LncRNA‐miRNA network analysis across the Th17 cell line reveals biomarker potency of lncRNA NEAT1 and KCNQ1OT1 in multiple sclerosis

Differentiation of CD⁴⁺ T cells into Th17 cells is an important factor in the onset and progression of multiple sclerosis (MS) and Th17/Treg imbalance. Little is known about the role of lncRNAs in the differentiation of CD⁴⁺ cells from Th17 cells. This study aimed to analyse the lncRNA‐miRNAs network involved in MS disease and its role in the differentiation of Th17 cells. The lncRNAs in Th17 differentiation were obtained from {"type":"entrez-geo","attrs":{"text":"GSE66261","term_id":"66261"}}GSE66261 using the GEO datasets. Differential expression of lncRNAs in Th17 primary cells compared to Th17 effector cells was investigated by RNA‐seq analysis. Next, the most highlighted lncRNAs in autoimmune diseases were downloaded from the lncRNAs disease database, and the most critical miRNA was extracted by literature search. Then, the lncRNA‐miRNA interaction was achieved by the Starbase database, and the ceRNA network was designed by Cytoscape. Finally, using the CytoHubba application, two hub lncRNAs with the most interactions with miRNAs were identified by the MCODE plug‐in. The expression level of genes was measured by qPCR, and the plasma level of cytokines was analysed by ELISA kits. The results showed an increase in the expression of NEAT1, KCNQ1OT1 and RORC and a decrease in the expression of FOXP3. In plasma, an upregulation of IL17 and a downregulation of TGFB inflammatory cytokines were detected. The dysregulated expression of these genes could be attributed to relapsing‐remitting MS (RR‐MS) patients and help us understand MS pathogenesis better.     

Light producing reporter plasmids for Escherichia coli K-12 that can be integrated into the chromosome

Plasmid vectors using the Photorhabdus luminescenslux operon can be used for real time measurements of promoter activity. We have generated a series of lux vectors that have a conditional origin of replication, different selectable markers and the attP sequence from λ. Single copies of these plasmids can be integrated into the λ attachment site in the Escherichia coli chromosome. We constructed reporter derivatives and compared light production when the plasmids were present in multiple copies and in single copies. We also demonstrated that expression could be induced under the appropriate conditions.     

An ensemble approach to accurately detect somatic mutations using SomaticSeq

SomaticSeq is an accurate somatic mutation detection pipeline implementing a stochastic boosting algorithm to produce highly accurate somatic mutation calls for both single nucleotide variants and small insertions and deletions. The workflow currently incorporates five state-of-the-art somatic mutation callers, and extracts over 70 individual genomic and sequencing features for each candidate site. A training set is provided to an adaptively boosted decision tree learner to create a classifier for predicting mutation statuses. We validate our results with both synthetic and real data. We report that SomaticSeq is able to achieve better overall accuracy than any individual tool incorporated.

Electronic supplementary material

The online version of this article (doi:10.1186/s13059-015-0758-2) contains supplementary material, which is available to authorized users.     

A hidden Markov model reliably characterizes ketamine-induced spectral dynamics in macaque local field potentials and human electroencephalograms

Ketamine is an NMDA receptor antagonist commonly used to maintain general anesthesia. At anesthetic doses, ketamine causes high power gamma (25-50 Hz) oscillations alternating with slow-delta (0.1-4 Hz) oscillations. These dynamics are readily observed in local field potentials (LFPs) of non-human primates (NHPs) and electroencephalogram (EEG) recordings from human subjects. However, a detailed statistical analysis of these dynamics has not been reported. We characterize ketamine’s neural dynamics using a hidden Markov model (HMM). The HMM observations are sequences of spectral power in seven canonical frequency bands between 0 to 50 Hz, where power is averaged within each band and scaled between 0 and 1. We model the observations as realizations of multivariate beta probability distributions that depend on a discrete-valued latent state process whose state transitions obey Markov dynamics. Using an expectation-maximization algorithm, we fit this beta-HMM to LFP recordings from 2 NHPs, and separately, to EEG recordings from 9 human subjects who received anesthetic doses of ketamine. Our beta-HMM framework provides a useful tool for experimental data analysis. Together, the estimated beta-HMM parameters and optimal state trajectory revealed an alternating pattern of states characterized primarily by gamma and slow-delta activities. The mean duration of the gamma activity was 2.2s([1.7,2.8]s) and 1.2s([0.9,1.5]s) for the two NHPs, and 2.5s([1.7,3.6]s) for the human subjects. The mean duration of the slow-delta activity was 1.6s([1.2,2.0]s) and 1.0s([0.8,1.2]s) for the two NHPs, and 1.8s([1.3,2.4]s) for the human subjects. Our characterizations of the alternating gamma slow-delta activities revealed five sub-states that show regular sequential transitions. These quantitative insights can inform the development of rhythm-generating neuronal circuit models that give mechanistic insights into this phenomenon and how ketamine produces altered states of arousal.     

In-solution staining and arraying method for the immunofluorescence detection of γH2AX foci optimized for clinical applications

Immunofluorescence quantification of γH2AX foci is a powerful approach to quantify DNA double-strand breaks induced by cancer therapy or accidental exposure to ionizing radiation. Here we report a modification to the γH2AX immunofluorescence labeling method, whereby cells are stained in-solution before being spotted and fixed onto microscope slides. Our modified method allows arraying of 16 patient samples/slide ready for foci counting in 2 h and demonstrated reliably detection of γH2AX foci in mononuclear cells prepared from patients who had undergone radiation therapy.