In silico experiment with an-antigen-toll like receptor-5 agonist fusion construct for immunogenic application to Helicobacter pylori

BACKGROUNDS:

Helicobacter pylori colonize the gastric mucosa of half of the world''s population. Although it is classified as a definitive type I carcinogen by World Health Organization, there is no effective vaccine against this bacterium. H. pylori evade the host immune response by avoiding toll-like detection, such as detection via toll-like receptor-5 (TLR-5). Thus, a chimeric construct consisting of selected epitopes from virulence factors that is incorporated into a TLR-5 ligand (Pseudomonas flagellin) could result in more potent innate and adaptive immune responses.

MATERIALS AND METHODS:

Based on the histocompatibility antigens of BALB/c mice, in silico techniques were used to select several fragments from H. pylori virulence factors with a high density of B- and T-cell epitopes.

RESULTS:

These segments consist of cytotoxin-associated geneA (residue 162-283), neutrophil activating protein (residue 30-135) and outer inflammatory protein A (residue 155-268). The secondary and tertiary structure of the chimeric constructs and other bioinformatics analyses such as stability, solubility, and antigenicity were performed. The chimeric construct containing antigenic segments of H. pylori proteins was fused with the D3 domain of Pseudomonas flagellin. This recombinant chimeric gene was optimized for expression in Escherichia coli. The in silico results showed that the conserved C- and N-terminal domains of flagellin and the antigenicity of selected fragments were retained.

DISCUSSION:

In silico analysis showed that Pseudomonas flagellin is a suitable platform for incorporation of an antigenic construct from H. pylori. This strategy may be an effective tool for the control of H. pylori and other persistent infections.     

Analysis of microRNA signatures using size-coded ligation-mediated PCR

The expression pattern and regulatory functions of microRNAs (miRNAs) are intensively investigated in various tissues, cell types and disorders. Differential miRNA expression signatures have been revealed in healthy and unhealthy tissues using high-throughput profiling methods. For further analyses of miRNA signatures in biological samples, we describe here a simple and efficient method to detect multiple miRNAs simultaneously in total RNA. The size-coded ligation-mediated polymerase chain reaction (SL-PCR) method is based on size-coded DNA probe hybridization in solution, followed-by ligation, PCR amplification and gel fractionation. The new method shows quantitative and specific detection of miRNAs. We profiled miRNAs of the let-7 family in a number of organisms, tissues and cell types and the results correspond with their incidence in the genome and reported expression levels. Finally, SL-PCR detected let-7 expression changes in human embryonic stem cells as they differentiate to neuron and also in young and aged mice brain and bone marrow. We conclude that the method can efficiently reveal miRNA signatures in a range of biological samples.     

Protein,fatty acid,and pigment content of <Emphasis Type="Italic">Chlorella vulgaris</Emphasis> under different light regimes

The effects of irradiance and photoperiod on growth rates, chlorophyll a, β-carotene, total protein, and fatty acid content of Chlorella vulgaris were determined. The maximum growth rate (1.13 day⁻¹) was at 100 μmol photons m⁻² s⁻¹ and 16:8-h light/dark photoperiod. Chlorophyll a and β-carotene contents significantly differed under different light regimes with chlorophyll a content lower at high irradiance and longer light duration, while β-carotene showed the inverse trend. The total protein and fatty acid content also significantly differed in different light regimes; the maximum percentage of protein (46%) was at 100 μmol photons m⁻² s⁻¹ and 16:8 h photoperiod, and minimum (33%) was at 37.5 μmol photons m⁻² s⁻¹ and 8:16 h photoperiod; the total saturated fatty acids increased, while monounsaturated and polyunsaturated fatty acids decreased with increasing irradiance and light duration.     

Effect of general practitioner education on adherence to antihypertensive drugs: cluster randomised controlled trial

Objective To determine the impact of a simple educational package for general practitioners on adherence to antihypertensive drugs.Design Cluster randomised controlled trial.Setting Six randomly selected communities in Karachi, Pakistan.Participants 200 patients with hypertension taking antihypertensive drugs; 78 general practitioners.Intervention Care by general practitioners specially trained in management of hypertension compared with usual care.Main outcome measure Correct dosing, defined as percentage of prescribed doses taken, measured with electronic medication event monitoring system (MEMS) bottle.Results 200 patients were enrolled, and 178 (89%) successfully completed six weeks of follow-up. Adherence was significantly greater in the special care group than in the usual care group (unadjusted mean percentage days with correct dose 48.1%, 95% confidence interval 35.8% to 60.4%, versus 32.4%, 22.6% to 42.3%; P=0.048). Adherence was also higher among patients who had higher levels of education (P<0.001), were encouraged by family members (P<0.001), believed in the effect of drugs (P<0.001), and had the purpose of the drugs explained to them (P<0.001).Conclusions Special training of general practitioners in management of hypertension, emphasising good communication between doctors and patients, is more effective than usual care provided in the communities in Karachi. Such simple interventions should be adopted by other developing countries that are now facing an increasing burden of hypertension.Trial registration Clinical trials {"type":"clinical-trial","attrs":{"text":"NCT00330408","term_id":"NCT00330408"}}NCT00330408.     

The immunomodulatory role of probiotics in allergy therapy

The increased incidence of allergic disorders may be the result of a relative fall in microbial induction in the intestinal immune system during infancy and early childhood. Probiotics have recently been proposed as viable microorganisms for the prevention and treatment of specific allergic diseases. Different mechanisms have been considered for this probiotic property, such as generation of cytokines from activated pro-T-helper type 1 after bacterial contact. However, the effects of its immunomodulatory potential require validation for clinical applications. This review will focus on the currently available data on the benefits of probiotics in allergy disease.     

Transplantation of miR-219 overexpressed human endometrial stem cells encapsulated in fibrin hydrogel in spinal cord injury

Oligodendrocyte (OL) loss and demyelination occur after spinal cord injury (SCI). Stimulation of remyelination through transplantation of myelinating cells may be effective in improving function. For the repair strategy to be successful, the selection of a suitable cell and maintaining cell growth when cells are injected directly to the site of injury is important. In addition to selecting the type of cell, fibrin hydrogel was used as a suitable tissue engineering scaffold for this purpose. To test the relationship between myelination and functional improvement, the human endometrial stem cells (hEnSCs) were differentiated toward oligodendrocyte progenitor cells (OPCs) using overexpression of miR-219. Adult female Wistar rats were used to induce SCI by using a compression model and were randomly assigned to the following four experimental groups: SCI, Vehicle, hEnSC, and OPC. Ten days after injury, miR-219 overexpressed hEnSC-derived OPCs encapsulated in fibrin hydrogel, as an injectable scaffold, were injected to the injury site. In this study, with a focus on promoting functional recovery after SCI, the Basso-Beattie-Bresnahan test was performed to evaluate the recovery of motor function every week for 10 weeks and the histological assay was then performed. Results showed that the rate of motor function recovery was significantly higher in OPC group compared to SCI and vehicle groups but no marked differences were found between OPC and hEnSC groups, although, the rate of myelination in the OPC group was significantly higher than the other groups. These results demonstrated that remyelination was not the cause of recovery of motor function.     

The deubiquitinating gene Usp29 is dispensable for fertility in male mice

The balanced actions between ubiquitination and deubiquitination precisely control the levels of various proteins vital for spermatogenesis. Ubiquitin-specific processing proteases(USPs) are the largest family of deubiquitinatingenzymes(DUBs),containing more than 50 members. So far, the functions of only a few USPs in male fertility have been studied, the roles of the majority are yet unknown. The present study aimed to explore the function of Usp29(ubiquitin-specific protease 29) in male fertility. We found that Usp29 showed predominant expression in mouse testis, and its m RNA expression started to increase at 14 days postpartum(dpp), with a peak at 28 and 35 dpp. Using CRISPR/Cas9 technology, we generated Usp29 knockout mice(Usp29~(–/–)). Usp29~(–/–)mice exhibited no overt developmental anomalies. Further examination revealed that Usp29~(–/–)mice had normal fertility and showed no detectable difference in the testis/body weight ratio, testicular and epididymal histology as well as epididymal sperm count from the wild-type littermates. Moreover, Usp29 is not a pseudogene in mice. Taken together, our study first reported that though Usp29 is predominantly expressed in the testis, it is not essential for male fertility in mice.