Dysfunctional tumor-infiltrating Vδ1 + T lymphocytes in microsatellite-stable colorectal cancer.

Although γδ T cells are known to participate in immune dysregulation in solid tumors, their relevance to human microsatellite-stable (MSS) colorectal cancer (CRC) is still undefined. Here, using integrated gene expression analysis and T cell receptor sequencing, we characterized γδ T cells in MSS CRC, with a focus on Vδ1 + T cells. We identified Vδ1 T cells with shared motifs in the third complementarity-determining region of the δ-chain, reflective of antigen recognition.

Secondary structure of the human mitochondrial genome affects formation of deletions.

Background: Aging in postmitotic tissues is associated with clonal expansion of somatic mitochondrial deletions, the origin of which is not well understood. Such deletions are often flanked by direct nucleotide repeats, but this alone does not fully explain their distribution. Here, we hypothesized that the close proximity of direct repeats on single-stranded mitochondrial DNA (mtDNA) might play a role in the formation of deletions.

Altered differentiation of endometrial mesenchymal stromal fibroblasts is associated with endometriosis susceptibility.

Cellular development is tightly regulated as mature cells with aberrant functions may initiate pathogenic processes. The endometrium is a highly regenerative tissue, shedding and regenerating each month. Endometrial stromal fibroblasts are regenerated each cycle from mesenchymal stem cells and play a pivotal role in endometriosis, a disease characterised by endometrial cells that grow outside the uterus.

Secreted Toxins From Strains Isolated From Keratinocyte Skin Cancers Mediate Pro-tumorigenic Inflammatory Responses in the Skin.

Squamous cell carcinoma (SCC) is a common type of skin cancer that typically arises from premalignant precursor lesions named actinic keratoses (AK). Chronic inflammation is a well-known promoter of skin cancer progression. AK and SCC have been associated with an overabundance of the bacterium ().

A model of impaired Langerhans cell maturation associated with HPV induced epithelial hyperplasia.

Langerhans cells (LC) are skin-resident antigen-presenting cells that regulate immune responses to epithelial microorganisms. Human papillomavirus (HPV) infection can promote malignant epithelial transformation. As LCs are considered important for controlling HPV infection, we compared the transcriptome of murine LCs from skin transformed by K14E7 oncoprotein and from healthy skin.

Acquisition of murine splenic myeloid cells for protein and gene expression profiling by advanced flow cytometry and CITE-seq.

Here, we outline detailed protocols to isolate and profile murine splenic dendritic cells (DCs) through advanced flow cytometry of the myeloid compartment and single-cell transcriptomic profiling with integrated cell surface protein expression through CITE-seq. This protocol provides a general transferrable road map for different tissues and species. For complete details on the use and execution of this protocol, please refer to Lukowski et al.

Absence of Batf3 reveals a new dimension of cell state heterogeneity within conventional dendritic cells.

Conventional dendritic cells (cDCs) are traditionally subdivided into cDC1 and cDC2 lineages. Batf3 is a cDC1-required transcription factor, and we observed that -/- mice harbor a population of cDC1-like cells co-expressing cDC2-associated surface molecules. Using single-cell RNA sequencing with integrated cell surface protein expression (CITE-seq), we found that -/- mitotic immature cDC1-like cells showed reduced expression of cDC1 features and increased levels of cDC2 features.

Single cell eQTL analysis identifies cell type-specific genetic control of gene expression in fibroblasts and reprogrammed induced pluripotent stem cells.

Background: The discovery that somatic cells can be reprogrammed to induced pluripotent stem cells (iPSCs) has provided a foundation for in vitro human disease modelling, drug development and population genetics studies. Gene expression plays a critical role in complex disease risk and therapeutic response. However, while the genetic background of reprogrammed cell lines has been shown to strongly influence gene expression, the effect has not been evaluated at the level of individual cells which would provide significant resolution.

Antigen Nonspecific Induction of Distinct Regulatory T Cell States in Oncogene-Driven Hyperproliferative Skin.

Regulatory T cells (Tregs) are recruited to nonlymphoid tissues in chronic disease, including cancer, and the tissue environment is held to shape the Treg phenotype diversity. Using single-cell RNA sequencing, we examined the transcriptomic and TCR profile of Tregs recruited to hyperproliferative HPV16 E7-expressing transgenic and control nontransgenic murine skin grafts. Tregs were more abundant in E7 transgenic skin grafts than control grafts, without evidence of E7 specificity.

Comparative performance of the BGI and Illumina sequencing technology for single-cell RNA-sequencing.

The libraries generated by high-throughput single cell RNA-sequencing (scRNA-seq) platforms such as the Chromium from 10× Genomics require considerable amounts of sequencing, typically due to the large number of cells. The ability to use these data to address biological questions is directly impacted by the quality of the sequence data. Here we have compared the performance of the Illumina NextSeq 500 and NovaSeq 6000 against the BGI MGISEQ-2000 platform using identical Single Cell 3' libraries consisting of over 70 000 cells generated on the 10× Genomics Chromium platform.

Chemical Modulators of Fibrinogen Production and Their Impact on Venous Thrombosis.

Thrombosis is a leading cause of morbidity and mortality. Fibrinogen, the soluble substrate for fibrin-based clotting, has a central role in haemostasis and thrombosis and its plasma concentration correlates with cardiovascular disease event risk and a prothrombotic state in experimental models. We aimed to identify chemical entities capable of changing fibrinogen production and test their impact on experimental thrombosis.

Dysregulation of Stemness Pathways in HPV Mediated Cervical Malignant Transformation Identifies Potential Oncotherapy Targets.

Human papillomavirus (HPV) infection is associated with a range of malignancies that affect anogenital and oropharyngeal sites. α-HPVs dominantly infect basal epithelial cells of mucosal tissues, where they dysregulate cell division and local immunity. The cervix is one of the mucosal sites most susceptible to HPV infections.

Tissue specific regulation of transcription in endometrium and association with disease.

Study Question: Are genetic effects on endometrial gene expression tissue specific and/or associated with reproductive traits and diseases?

Summary Answer: Analyses of RNA-sequence data and individual genotype data from the endometrium identified novel and disease associated, genetic mechanisms regulating gene expression in the endometrium and showed evidence that these mechanisms are shared across biologically similar tissues.

What Is Known Already: The endometrium is a complex tissue vital for female reproduction and is a hypothesized source of cells initiating endometriosis. Understanding genetic regulation specific to, and shared between, tissue types can aid the identification of genes involved in complex genetic diseases.

Dynamics of human monocytes and airway macrophages during healthy aging and after transplant.

The ontogeny of airway macrophages (AMs) in human lung and their contribution to disease are poorly mapped out. In mice, aging is associated with an increasing proportion of peripherally, as opposed to perinatally derived AMs. We sought to understand AM ontogeny in human lung during healthy aging and after transplant.

ascend: R package for analysis of single-cell RNA-seq data.

Background: Recent developments in single-cell RNA sequencing (scRNA-seq) platforms have vastly increased the number of cells typically assayed in an experiment. Analysis of scRNA-seq data is multidisciplinary in nature, requiring careful consideration of the application of statistical methods with respect to the underlying biology. Few analysis packages exist that are at once robust, are computationally fast, and allow flexible integration with other bioinformatics tools and methods.

A single-cell transcriptome atlas of the adult human retina.

The retina is a specialized neural tissue that senses light and initiates image processing. Although the functional organization of specific retina cells has been well studied, the molecular profile of many cell types remains unclear in humans. To comprehensively profile the human retina, we performed single-cell RNA sequencing on 20,009 cells from three donors and compiled a reference transcriptome atlas.

Single-cell RNA sequencing reveals cell type-specific HPV expression in hyperplastic skin lesions.

Human Papillomavirus infection is highly prevalent worldwide. While most types of HPV cause benign warts, some high-risk types are known to cause cervical cancer, as well as cancer of the oral cavity and head and neck. Persistent cutaneous HPV infection can be particularly problematic in patients with chronic immunosuppression, for example following organ transplantation.

Single-Cell Transcriptional Profiling of Aortic Endothelium Identifies a Hierarchy from Endovascular Progenitors to Differentiated Cells.

The cellular and molecular profiles that govern the endothelial heterogeneity of the circulatory system have yet to be elucidated. Using a data-driven approach to study the endothelial compartment via single-cell RNA sequencing, we characterized cell subpopulations within and assigned them to a defined endothelial hierarchy. We show that two transcriptionally distinct endothelial populations exist within the aorta and, using two independent trajectory analysis methods, confirm that they represent transitioning cells rather than discrete cell types.

Genetic regulation of methylation in human endometrium and blood and gene targets for reproductive diseases.

Background: Major challenges in understanding the functional consequences of genetic risk factors for human disease are which tissues and cell types are affected and the limited availability of suitable tissue. The aim of this study was to evaluate tissue-specific genotype-epigenetic characteristics in DNA samples from both endometrium and blood collected from women at different stages of the menstrual cycle and relate results to genetic risk factors for reproductive traits and diseases.

Results: We analysed DNA methylation (DNAm) data from endometrium and blood samples from 66 European women.

Single-Cell Transcriptomic Analysis of Cardiac Differentiation from Human PSCs Reveals HOPX-Dependent Cardiomyocyte Maturation.

Cardiac differentiation of human pluripotent stem cells (hPSCs) requires orchestration of dynamic gene regulatory networks during stepwise fate transitions but often generates immature cell types that do not fully recapitulate properties of their adult counterparts, suggesting incomplete activation of key transcriptional networks. We performed extensive single-cell transcriptomic analyses to map fate choices and gene expression programs during cardiac differentiation of hPSCs and identified strategies to improve in vitro cardiomyocyte differentiation. Utilizing genetic gain- and loss-of-function approaches, we found that hypertrophic signaling is not effectively activated during monolayer-based cardiac differentiation, thereby preventing expression of HOPX and its activation of downstream genes that govern late stages of cardiomyocyte maturation.

Transcriptomics and single-cell RNA-sequencing.

The past four decades have yielded advances in molecular biology allowing detailed characterization of the cellular genome and the transcriptome: the complete set of RNA species transcribed by a cell or tissue. Through transcriptomics and next-generation sequencing, we can now attain an unprecedented level of detail in understanding cellular phenotypes through examining the genes expressed in specific physiological and pathological states. In this review, we provide an overview of transcriptomics and RNA-sequencing in the analysis of whole tissue and single cells.

Genetic regulation of disease risk and endometrial gene expression highlights potential target genes for endometriosis and polycystic ovarian syndrome.

Gene expression varies markedly across the menstrual cycle and expression levels for many genes are under genetic control. We analyzed gene expression and mapped expression quantitative trait loci (eQTLs) in endometrial tissue samples from 229 women and then analyzed the overlap of endometrial eQTL signals with genomic regions associated with endometriosis and other reproductive traits. We observed a total of 45,923 cis-eQTLs for 417 unique genes and 2,968 trans-eQTLs affecting 82 unique genes.

Hepatic expression profiling identifies steatosis-independent and steatosis-driven advanced fibrosis genes.

Chronic liver disease (CLD) is associated with tissue-destructive fibrosis. Considering that common mechanisms drive fibrosis across etiologies, and that steatosis is an important cofactor for pathology, we performed RNA sequencing on liver biopsies of patients with different fibrosis stages, resulting from infection with hepatitis C virus (HCV) (with or without steatosis) or fatty liver disease. In combination with enhanced liver fibrosis score correlation analysis, we reveal a common set of genes associated with advanced fibrosis, as exemplified by those encoding the transcription factor ETS-homologous factor (EHF) and the extracellular matrix protein versican (VCAN).

Longitudinal expression profiling of CD4+ and CD8+ cells in patients with active to quiescent giant cell arteritis.

Background: Giant cell arteritis (GCA) is the most common form of vasculitis affecting elderly people. It is one of the few true ophthalmic emergencies but symptoms and signs are variable thereby making it a challenging disease to diagnose. A temporal artery biopsy is the gold standard to confirm GCA, but there are currently no specific biochemical markers to aid diagnosis.

Detection of HPV E7 Transcription at Single-Cell Resolution in Epidermis.

Persistent human papillomavirus (HPV) infection is responsible for at least 5% of human malignancies. Most HPV-associated cancers are initiated by the HPV16 genotype, as confirmed by detection of integrated HPV DNA in cells of oral and anogenital epithelial cancers. However, single-cell RNA sequencing may enable prediction of HPV involvement in carcinogenesis at other sites.