We have developed a PCR assay that can detect a single Epstein-Barr virus (EBV) genome in the presence of 10(6) uninfected cells. Using this assay, we demonstrate that EBV persists, in the peripheral blood of all seropositive individuals tested, in CD19+, CD23-, and CD80 (B7)- B cells. We further show that the virus in these cells is latent, but readily reactivated to produce infectious immortalizing virus; therefore, these cells represent a true site of latent persistence. EBV was not significantly detected in monocytes or T cells. The frequency of infected cells in nine healthy donors varied from 23 to 625 per 10(7) B cells, but was relatively stable for each individual over the course of 2 years. We conclude that the EBV-infected cells in vivo are B cells with a nonactivated phenotype. This represents a novel form of latency in normal B cells.

We have measured the absolute numbers of EBV-infected B cells in the peripheral blood of healthy persistently infected individuals. Single measurements on a panel of 15 healthy individuals demonstrate that the frequency varies over a wide range from 1-50 per 10(6) B cells. Repeat measurements over 1-3.5 years on several individuals whose frequencies varied over a 10-fold range showed that the variation does not represent the fluctuation in the frequency that can occur within an individual; rather, the frequencies are specific to the individual. The frequency within an individual measured over time is stable and contributes less than 10% to the variance seen in the whole population. These measurements suggest that the level of EBV-infected B cells is tightly regulated and we propose that the same homeostatic mechanisms that regulate the levels of normal B cells also regulate B cells latently infected with EBV.

BackgroundDysfunctions of long non-coding RNA (lncRNAs) have been associated with the initiation and progression of hepatocellular carcinoma (HCC), but the clinicopathologic significance and potential role of lncRNA PTTG3P (pituitary tumor-transforming 3, pseudogene) in HCC remains largely unknown.MethodsWe compared the expression profiles of lncRNAs in 3 HCC tumor tissues and adjacent non-tumor tissues by microarrays. In situ hybridization (ISH) and quantitative real-time polymerase chain reaction (qRT-PCR) were applied to assess the level of PTTG3P and prognostic values of PTTG3P were assayed in two HCC cohorts (n = 46 and 90). Artificial modulation of PTTG3P (down- and over-expression) was performed to explore the role of PTTG3P in tumor growth and metastasis in vitro and in vivo. Involvement of PTTG1 (pituitary tumor-transforming 1), PI3K/AKT signaling and its downstream signals were validated by qRT-PCR and western blot.ResultsWe found that PTTG3P was frequently up-regulated in HCC and its level was positively correlated to tumor size, TNM stage and poor survival of patients with HCC. Enforced expression of PTTG3P significantly promoted cell proliferation, migration, and invasion in vitro, as well as tumorigenesis and metastasis in vivo. Conversely, PTTG3P knockdown had opposite effects. Mechanistically, over-expression of PTTG3P up-regulated PTTG1, activated PI3K/AKT signaling and its downstream signals including cell cycle progression, cell apoptosis and epithelial-mesenchymal transition (EMT)-associated genes.ConclusionsOur findings suggest that PTTG3P, a valuable marker of HCC prognosis, promotes tumor growth and metastasis via up-regulating PTTG1 and activating PI3K/AKT signaling in HCC and might represent a potential target for gene-based therapy.Electronic supplementary materialThe online version of this article (10.1186/s12943-018-0841-x) contains supplementary material, which is available to authorized users.

Many types of fluorescent sensing systems have been reported for biological small molecules. Particularly, several methods have been developed for the recognition of ATP or NAD+, but they only show moderate sensitivity, and they cannot discriminate either ATP or NAD+ from their respective analogues. We have addressed these limitations and report here a dual strategy which combines split DNAzyme-based background reduction with catalytic and molecular beacon (CAMB)-based amplified detection to develop a ligation-triggered DNAzyme cascade, resulting in ultrahigh sensitivity. First, the 8–17 DNAzyme is split into two separate oligonucleotide fragments as the building blocks for the DNA ligation reaction, thereby providing a zero-background signal to improve overall sensitivity. Next, a CAMB strategy is further employed for amplified signal detection achieved through cycling and regenerating the DNAzyme to realize the true enzymatic multiple turnover (one enzyme catalyzes the cleavage of several substrates) of catalytic beacons. This combination of zero-background signal and signal amplification significantly improves the sensitivity of the sensing systems, resulting in detection limits of 100 and 50 pM for ATP and NAD+, respectively, much lower than those of previously reported biosensors. Moreover, by taking advantage of the highly specific biomolecule-dependence of the DNA ligation reaction, the developed DNAzyme cascades show significantly high selectivity toward the target cofactor (ATP or NAD+), and the target biological small molecule can be distinguished from its analogues. Therefore, as a new and universal platform for the design of DNA ligation reaction-based sensing systems, this novel ligation-triggered DNAzyme cascade method may find a broad spectrum of applications in both environmental and biomedical fields.

Interstitial fluid (ISF), as an emerging source of biomarkers, is unmistakably significant for disease diagnosis. Microneedles (MNs) provide a minimally invasive approach for extracting the desired molecules from ISF. However, existing MNs are limited by their capture efficiency and sensitivity, which impedes early disease diagnosis. Herein, an engineered wearable epidermal system is presented with a combination of reverse iontophoresis and MNs for rapid capture and sensing of Epstein‐Barr virus cell‐free DNA (an important biomarker of nasopharyngeal carcinoma). Owing to a dual‐extraction effect demonstrated by reverse iontophoresis and MNs, the engineered wearable platform successfully isolates the cell‐free DNA target from ISF within 10 min, with a threshold of 5 copies per µL and a maximum capture efficiency of 95.4%. The captured cell‐free DNA is also directly used in a recombinase polymerase amplification electrochemical microfluidic biosensor with a detection limit of 1.1 copies per µL (or a single copy). The experimental data from immunodeficient mouse models rationalizes the feasibility and practicality of the wearable system. Collectively, the developed method opens an innovative route for minimally invasive sampling of ISF for cell‐free DNA‐related cancer screening and prognosis.

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The information regarding the effect of hepatitis B virus (HBV) infection on gut microbiota and the relationship between gut microbiota dysbiosis and hepatitis B virus-induced chronic liver disease (HBVCLD) is limited. In this study, we aimed at characterizing the gut microbiota composition in the three different stages of hepatitis B virus-induced chronic liver disease patients and healthy individuals. Faecal samples and clinical data were collected from HBVCLD patients and healthy individuals.The 16S rDNA gene amplification products were sequenced. Bioinformatic analysis including alpha diversity and PICRUSt was performed. A total of 19 phyla, 43 classes, 72 orders, 126 families and 225 genera were detected. The beta-diversity showed a separate clustering of healthy controls and HBVCLD patients covering chronic hepatitis (CHB), liver cirrhosis (LC) and hepatocellular carcinoma (HCC); and gut microbiota of healthy controls was more consistent, whereas those of CHB, LC and HCC varied substantially. The abundance of Firmicutes was lower, and Bacteroidetes was higher in patients with CHB, LC and HCC than in healthy controls. Predicted metagenomics of microbial communities showed an increase in glycan biosynthesis and metabolism-related genes and lipid metabolism-related genes in HBVCLD than in healthy individuals. Our study suggested that HBVCLD is associated with gut dysbiosis, with characteristics including, a gain in potential bacteria and a loss in potential beneficial bacteria or genes. Further study of CHB, LC and HCC based on microbiota may provide a novel insight into the pathogenesis of HBVCLD as well as a novel treatment strategy. K E Y W O R D S16S rDNA, dysbiosis, Gut Microbiota, hepatitis B virus, progression

Exosomes, known as nanometer‐sized vesicles (30–200 nm), are secreted by many types of cells. Cancer‐derived exosomes have great potential to be biomarkers for early clinical diagnosis and evaluation of cancer therapeutic efficacy. Conventional detection methods are limited to low sensitivity and reproducibility. There are hundreds of papers published with different detection methods in recent years to address these challenges. Therefore, in this review, pioneering researches about various detection strategies are comprehensively summarized and the analytical performance of these tests is evaluated. Furthermore, the exosome molecular composition (protein and nucleic acid) profiling, a single exosome profiling, and their application in clinical cancer diagnosis are reviewed. Finally, the principles and applications of machine learning method in exosomes researches are presented.