Background: Broad-range PCR provides valuable information for detecting bacterial infections. This study assesses the combined use of broad-range real-time PCR and high-resolution melting analysis for rapid detection and identification of clinically important bacteria. Methods: We subjected 46 bacterial culture colonies representing 25 clinically important bacterial species to LightCycler real-time PCR amplification of the 16S rRNA gene in the presence of LCGreen I fluorescent dye. We performed high-resolution melting analysis of the PCR products with the HR-1 instrument and used melting profiles as molecular fingerprints for bacterial species identification. We validated this method via assessment of 54 consecutive bacteria culture colonies obtained from a clinical microbiology laboratory. Results: The 16S rRNA gene of all 25 bacterial species was amplifiable by this method, with PCR product
In studies of the phosphorylated proteins in rat liver and Walker-256, it was established that the ratio of various fractions of P-N linkages to P-O linkages varies from 0.6 to 3.1. In rat regenerating liver nuclei, the ratio of P-N and P-O varies with time after partial hepatectomy. Using [3H]-lysine and 32Pi, it is shown that phosphoryllysine forms in some new and, presumably, some preexisting H1 molecules. Using [3H]histidine and 32Pi, it is shown that phosphohistidine forms exclusively in preexisting H4. The half-life of H4 phosphohistidine appears to be about 2 h.
Valproic acid (VPA), with inhibition activity mainly toward histone deacetylase (HDAC) and Glycogen Synthase Kinase (GSK)-3, and lithium, with inhibition activity mainly toward GSK-3, are both prescribed in clinical as mood-stabilizers and anticonvulsants for the control of bipolar disorder. This study aims to compare the immuno-modulation activities of VPA and lithium, especially on the differentiation and functions of dendritic cells (DC). Our data show that treatment with VPA or lithium effectively alleviated the severity of collagen-induced arthritis triggered by LPS in mice. Both agents reduced the serum level of IL-6 and IL-10 after LPS challenge in mice. VPA and lithium both induce significant down-regulation of group I CD1 expression and secretion of IL-6 during differentiation of human monocyte-derived immature DC, while they differ in the induction of CD83 and CD86 expression, secretion of IL-8, IL-10, and TNF-α. Upon stimulation of immature DC with LPS, VPA, and lithium both reduced the secretion of IL-6 and TNF-α. However, only lithium significantly increased the production of IL-10, while VPA increased the production of IL-8 but substantially reduce the secretion of IL-10 and IL-23. Treatment with VPA resulted in a reduced capacity of LPS-stimulated DC to promote the differentiation of T helper 17 cells that are critical in the promotion of inflammatory responses. Taken together, our results suggest that VPA and lithium may differentially modulate inflammation through regulating the capacity of DC to mediate distinct T cell responses, and they may provide a complementary immunomodulatory effects for the treatment of inflammation-related diseases. J. Cell. Physiol. 232: 1176-1186, 2017. © 2016 Wiley Periodicals, Inc.
BackgroundIn view of the limited knowledge of plasma biomarkers relating to cancer resistance to radiotherapy, we have set up screening, training and testing stages to investigate the microRNAs (miRNAs) expression profile in plasma to predict between the poor responsive and responsive groups after 6 months of radiotherapy.MethodsPlasma was collected prior to and after radiotherapy, and the microRNA profiles were analyzed by quantitative reverse transcription polymerase chain reaction (qRT-PCR) arrays. Candidate miRNAs were validated by single qRT-PCR assays from the training and testing set. The classifier for ancillary prognosis was developed by multiple logistic regression analysis to correlate the ratios of miRNAs expression levels with clinical data.ResultsWe revealed that eight miRNAs expressions had significant changes after radiotherapy and the expression levels of miR-374a-5p, miR-342-5p and miR-519d-3p showed significant differences between the responsive and poor responsive groups in the pre-radiotherapy samples. The Kaplan–Meier curve analysis also showed that low miR-342-5p and miR-519d-3p expressions were associated with worse prognosis. Our results revealed two miRNA classifiers from the pre- and post-radiotherapy samples to predict radiotherapy response with area under curve values of 0.8923 and 0.9405.ConclusionsThe expression levels of miR-374a-5p, miR-342-5p and miR-519d-3p in plasma are associated with radiotherapy responses. Two miRNA classifiers could be developed as a potential non-invasive ancillary tool for predicting patient response to radiotherapy.Electronic supplementary materialThe online version of this article (10.1186/s12967-018-1711-4) contains supplementary material, which is available to authorized users.
Traditional, horse-derived antivenin is currently the most efficient treatment against snake bites. However, it is costly and has unpredictable side effects. Thus, alternative, cost-effective strategies for producing antivenin are needed. In this study, we immunized hens with inactivated NNA venom proteins from the cobra Naja naja atra (NNA). Purified yolk IgY antibodies showed specific anti-NNA binding activity comparable to that of the equine-derived antivenin. We used phage display technology to generate two antibody libraries containing 9.0 × 108 and 8.4 × 108 clones with a short or long linker, respectively. The phage ELISA indicated that anti-NNA clones displaying single-chain variable fragments (scFv) were significantly enriched after biopanning. The nucleotide sequences of the light and heavy chain genes of 30 monoclonal scFv antibodies were determined and classified into six groups with the short linker and nine groups with the long linker. These scFv clones specifically bound to NNA proteins but not to venom proteins from other snakes. Their binding affinities were further determined by competitive ELISA. Animal model studies showed that anti-NNA IgY antibodies exhibited complete protective effects, while a combination of scFv antibodies raised the survival rates and times of mice challenged with lethal doses of NNA venom proteins.
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