The advance of next generation sequencing (NGS) techniques provides an unprecedented opportunity to probe the enormous diversity of the immune repertoire by deep sequencing T-cell receptors (TCRs) and B-cell receptors (BCRs). However, an efficient and accurate analytical tool is still on demand to process the huge amount of data. We have developed a high-resolution analytical pipeline, Immune Monitor ("IMonitor") to tackle this task. This method utilizes realignment to identify V(D)J genes and alleles after common local alignment. We compare IMonitor with other published tools by simulated and public rearranged sequences, and it demonstrates its superior performance in most aspects. Together with this, a methodology is developed to correct the PCR and sequencing errors and to minimize the PCR bias among various rearranged sequences with different V and J gene families. IMonitor provides general adaptation for sequences from all receptor chains of different species and outputs useful statistics and visualizations. In the final part of this article, we demonstrate its application on minimal residual disease detection in patients with B-cell acute lymphoblastic leukemia. In summary, this package would be of widespread usage for immune repertoire analysis.KEYWORDS next generation sequencing; bioinformatics; immune repertoire; TCR/BCR T HE diversity of T-cell receptors (TCRs), B-cell receptors (BCRs), and secreting form antibodies makes up the core of the complicated immune system and serves as pivotal defensive components to protect the body against invading virus, bacteria, and other pathogens. The TCR consists of a heterodimeric ab chain (95%, TRA, TRB) or gd chain (5%), while the BCR is assembled with two heavy chains (IGH) and two light chains (IGK or IGL). Structurally, each chain can be divided into the variable domain and the constant domain (Lefranc and Lefranc 2001a,b). The diversity of the TCR and BCR repertoire is enormous, owing to the process of V(D)J gene rearrangement, random deletion of germline nucleotides, and insertion of uncertain length of nontemplate nucleotides between V-D and D-J junctions (TRB, IGH) or V-J junctions (TRA, IGK, IGL). In humans, it has been estimated theoretically that the diversity of TCR-ab receptors exceeds 10 18 in the thymus, and the diversity of the B-cell repertoire is even larger, considering the somatic hypermutation (Janeway 2005;Benichou et al. 2012). The T-and B-cell repertoire could undergo dynamic changes under different phenotypic status. Recently, deep sequencing enabled by different platforms including Roche 454 and Illumina Hiseq (Freeman et al. 2009;Robins et al. 2009;Wang et al. 2010;Fischer 2011;Venturi et al. 2011) has been applied to unravel the dynamics of the TCR and BCR repertoire and extended to various translational applications such as vaccination, cancer, and autoimmune diseases.Several tools and software have been developed for TCR and BCR sequence analysis, including iHMMune-align (Gaeta et al. 2007), HighV-QEUST (Li et al. 2013), IgBLA...

Background Since the start of the COVID-19 outbreak, a large number of COVID-19-related papers have been published. However, concerns about the risk of expedited science have been raised. We aimed at reviewing and categorizing COVID-19-related medical research and to critically appraise peer-reviewed original articles. Methods The data sources were Pubmed, Cochrane COVID-19 register study, arXiv, medRxiv and bioRxiv, from 01/11/2019 to 01/05/2020. Peer-reviewed and preprints publications related to COVID-19 were included, written in English or Chinese. No limitations were placed on study design. Reviewers screened and categorized studies according to i) publication type, ii) country of publication, and iii) topics covered. Original articles were critically appraised using validated quality assessment tools. Results Among the 11,452 publications identified, 10,516 met the inclusion criteria, among which 7468 (71.0%) were peer-reviewed articles. Among these, 4190 publications (56.1%) did not include any data or analytics (comprising expert opinion pieces). Overall, the most represented topics were infectious disease (n = 2326, 22.1%), epidemiology (n = 1802, 17.1%), and global health (n = 1602, 15.2%). The top five publishing countries were China (25.8%), United States (22.3%), United Kingdom (8.8%), Italy (8.1%) and India (3.4%). The dynamic of publication showed that the exponential growth of COVID-19 peer-reviewed articles was mainly driven by publications without original data (mean 261.5 articles ± 51.1 per week) as compared with original articles (mean of 69.3 ± 22.3 articles per week). Original articles including patient data accounted for 713 (9.5%) of peer-reviewed studies. A total of 576 original articles (80.8%) showed intermediate to high risk of bias. Last, except for simulation studies that mainly used large-scale open data, the median number of patients enrolled was of 102 (IQR = 37–337). Conclusions Since the beginning of the COVID-19 pandemic, the majority of research is composed by publications without original data. Peer-reviewed original articles with data showed a high risk of bias and included a limited number of patients. Together, these findings underscore the urgent need to strike a balance between the velocity and quality of research, and to cautiously consider medical information and clinical applicability in a pressing, pandemic context. Systematic review registration https://osf.io/5zjyx/

are current employees of Geneplus-Beijing. C. Wang and Z. Yu are current employees of Geneplus-Shenzhen. L. Yang holds leadership positions of Geneplus-Beijing. The other authors declare no conflicts of interest. Translational Relevance Neoadjuvant chemoradiotherapy (nCRT) was the standard of care for patients with locally advanced rectal cancer (LARC), however the uniform regimen may not be applicable for all patients with different tumor loads and heterogeneous biological behaviors. In this study, the preoperative ctDNA status was significantly consistent with the postoperative pathological results, showing that ctDNA can accurately reflect the real-time tumor burden. Additionally, ctDNA showed a predictive ability for distant metastasis as early as prior to treatment. Besides, tumors with POLD1 mutation had significantly better response to nCRT than those without POLD1 mutation. These findings imply that ctDNA and tumor mutational information may potentially be powerful tools to guide the individualized multidisciplinary therapy for patients with LARC by assisting the selection of initial treatment strategies and regimens, or guiding the adjustment of treatment methods.

Rationale: Ischemia/reperfusion injury (IRI) is a major cause of acute kidney injury (AKI) that is associated with high morbidity and mortality, and for which specific treatments are lacking. In this study, we investigated the protective effect of human urine-derived stem cells (USCs) and their exosomes against IRI-induced AKI to explore the potential of these cells as a new therapeutic strategy. Methods: USCs were derived from fresh human urine. Cell surface marker expression was analyzed by flow cytometry to determine the characteristics of the stem cells. Adult male Sprague-Dawley rats were used to generate a lethal renal IRI model. One dose of USCs (2×10 6 cells/ml) or exosomes (20 µg/1 ml) in the experimental groups or saline (1 ml) in the control group was administered intravenously immediately after blood reperfusion. Blood was drawn every other day for measurement of serum creatinine (sCr) and blood urea nitrogen (BUN) levels. The kidneys were harvested for RNA and protein extraction to examine the levels of apoptosis and tubule injury. In vitro , the hypoxia-reoxygenation (H/R) model in human kidney cortex/proximal tubule cells (HK2) was used to analyze the protective ability of USC-derived exosomes (USC-Exo). Quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR), western blotting, superoxide dismutase activity, and malonaldehyde content analyses were used to evaluate oxidative stress in HK2 cells treated with USC-Exo after H/R. Exosomal microRNA sequencing techniques and bioinformatics analysis were used to search for enriched miRNAs in the exosomes and interacting genes. The interaction between miRNAs and the 3' untranslated region of the target gene was detected using a dual luciferase reporting system. The miRNA mimic and inhibitor were used to regulate the miRNA level in HK2 cells. Results: Treatment with USCs led to reductions in the levels of sCr, BUN, and renal tubular cell apoptosis; inhibited the infiltration of inflammatory cells; and protected renal function in the rat IRI model. Additionally, USC-derived exosomes protected against IRI-induced renal damage. miR-146a-5p was the most abundant miRNA in exosomes obtained from the conditioned medium (CM) of USCs. miR-146a-5p targeted and degraded the 3'UTR of interleukin-1 receptor-associated kinase 1 (IRAK1) mRNA, subsequently inhibited the activation of nuclear factor (NF)-κB signaling, and protected HK2 cells from H/R injury. USC transplantation also upregulated miR-146a-5p expression, downregulated IRAK1 expression and inhibited nuclear translocation of NF-κB p65 in the kidney of the rat IRI model. Conclusions: According to our experimental results, USCs could protect against renal IRI via exosomal miR-146a-5p , which could target the 3'UTR of I...

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Bladder cancer is one of the most common and highly vascularized cancers. To better understand its genomic structure and underlying etiology, we conduct whole-genome and targeted sequencing in urothelial bladder carcinomas (UBCs, the most common type of bladder cancer). Recurrent mutations in noncoding regions affecting gene regulatory elements and structural variations (SVs) leading to gene disruptions are prevalent. Notably, we find recurrent ADGRG6 enhancer mutations and FRS2 duplications which are associated with higher protein expression in the tumor and poor prognosis. Functional assays demonstrate that depletion of ADGRG6 or FRS2 expression in UBC cells compromise their abilities to recruit endothelial cells and induce tube formation. Moreover, pathway assessment reveals recurrent alterations in multiple angiogenesis-related genes. These results illustrate a multidimensional genomic landscape that highlights noncoding mutations and SVs in UBC tumorigenesis, and suggest ADGRG6 and FRS2 as novel pathological angiogenesis regulators that would facilitate vascular-targeted therapies for UBC.

BackgroundThe dramatic growth of mobile phone (MP) use among young people has increased interest in its possible health hazards in this age group. The aim of this cross-sectional study was to investigate the association between MP use and inattention in adolescents.MethodsA total of 7720 middle school students were involved in this cross-sectional study. Inattention was assessed as defined for the Attention Deficit component of Attention deficit/Hyperactivity disorder (ADHD) by the Diagnostic and Statistical Manual of Mental Disorders (4th ed., text rev. [DSM-IV-TR]). The demographic characteristics and information on MP use were included in the questionnaire. Chi-square tests and logistic regression models were used to analyze the data.ResultsIn total, 7102 (91.99%) valid questionnaires were obtained. After adjusted for confounders, inattention in adolescents was significantly associated with MP ownership, the time spent on entertainment on MP per day, the position of the MP during the day and the mode of the MP at night. The strongest association between inattention and the time spent on the MP was among students who spent more than 60 minutes per day playing on their MP.ConclusionsOur study shows some associations between MP use and inattention in Chinese adolescents. Decreasing MP usage to less than 60 minutes per day may help adolescents to stay focused and centered.

T lymphocytes infiltrate the microenvironment of breast cancer tumors and play a pivotal role in tumor immune surveillance. Relationships between the T-cell receptors (TCR) borne by T cells within tumors, in the surrounding tissues, and in draining lymph nodes are largely unexplored in human breast cancer. Consequently, information about the relative extent of possible T-cell exchange between these tissues is also lacking. Here, we have analyzed the TCR repertoire of T cells using multiplex PCR and high-throughput sequencing of the TCRβ chain in the tissues of tumor, adjacent nontumor, and axillary lymph nodes of breast cancer patients. T-cell repertoire diversity in tumors was lower than in lymph nodes, but higher than in nontumor tissue, with a preferential use of variable and joining genes. These data are consistent with the hypothesis that most of the T cells in tumors derive from the lymph node, followed by their expansion in tumor tissue. Positive nodes appeared to enhance T-cell infiltration into tumors and T-cell clonal expansion in lymph nodes. Additionally, the similarity in TCR repertoire between tumor and nontumor tissue was significantly higher in luminal-like, rather than basal-like, breast cancer. Our study elucidated the high heterogeneity of the TCR repertoire and provides potential for future improvements in immune-related diagnosis, therapy, and prognosis for breast cancer patients. Cancer Immunol Res; 5(2); 148-56. ©2016 AACR.