseline. Skin biopsies were obtained for differential gene expression and pathway enrichment analyses and intrinsic gene expression subset assignment.ResultsTen subjects were randomized to abatacept (n = 7) or placebo (n = 3). Disease duration from first non-Raynaud’s symptom was significantly longer (8.8 ± 3.8 years vs. 2.4 ± 1.6 years, p = 0.004) and median mRSS was higher (30 vs. 22, p = 0.05) in the placebo compared to abatacept group. Adverse events were similar in the two groups. Five out of seven patients (71 %) randomized to abatacept and one out of three patients (33 %) randomized to placebo experienced ≥30 % improvement in skin score. Subjects receiving abatacept showed a trend toward improvement in mRSS at week 24 (−8.6 ± 7.5, p = 0.0625) while those in the placebo group did not (−2.3 ± 15, p = 0.75). After adjusting for disease duration, mRSS significantly improved in the abatacept compared with the placebo group (abatacept vs. placebo mRSS decrease estimate −9.8, 95 % confidence interval −16.7 to −3.0, p = 0.0114). In the abatacept group, the patients in the inflammatory intrinsic subset showed a trend toward greater improvement in skin score at 24 weeks compared with the patients in the normal-like intrinsic subset (−13.5 ± 3.1 vs. −4.5 ± 6.4, p = 0.067). Abatacept resulted in decreased CD28 co-stimulatory gene expression in improvers consistent with its mechanism of action. Improvers mapped to the inflammatory intrinsic subset and showed decreased gene expression in inflammatory pathways, while non-improver and placebos showed stable or reverse gene expression over 24 weeks.ConclusionsClinical improvement following abatacept therapy was associated with modulation of inflammatory pathways in skin.

Trial registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00442611","term_id":"NCT00442611"}}NCT00442611. Registered 1 March 2007.

Electronic supplementary material

The online version of this article (doi:10.1186/s13075-015-0669-3) contains supplementary material, which is available to authorized users.