scholarly journals Identification of Specific Cell Subpopulations and Marker Genes in Ovarian Cancer Using Single-Cell RNA Sequencing

2021 ◽  
Vol 2021 ◽  
pp. 1-27
Author(s):  
Yan Li ◽  
Juan Wang ◽  
Fang Wang ◽  
Chengzhen Gao ◽  
Yuanyuan Cao ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Single Cell ◽  
Differentially Expressed Genes ◽  
Single Cells ◽  
Differentially Expressed ◽  
Ovarian Cancer Cells ◽  
Marker Genes ◽  
Specific Cell ◽  
Rna Seq ◽  
Cell Subpopulations

Objective. Ovarian cancer is the deadliest gynaecological cancer globally. In our study, we aimed to analyze specific cell subpopulations and marker genes among ovarian cancer cells by single-cell RNA sequencing (RNA-seq). Methods. Single-cell RNA-seq data of 66 high-grade serous ovarian cancer cells were employed from the Gene Expression Omnibus (GEO). Using the Seurat package, we performed quality control to remove cells with low quality. After normalization, we detected highly variable genes across the single cells. Then, principal component analysis (PCA) and cell clustering were performed. The marker genes in different cell clusters were detected. A total of 568 ovarian cancer samples and 8 normal ovarian samples were obtained from The Cancer Genome Atlas (TCGA) database. Differentially expressed genes were identified according to ∣ log 2 fold   change   FC ∣ > 1 and adjusted p value <0.05. To explore potential biological processes and pathways, functional enrichment analyses were performed. Furthermore, survival analyses of differentially expressed marker genes were performed. Results. After normalization, 6000 highly variable genes were identified across the single cells. The cells were divided into 3 cell populations, including G1, G2M, and S cell cycles. A total of 1,124 differentially expressed genes were identified in ovarian cancer samples. These differentially expressed genes were enriched in several pathways associated with cancer, such as metabolic pathways, pathways in cancer, and PI3K-Akt signaling pathway. Furthermore, marker genes, STAT1, ANP32E, GPRC5A, and EGFL6, were highly expressed in ovarian cancer, while PMP22, FBXO21, and CYB5R3 were lowly expressed in ovarian cancer. These marker genes were positively associated with prognosis of ovarian cancer. Conclusion. Our findings revealed specific cell subpopulations and marker genes in ovarian cancer using single-cell RNA-seq, which provided a novel insight into the heterogeneity of ovarian cancer.

scholarly journals Comprehensive benchmarking of single cell RNA sequencing technologies for characterizing cellular perturbation

2020 ◽  
Author(s):  
Verboom Karen ◽  
Alemu T Assefa ◽  
Nurten Yigit ◽  
Jasper Anckaert ◽  
Niels Vandamme ◽  
...  
Keyword(s):  
Single Cell ◽  
Differentially Expressed Genes ◽  
Single Cells ◽  
Neuroblastoma Cells ◽  
Transcriptional Response ◽  
Differentially Expressed ◽  
Cellular Heterogeneity ◽  
Rna Seq ◽  
Library Size ◽  
Cell Subpopulations

ABSTRACTTechnological advances in transcriptome sequencing of single cells continues to provide an unprecedented view on tissue composition and cellular heterogeneity. While several studies have compared different single cell RNA-seq methods with respect to data quality and their ability to distinguish cell subpopulations, none of these studies investigated the heterogeneity of the cellular transcriptional response upon a chemical perturbation. In this study, we evaluated the transcriptional response of NGP neuroblastoma cells upon nutlin-3 treatment using the C1, ddSeq and Chromium single cell systems. These devices and library preparation methods are representative for the wide variety of platforms, ranging from microfluid chips to droplet-based systems and from full transcript sequencing to 3-prime end sequencing. In parallel, we used bulk RNA-seq for molecular characterization of the transcriptional response. Two complementary metrics to evaluate performance were applied: the first is the number and identity of differentially expressed genes as defined in consensus by two statistical models, and the second is the enrichment analysis of biological signals. Where relevant, to make the data more comparable, we downsampled sequencing library size, selected cell subpopulations based on specific RNA abundance features, or created pseudobulk samples. While the C1 detects the highest number of genes per cell and better resembles bulk RNA-seq, the Chromium identifies most differentially expressed genes, albeit still substantially fewer than bulk RNA-seq. Gene set enrichment analyses reveals that detection of a limited set of the most abundant genes in single cell RNA-seq experiments is sufficient for molecular phenotyping. Finally, single cell RNA-seq reveals a heterogeneous response of NGP neuroblastoma cells upon nutlin-3 treatment, revealing putative late-responder or resistant cells, both undetected in bulk RNA-seq experiments.

scholarly journals Single cell-derived spheroids capture the self-renewing subpopulations of metastatic ovarian cancer

2021 ◽  
Author(s):  
Tania Velletri ◽  
Carlo Emanuele Villa ◽  
Domenica Cilli ◽  
Bianca Barzaghi ◽  
Pietro Lo Riso ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Single Cell ◽  
Single Cells ◽  
Unmet Need ◽  
3D Culture ◽  
Patient Specific ◽  
Precision Oncology ◽  
Cell Subpopulations ◽  
Cancer Spheroids ◽  
2D And 3D

AbstractHigh Grade Serous Ovarian cancer (HGSOC) is a major unmet need in oncology, due to its precocious dissemination and the lack of meaningful human models for the investigation of disease pathogenesis in a patient-specific manner. To overcome this roadblock, we present a new method to isolate and grow single cells directly from patients’ metastatic ascites, establishing the conditions for propagating them as 3D cultures that we refer to as single cell-derived metastatic ovarian cancer spheroids (sMOCS). By single cell RNA sequencing (scRNAseq) we define the cellular composition of metastatic ascites and trace its propagation in 2D and 3D culture paradigms, finding that sMOCS retain and amplify key subpopulations from the original patients’ samples and recapitulate features of the original metastasis that do not emerge from classical 2D culture, including retention of individual patients’ specificities. By enabling the enrichment of uniquely informative cell subpopulations from HGSOC metastasis and the clonal interrogation of their diversity at the functional and molecular level, this method provides a powerful instrument for precision oncology in ovarian cancer.

ColorCells: a database of expression, classification and functions of lncRNAs in single cells

2020 ◽  
Author(s):  
Ling-Ling Zheng ◽  
Jing-Hua Xiong ◽  
Wu-Jian Zheng ◽  
Jun-Hao Wang ◽  
Zi-Liang Huang ◽  
...  
Keyword(s):  
Single Cell ◽  
Tissue Specificity ◽  
Single Cells ◽  
Noncoding Rnas ◽  
Cell Types ◽  
Marker Genes ◽  
Rna Seq ◽  
Expression Variability ◽  
Expression Classification

Abstract Although long noncoding RNAs (lncRNAs) have significant tissue specificity, their expression and variability in single cells remain unclear. Here, we developed ColorCells (http://rna.sysu.edu.cn/colorcells/), a resource for comparative analysis of lncRNAs expression, classification and functions in single-cell RNA-Seq data. ColorCells was applied to 167 913 publicly available scRNA-Seq datasets from six species, and identified a batch of cell-specific lncRNAs. These lncRNAs show surprising levels of expression variability between different cell clusters, and has the comparable cell classification ability as known marker genes. Cell-specific lncRNAs have been identified and further validated by in vitro experiments. We found that lncRNAs are typically co-expressed with the mRNAs in the same cell cluster, which can be used to uncover lncRNAs’ functions. Our study emphasizes the need to uncover lncRNAs in all cell types and shows the power of lncRNAs as novel marker genes at single cell resolution.

scholarly journals Single-cell mapper (scMappR): using scRNA-seq to infer the cell-type specificities of differentially expressed genes

2021 ◽  
Vol 3 (1) ◽  
Author(s):  
Dustin J Sokolowski ◽  
Mariela Faykoo-Martinez ◽  
Lauren Erdman ◽  
Huayun Hou ◽  
Cadia Chan ◽  
...  
Keyword(s):  
Single Cell ◽  
Differentially Expressed Genes ◽  
Cell Types ◽  
Differentially Expressed ◽  
Rna Seq ◽  
Kidney Regeneration ◽  
Cell Type ◽  
Cell Type Specificity ◽  
Cost Constraints ◽  
Mouse Tissues

Abstract RNA sequencing (RNA-seq) is widely used to identify differentially expressed genes (DEGs) and reveal biological mechanisms underlying complex biological processes. RNA-seq is often performed on heterogeneous samples and the resulting DEGs do not necessarily indicate the cell-types where the differential expression occurred. While single-cell RNA-seq (scRNA-seq) methods solve this problem, technical and cost constraints currently limit its widespread use. Here we present single cell Mapper (scMappR), a method that assigns cell-type specificity scores to DEGs obtained from bulk RNA-seq by leveraging cell-type expression data generated by scRNA-seq and existing deconvolution methods. After evaluating scMappR with simulated RNA-seq data and benchmarking scMappR using RNA-seq data obtained from sorted blood cells, we asked if scMappR could reveal known cell-type specific changes that occur during kidney regeneration. scMappR appropriately assigned DEGs to cell-types involved in kidney regeneration, including a relatively small population of immune cells. While scMappR can work with user-supplied scRNA-seq data, we curated scRNA-seq expression matrices for ∼100 human and mouse tissues to facilitate its stand-alone use with bulk RNA-seq data from these species. Overall, scMappR is a user-friendly R package that complements traditional differential gene expression analysis of bulk RNA-seq data.

scholarly journals Single cell derived organoids capture the self-renewing subpopulations of metastatic ovarian cancer

2018 ◽  
Author(s):  
Tania Velletri ◽  
Emanuele Carlo Villa ◽  
Michela Lupia ◽  
Pietro Lo Riso ◽  
Raffaele Luongo ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Single Cell ◽  
Molecular Level ◽  
Single Cells ◽  
Unmet Need ◽  
3D Culture ◽  
Patient Specific ◽  
Precision Oncology ◽  
Cell Subpopulations ◽  
2D And 3D

AbstractHigh Grade Serous Ovarian cancer (HGSOC) is a major unmet need in oncology, due to its precocious dissemination and the lack of meaningful human models for the investigation of disease pathogenesis in a patient-specific manner. To overcome this roadblock, we present a new method to isolate and grow single cells directly from patients’ ascites, establishing the conditions for propagating them as single-cell derived ovarian cancer organoids (scOCOs). By single cell RNA sequencing (scRNAseq) we define the cellular composition of metastatic ascites and trace its propagation in 2D and 3D culture paradigms, finding that scOCOs retain and amplify key subpopulations from the original patients’ samples and recapitulate features of the original metastasis that do not emerge from classical 2D culture, including retention of individual patients’ specificities. By enabling the enrichment of uniquely informative cell subpopulations from HGSOC metastasis and the clonal interrogation of their diversity at the functional and molecular level, this method transforms the prospects of precision oncology for ovarian cancer.

scholarly journals powsimR: Power analysis for bulk and single cell RNA-seq experiments

2017 ◽  
Author(s):  
Beate Vieth ◽  
Christoph Ziegenhain ◽  
Swati Parekh ◽  
Wolfgang Enard ◽  
Ines Hellmann
Keyword(s):  
Single Cell ◽  
Differential Expression ◽  
Differentially Expressed Genes ◽  
Power Analysis ◽  
A Priori ◽  
Differentially Expressed ◽  
Rna Seq ◽  
Flexible Tool ◽  
Power Analyses

AbstractPower analysis is essential to optimize the design of RNA-seq experiments and to assess and compare the power to detect differentially expressed genes in RNA-seq data. PowsimR is a flexible tool to simulate and evaluate differential expression from bulk and especially single-cell RNA-seq data making it suitable for a priori and posterior power analyses.

scholarly journals Single-cell mapper (scMappR): using scRNA-seq to infer cell-type specificities of differentially expressed genes

2020 ◽  
Author(s):  
Dustin J. Sokolowski ◽  
Mariela Faykoo-Martinez ◽  
Lauren Erdman ◽  
Huayun Hou ◽  
Cadia Chan ◽  
...  
Keyword(s):  
Single Cell ◽  
Differential Expression ◽  
Differentially Expressed Genes ◽  
Cell Types ◽  
R Package ◽  
Differentially Expressed ◽  
Rna Seq ◽  
Kidney Regeneration ◽  
Cell Type ◽  
User Friendly

AbstractRNA sequencing (RNA-seq) is widely used to identify differentially expressed genes (DEGs) and reveal biological mechanisms underlying complex biological processes. RNA-seq is often performed on heterogeneous samples and the resulting DEGs do not necessarily indicate the cell types where the differential expression occurred. While single-cell RNA-seq (scRNA-seq) methods solve this problem, technical and cost constraints currently limit its widespread use. Here we present single cell Mapper (scMappR), a method that assigns cell-type specificity scores to DEGs obtained from bulk RNA-seq by integrating cell-type expression data generated by scRNA-seq and existing deconvolution methods. After benchmarking scMappR using RNA-seq data obtained from sorted blood cells, we asked if scMappR could reveal known cell-type specific changes that occur during kidney regeneration. We found that scMappR appropriately assigned DEGs to cell-types involved in kidney regeneration, including a relatively small proportion of immune cells. While scMappR can work with any user supplied scRNA-seq data, we curated scRNA-seq expression matrices for ∼100 human and mouse tissues to facilitate its use with bulk RNA-seq data alone. Overall, scMappR is a user-friendly R package that complements traditional differential expression analysis available at CRAN.HighlightsscMappR integrates scRNA-seq and bulk RNA-seq to re-calibrate bulk differentially expressed genes (DEGs).scMappR correctly identified immune-cell expressed DEGs from a bulk RNA-seq analysis of mouse kidney regeneration.scMappR is deployed as a user-friendly R package available at CRAN.

scholarly journals MarcoPolo: a clustering-free approach to the exploration of differentially expressed genes along with group information in single-cell RNA-seq data

2020 ◽  
Author(s):  
Chanwoo Kim ◽  
Hanbin Lee ◽  
Juhee Jeong ◽  
Keehoon Jung ◽  
Buhm Han
Keyword(s):  
Single Cell ◽  
Differentially Expressed Genes ◽  
Differential Expression Analysis ◽  
Feature Selection Method ◽  
Real Data ◽  
Cell Types ◽  
Differentially Expressed ◽  
Rna Seq ◽  
Sequencing Data ◽  
Group Information

ABSTRACTA common approach to analyzing single-cell RNA-sequencing data is to cluster cells first and then identify differentially expressed genes based on the clustering result. However, clustering has an innate uncertainty and can be imperfect, undermining the reliability of differential expression analysis results. To overcome this challenge, we present MarcoPolo, a clustering-free approach to exploring differentially expressed genes. To find informative genes without clustering, MarcoPolo exploits the bimodality of gene expression to learn the group information of the cells with respect to the expression level directly from given data. Using simulations and real data analyses, we showed that our method puts biologically informative genes at higher ranks more accurately and robustly than other existing methods. As our method provides information on how cells can be grouped for each gene, it can help identify cell types that are not separated well in the standard clustering process. Our method can also be used as a feature selection method to improve the robustness against changes in the number of genes used in clustering.

scholarly journals Autophagy Regulatory Genes MET and RIPK2 Play a Prognostic Role in Pancreatic Ductal Adenocarcinoma: A Bioinformatic Analysis Based on GEO and TCGA

2020 ◽  
Vol 2020 ◽  
pp. 1-15
Author(s):  
Xingyu Li ◽  
Zhiqiang Li ◽  
Hongwei Zhu ◽  
Xiao Yu
Keyword(s):  
Single Cell ◽  
Pancreatic Ductal Adenocarcinoma ◽  
Cancer Cells ◽  
Differentially Expressed Genes ◽  
Clinical Application ◽  
Regulatory Genes ◽  
Gene Set Enrichment Analysis ◽  
Differentially Expressed ◽  
Ductal Adenocarcinoma ◽  
Rna Seq

Pancreatic ductal adenocarcinoma is a common malignant tumor with a poor prognosis. Autophagy activity changes in both cancer cells and microenvironment and affects the progression of pancreatic ductal adenocarcinoma. The purpose of this study was to predict the prognostic autophagy regulatory genes and their role in the regulation of autophagy in pancreatic ductal adenocarcinoma. We draw conclusions based on gene expression data from different platforms: GSE62165 and GSE85916 from the array platform, TCGA from the bulk RNA-seq platform, and GSE111672 from the single-cell RNA-seq platform. At first, we detected differentially expressed genes in pancreatic ductal adenocarcinoma compared with normal pancreatic tissue based on GSE62165. Then, we screened prognostic genes based on GSE85916 and TCGA. Furthermore, we constructed a risk signature composed of the prognostic differentially expressed genes. Finally, we predicted the probable role of these genes in regulating autophagy and the types of cell expressing these genes. According to our screening criteria, there were only two genes: MET and RIPK2, selected into the development of the risk signature. However, evaluated by log-rank tests, receiver operating characteristic curves, and calibration curves, the risk signature was worth considering its clinical application because of good sensitivity, specificity, and stability. Besides, we predicted that both MET and RIPK2 promote autophagy in pancreatic ductal adenocarcinoma by gene set enrichment analysis. Analysis of single-cell RNA-seq data from GSE111672 revealed that both MET and RIPK2 were expressed in cancer cells while RIPK2 was also expressed in monocytes and neutrophils. After comprehensive analysis, we found that both MET and RIPK2 are related to the prognosis of pancreatic ductal adenocarcinoma and provided some associated clues for clinical application and basic experiment research.

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