Dimethyl sulfoxide blocks herpes simplex virus-1 productive infection in vitro acting at different stages with positive cooperativity. Application of micro-array analysis

BACKGROUND: Dimethyl sulfoxide (DMSO) is frequently used at a concentration of up to 95% in the formulation of antiherpetic agents because of its properties as a skin penetration enhancer. Here, we have analyzed the effect of DMSO on several parameters of Herpes Simplex Virus replication. METHODS: Productive infection levels of HSV-1 were determined by plaque assay or by reporter gene activity, and its DNA replication was estimated by PCR. Transcript levels were evaluated with HSV-specific DNA micro-arrays. RESULTS: DMSO blocks productive infection in vitro in different cell types with a 50% inhibitory concentration (IC(50)) from 0.7 to 2% depending upon the multiplicity of infection. The concentration dependence exhibits a Hill coefficient greater than 1, indicating that DMSO blocks productive infection by acting at multiple different points (mechanisms of action) with positive cooperativity. Consistently, we identified at least three distinct temporal target mechanisms for inhibition of virus growth by DMSO. At late stages of infection, DMSO reduces virion infectivity, and markedly inhibits viral DNA replication. A third mode of action was revealed using an oligonucleotide-based DNA microarray system for HSV. These experiments showed that DMSO reduced the transcript levels of many HSV-1 genes; including several genes coding for proteins involved in forming and assembling the virion. Also, DMSO markedly inhibited some but not all early transcripts indicating a previously unknown mode for inhibiting the early phase of HSV transcription-replication cycle. CONCLUSION: These observations suggest that DMSO itself may have a role in the anti-herpetic activity of formulations utilizing it as a dispersant

Outbreak of pandemic influenza A/H1N1 2009 in Nepal

<p>Abstract</p> <p>Background</p> <p>The 2009 flu pandemic is a global outbreak of a new strain of H1N1 influenza virus. Pandemic influenza A (H1N1) 2009 has posed a serious public health challenge world-wide. Nepal has started Laboratory diagnosis of Pandemic influenza A/H1N1 from mid June 2009 though active screening of febrile travellers with respiratory symptoms was started from April 27, 2009.</p> <p>Results</p> <p>Out of 609 collected samples, 302 (49.6%) were Universal Influenza A positive. Among the influenza A positive samples, 172(28.3%) were positive for Pandemic influenza A/H1N1 and 130 (21.3%) were Seasonal influenza A. Most of the pandemic cases (53%) were found among young people with ≤ 20 years. Case Fatality Ratio for Pandemic influenza A/H1N1 in Nepal was 1.74%. Upon Molecular characterization, all the isolated pandemic influenza A/H1N1 2009 virus found in Nepal were antigenically and genetically related to the novel influenza A/CALIFORNIA/07/2009-LIKE (H1N1)v type.</p> <p>Conclusion</p> <p>The Pandemic 2009 influenza virus found in Nepal were antigenically and genetically related to the novel A/CALIFORNIA/07/2009-LIKE (H1N1)v type.</p

Down-regulation of estrogen receptor-alpha and rearranged during transfection tyrosine kinase is associated with withaferin a-induced apoptosis in MCF-7 breast cancer cells

<p>Abstract</p> <p>Background</p> <p>Withaferin A (WA), a naturally occurring withanolide, induces apoptosis in both estrogen-responsive MCF-7 and estrogen-independent MDA-MB-231 breast cancer cell lines with higher sensitivity in MCF-7 cells, but the underlying mechanisms are not well defined. The purpose of this study was to determine the anti-cancer effects of WA in MCF-7 breast cancer cells and explore alterations in estrogen receptor alpha (ERα) and its associated molecules <it>in vitro </it>as novel mechanisms of WA action.</p> <p>Methods</p> <p>The effects of WA on MCF-7 viability and proliferation were evaluated by 3-(4, 5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay and trypan blue exclusion assays. Apoptosis was evaluated by Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) flow cytometry and Western blot analysis of poly (ADP-ribose) polymerase (PARP) cleavage. Cell cycle effects were analyzed by PI flow cytometry. Western blotting was also conducted to examine alterations in the expression of ERα and pathways that are associated with ERα function.</p> <p>Results</p> <p>WA resulted in growth inhibition and decreased viability in MCF-7 cells with an IC50 of 576 nM for 72 h. It also caused a dose- and time-dependent apoptosis and G2/M cell cycle arrest. WA-induced apoptosis was associated with down-regulation of ERα, REarranged during Transfection (RET) tyrosine kinase, and heat shock factor-1 (HSF1), as well as up-regulation of phosphorylated p38 mitogen-activated protein kinase (phospho-p38 MAPK), p53 and p21 protein expression. Co-treatment with protein synthesis inhibitor cycloheximide or proteasome inhibitor MG132 revealed that depletion of ERα by WA is post-translational, due to proteasome-dependent ERα degradation.</p> <p>Conclusions</p> <p>Taken together, down-regulation of ERα, RET, HSF1 and up-regulation of phospho-p38 MAPK, p53, p21 are involved in the pro-apoptotic and growth-inhibitory effects of WA in MCF-7 breast cancer cells <it>in vitro</it>. Down-regulation of ERα protein levels by WA is caused by proteasome-dependent ERα degradation.</p

Short and long term treatment of asthma with intravenous nutrients

BACKGROUND: Asthma is an increasing problem in this country and others. Although medications for the treatment of asthma abound and are improving, there are inherent risks and side effects with all of them. Intravenous magnesium has been employed in the treatment of acute asthma, but its use has not become universal, nor has it been studied for the treatment of chronic asthma. It is known to be a safe drug with minimal side effects. In this study, the author investigates the use of magnesium and other nutrients in the treatment of both acute and chronic asthma. METHODS: In this non-blinded outcome study, following informed consent, forty-three (43) randomly selected volunteer patients with both acute and chronic asthma were treated with IV infusions described herein. All patients were observed with spirometry 10 minutes post-infusion; two sub-groups of patients were also observed after multiple infusions over a short period of time (less than one month) and a longer period of time (average 5.8 months). Pulmonary function was analyzed by spirometric testing with pre- and post-infusion spirometric measurements with the pre/post group. For longer term (Trend) patients, baseline spirometry measurements were compared to spirometry measurements after patients had received multiple infusions over a period of time. Eight (8) patients were measured for both pre/post and Trend data. RESULTS: The 38 pre-infusion/post-infusion patients with acute and chronic asthma demonstrated an overall average improvement (percentage improvement in percent predicted) of 45%. The 13 patients measured for improvement over time (Trend data, average duration 5.82 months), demonstrated an overall average improvement (percentage improvement in percent predicted) of 57%. Of the 13 patients in the multiple infusion group, 9 patients who received longer-term therapy (average duration of 12.58 months) for chronic asthma demonstrated an overall average improvement of 95% (percentage improvement in percent predicted). CONCLUSION: The use of intravenous treatment with multiple nutrients, including magnesium, for acute and chronic asthma may be of considerable benefit. Pulmonary function improved progressively the longer patients received treatment

Dengue Virus Activates Polyreactive, Natural IgG B Cells after Primary and Secondary Infection

BACKGROUND: Dengue virus is transmitted by mosquitoes and has four serotypes. Cross-protection to other serotypes lasting for a few months is observed following infection with one serotype. There is evidence that low-affinity T and/or B cells from primary infections contribute to the severe syndromes often associated with secondary dengue infections. such pronounced immune-mediated enhancement suggests a dengue-specific pattern of immune cell activation. This study investigates the acute and early convalescent B cell response leading to the generation of cross-reactive and neutralizing antibodies following dengue infection. METHODOLOGY/PRINCIPAL FINDINGS: We assayed blood samples taken from dengue patients with primary or secondary infection during acute disease and convalescence and compared them to samples from patients presenting with non-dengue related fever. Dengue induced massive early plasmablast formation, which correlated with the appearance of polyclonal, cross-reactive IgG for both primary and secondary infection. Surprisingly, the contribution of IgG to the neutralizing titer 4-7 days after fever onset was more than 50% even after primary infection. CONCLUSIONS/SIGNIFICANCE: Poly-reactive and virus serotype cross-reactive IgG are an important component of the innate response in humans during both primary and secondary dengue infection, and "innate specificities" seem to constitute part of the adaptive response in dengue. While of potential importance for protection during secondary infection, cross-reactive B cells will also compete with highly neutralizing B cells and possibly interfere with their development

Evolutionary Sequence Modeling for Discovery of Peptide Hormones

There are currently a large number of “orphan” G-protein-coupled receptors (GPCRs) whose endogenous ligands (peptide hormones) are unknown. Identification of these peptide hormones is a difficult and important problem. We describe a computational framework that models spatial structure along the genomic sequence simultaneously with the temporal evolutionary path structure across species and show how such models can be used to discover new functional molecules, in particular peptide hormones, via cross-genomic sequence comparisons. The computational framework incorporates a priori high-level knowledge of structural and evolutionary constraints into a hierarchical grammar of evolutionary probabilistic models. This computational method was used for identifying novel prohormones and the processed peptide sites by producing sequence alignments across many species at the functional-element level. Experimental results with an initial implementation of the algorithm were used to identify potential prohormones by comparing the human and non-human proteins in the Swiss-Prot database of known annotated proteins. In this proof of concept, we identified 45 out of 54 prohormones with only 44 false positives. The comparison of known and hypothetical human and mouse proteins resulted in the identification of a novel putative prohormone with at least four potential neuropeptides. Finally, in order to validate the computational methodology, we present the basic molecular biological characterization of the novel putative peptide hormone, including its identification and regional localization in the brain. This species comparison, HMM-based computational approach succeeded in identifying a previously undiscovered neuropeptide from whole genome protein sequences. This novel putative peptide hormone is found in discreet brain regions as well as other organs. The success of this approach will have a great impact on our understanding of GPCRs and associated pathways and help to identify new targets for drug development

Malaria vaccines and their potential role in the elimination of malaria

Research on malaria vaccines is currently directed primarily towards the development of vaccines that prevent clinical malaria. Malaria elimination, now being considered seriously in some epidemiological situations, requires a different vaccine strategy, since success will depend on killing all parasites in the community in order to stop transmission completely

Differential transcriptomic profiles effected by oil palm phenolics indicate novel health outcomes

Abstract Background Plant phenolics are important nutritional antioxidants which could aid in overcoming chronic diseases such as cardiovascular disease and cancer, two leading causes of death in the world. The oil palm (Elaeis guineensis) is a rich source of water-soluble phenolics which have high antioxidant activities. This study aimed to identify the in vivo effects and molecular mechanisms involved in the biological activities of oil palm phenolics (OPP) during healthy states via microarray gene expression profiling, using mice supplemented with a normal diet as biological models. Results Having confirmed via histology, haematology and clinical biochemistry analyses that OPP is not toxic to mice, we further explored the gene expression changes caused by OPP through statistical and functional analyses using Illumina microarrays. OPP showed numerous biological activities in three major organs of mice, the liver, spleen and heart. In livers of mice given OPP, four lipid catabolism genes were up-regulated while five cholesterol biosynthesis genes were down-regulated, suggesting that OPP may play a role in reducing cardiovascular disease. OPP also up-regulated eighteen blood coagulation genes in spleens of mice. OPP elicited gene expression changes similar to the effects of caloric restriction in the hearts of mice supplemented with OPP. Microarray gene expression fold changes for six target genes in the three major organs tested were validated with real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR), and the correlation of fold changes obtained with these two techniques was high (R2 = 0.9653). Conclusions OPP showed non-toxicity and various pleiotropic effects in mice. This study implies the potential application of OPP as a valuable source of wellness nutraceuticals, and further suggests the molecular mechanisms as to how dietary phenolics work in vivo.</p