Biology subjects

Yan, M.

Publications and source records attributed to Yan, M..

62 records · Page 4Linked to original sources

bioRxiv2019

Cloning human dental pulp cells and studying inter-clone diversity

Heterogeneity within a putative stem cell population presents a challenge for studies and applications of such cells. Cloning may provide a strategy for reducing heterogeneity. However, previous studies have the weakness in reliability of single-cell-origin of the colonies. The present study aims to apply an alternative method to obtain clonal dental pulp cells with increased reliability of single cell origin. Dental pulp cells were cultured from 13 human wisdom teeth. Primary cultures of 3 human benign tumors were included as comparison. Cells were seeded into wells of a 96-plate at a mean density of 1 cell/well. On the next day, wells were inspected one by one to identify wells with single cells which were followed for 3 weeks. Survived clones were expanded and further characterized. Single cells were observed in all cases, the number of single-cell-wells varied from 16 to 33. Three weeks later, survived and grown clonal cells were observed in 10 to 29 wells, giving surviving rates of 33-91%. By contrast, though single tumor cells were also observed, none of them survived. Expanded clones exhibited diversity in viability and osteogenic differentiation which also differed from their parental cells. Seeding cells at clonal density into physically separated compartments like wells in a 96 plate and comprehensive observation provides a practical strategy for increasing reliability of single-cell origin of clonal cells. Cellular heterogeneity seems to be an intrinsic feature of dental pulp cells.

cell biology↗

bioRxiv2019

High-throughput multiplexed tandem repeat genotyping using targeted long-read sequencing

Tandem repeats (TRs) are highly prone to variation in copy numbers due to their repetitive and unstable nature, which makes them a major source of genomic variation between individuals. However, population variation of TRs have not been widely explored due to the limitations of existing tools, which are either low-throughput or restricted to a small subset of TRs. Here, we used SureSelect targeted sequencing approach combined with Nanopore sequencing to overcome these limitations. We achieved an average of 3062-fold target enrichment on a panel of 142 TR loci, generating an average of 97X sequence coverage on 7 samples utilizing 2 MinION flow-cells with 200ng of input DNA per sample. We identified a subset of 110 TR loci with length less than 2kb, and GC content greater than 25% for which we achieved an average genotyping rate of 75% and increasing to 91% for the highest-coverage sample. Alleles estimated from targeted long-read sequencing were concordant with gold standard PCR sizing analysis and moreover highly correlated with alleles estimated from whole genome long-read sequencing. We demonstrate a targeted long-read sequencing approach that enables simultaneous analysis of hundreds of TRs and accuracy is comparable to PCR sizing analysis. Our approach is feasible to scale for more targets and more samples facilitating large-scale analysis of TRs.

genomics↗

bioRxiv2019

Global analysis of epigenetic heterogeneity identifies divergent drivers of esophageal squamous cell carcinoma

Epigenetic landscapes can shape physiologic and disease phenotypes. We used integrative, high resolution multi-omics methods to characterize the oncogenic drivers of esophageal squamous cell carcinoma (ESCC). We found 98% of CpGs are hypomethylated across the ESCC genome and two-thirds occur in long non-coding (lnc)RNA regions. DNA methylation and epigenetic heterogeneity both coincide with chromosomal topological alterations. Gene body methylation, polycomb repressive complex occupancy, and CTCF binding sites associate with cancer-specific gene regulation. Epigenetically-mediated activation of non-canonical WNT signaling and the lncRNA ESCCAL-1 were validated as potential ESCC driver alterations. Gene-specific cancer driver roles of epigenetic alterations and heterogeneity are identified.

cell biology↗

bioRxiv2019

Pharmacokinetics Modeling and Simulation of Voriconazole Dosing and Safety in Patients with Liver Cirrhosis

Voriconazole is used to treat invasive fungal disease and the optimal dose regimens are still unknown in cirrhotic Patients. The aim of this study was to determine the safety, to describe pharmacokinetics characteristics, and to optimize dosage regimens of voriconazole in cirrhotic patients. Data pertaining to voriconazole were collected retrospectively and analyzed using a population pharmacokinetics model. A total of 219 trough concentrations (Cmin) from 120 patients were analyzed. Voriconazole-related adverse events developed in 29 patients, with 69.0% of AEs developing within the first week after voriconazole treatment. The threshold Cmin for AEs was 5.12 mg/L. A one-compartment model with first-order absorption and elimination adequately described the data. The Child-Pugh class was the only covariate in final model. Voriconazole clearance in patients with Child-pugh A and B cirrhosis (CP-A/CP-B) and Child-pugh C cirrhosis (CP-C) were 1.79 L/h and 0.99 L/h, respectively, the volume of distribution was 159.6 L, and the oral bioavailability was 91.8%. The elimination half-life was significantly extended for up to 61.8 - 111.7 h in cirrhotic patients. Model-based simulations showed that the appropriate maintenance doses are 75 mg/12 h and 150 mg/24 h intravenously or orally for CP-A/CP-B patients, and 50 mg/12 h and 100 mg/24 h intravenously or orally for CP-C patients. The results support voriconazole maintenance doses in LC patients should be reduced to one-fourth for CP-C patients and to one-third for CP-A/CP-B patients compared to that for patients with normal liver function. Monitoring Cmin early could be a useful strategy to ensure the safety.

pharmacology and toxicology↗

bioRxiv2018

Fast and Robust Deconvolution of Tumor Infiltrating Lymphocyte from Expression Profiles using Least Trimmed Squares

Gene-expression deconvolution is used to quantify different types of cells in a mixed population. It provides a highly promising solution to rapidly characterize the tumor-infiltrating immune landscape and identify cold cancers. However, a major challenge is that gene-expression data are frequently contaminated by many outliers that decrease the estimation accuracy. Thus, it is imperative to develop a robust deconvolution method that automatically decontaminates data by reliably detecting and removing outliers. We developed a new machine learning tool, Fast And Robust DEconvolution of Expression Profiles (FARDEEP), to enumerate immune cell subsets from whole tumor tissue samples. To reduce noise in the tumor gene expression datasets, FARDEEP utilizes an adaptive least trimmed square to automatically detect and remove outliers before estimating the cell compositions. We show that FARDEEP is less susceptible to outliers and returns a better estimation of coefficients than the existing methods with both numerical simulations and real datasets. FARDEEP provides the absolute quantitation of each immune cell subset in addition to relative percentages. Hence, FARDEEP represents a novel robust algorithm to complement the existing toolkit for the characterization of tissue-infiltrating immune cell landscape. The source code for FARDEEP as implemented in R is available for download at https://goo.gl/SqGKuo.

bioinformatics↗

bioRxiv2017

Active site alanine substitutions can convert deubiquitinating enzymes into avid ubiquitin-binding domains

A common strategy for studying the biological role of deubiquitinating enzymes (DUBs) in different pathways is to study the effects of replacing the wild type DUB with a catalytically inactive mutant in cells. We report here that a commonly studied DUB mutation, in which the catalytic cysteine is replaced with alanine, can dramatically increase the affinity of some DUBs for ubiquitin. Overexpression of these tight-binding mutants thus has the potential to sequester cellular pools of monoubiquitin and ubiquitin chains. As a result, cells expressing these mutants may display unpredictable dominant negative physiological effects that are not related to loss of DUB activity. The structure of the SAGA DUB module bound to free ubiquitin reveals the structural basis for the 30-fold higher affinity of Ubp8C146A for ubiquitin. We show that an alternative option, substituting the active site cysteine with arginine, can inactivate DUBs while also decreasing the affinity for ubiquitin.

biochemistry↗

bioRxiv2017

The pomegranate (Punica granatum L.) genome provides insights into fruit quality and ovule developmental biology

Pomegranate (Punica granatum L.) with an uncertain taxonomic status has an ancient cultivation history, and has become an emerging fruit due to its attractive features such as the bright red appearance and the high abundance of medicinally valuable ellagitannin-based compounds in its peel and aril. However, the absence of genomic resources has restricted further elucidating genetics and evolution of these interesting traits. Here we report a 274-Mb high-quality draft pomegranate genome sequence, which covers approximately 81.5% of the estimated 336 Mb genome, consists of 2,177 scaffolds with an N50 size of 1.7 Mb, and contains 30,903 genes. Phylogenomic analysis supported that pomegranate belongs to the Lythraceae family rather than the monogeneric Punicaceae family, and comparative analyses showed that pomegranate and Eucalyptus grandis shares the paleotetraploidy event. Integrated genomic and transcriptomic analyses provided insights into the molecular mechanisms underlying the biosynthesis of ellagitannin-based compounds, the color formation in both peels and arils during pomegranate fruit development, and the unique ovule development processes that are characteristic of pomegranate. This genome sequence represents the first reference in Lythraceae, providing an important resource to expand our understanding of some unique biological processes and to facilitate both comparative biology studies and crop breeding.

genomics↗

bioRxiv2017

Macrophages are required to coordinate mouse digit tip regeneration

In mammals, macrophages are known to play a major role in tissue regeneration. These cells contribute to inflammation, histolysis, re-epithelialization, re-vascularization and cell proliferation. While macrophages have been shown to be essential for epimorphic regeneration in salamanders and fish, their role has not been elucidated in mammalian epimorphic regeneration. Here, using the mouse digit tip model as a mammalian model of epimorphic regeneration, we demonstrate that macrophages are essential for the regeneration process. Using cell depletion strategies, we show that regeneration is completely inhibited; bone histolysis does not occur, wound re-epithelization is inhibited and the blastema does not form. Rescue of epidermal wound closure, in the absence of macrophages, promotes blastema accumulation but not differentiation indicating that macrophage play a role in re-differentiation. Additionally, inhibition of osteoclasts and the degradation process is not sufficient to inhibit regeneration. These findings show that macrophages play an essential role in coordinating the epimorphic regenerative response in mammals.

developmental biology↗