Dataset Information

Single-Cell RNA Sequencing of Mouse Fibroblasts Grown as Spheroids Reveals Presence of Mesenchymal Stem-Like Cells

ABSTRACT: Single-Cell RNA Sequencing of Mouse Fibroblasts Grown as Spheroids Reveals Presence of Mesenchymal Stem-Like Cells

PROVIDER: PRJNA1010165 | ENA |

REPOSITORIES: ENA

ACCESS DATA

Dataset's files

Source:

			Action	DRS
	SRR25777306_1.fastq.gz	Fastqsanger.gz
	SRR25777306_2.fastq.gz	Fastqsanger.gz
	SRR25777307_1.fastq.gz	Fastqsanger.gz
	SRR25777307_2.fastq.gz	Fastqsanger.gz

Items per page:

1 - 4 of 4

Similar Datasets

Project description:Compared to conventional monolayer cell culture, three-dimensional (or spheroid) cultures are more reflective of the in vivo environment and represent a better means of representing normal tissue. Therefore, understanding the biology of spheroid-derived cells is important for a more complete appreciation of in vivo tissue function. Although it has been shown that culture conditions such as nutrients, oxygen, cell passage etc. alter gene expression patterns of the cells, the gene expression profile of spheroid as compared to conventional monolayer culture in primary cells has not been studied. To investigate the effects of spheroid culture system on gene expression of primary cells, single cell RNA sequencing (scRNAseq) was performed on mouse dermal fibroblasts (tail/ear fibroblasts, TEFs) cultured as monolayers on cell culture dishes or grown as spheroids. After quality control, a total of 11,515 cells (8,022 spheroid cells and 3,493 cells of monolayer fibroblasts) were analyzed using Uniform Manifold Approximation and Projection (UMAP) which identified 2 largely separated libraries. These libraries contained cells with 2,491 and 8722 detected genes (nFeature_RNA) for monolayer and spheroid cells respectively. UMAP clustering of the integrated datasets identified 8 significant cell clusters (resolution=0.3) in which clusters 0, 1, 2, 4 and 7 had a higher number of cells in the spheroid library. Differential gene expression analyses revealed that each cluster was characterized by a specific transcriptional profile. Among cell clusters, cluster 4 exhibited significantly increased expression of collagen family genes, including Col1a2, Col3a1, Col1a1, Col5a2, Col4a, Col4a2, Col5a3, Col5a1, Col6a1, Col12a1 and Col15a1. Furthermore, expression of mesenchymal stem cell genes [Sca-1 (Ly6a), CD29 (Itgb1), CD44, CD90.1 (Thy1)] as well as a group of genes important for stem cell self-renewal including Notch2, Sox4, Sox9 Klf2, and Foxp1 were significantly increased in cluster 4. Immunofluorescence (IF) staining was performed to validate the protein expression of selected genes. Our work provides a significant advance in characterizing the global gene expression profile of spheroid culture of mouse TEFs compared to monolayer culture. We reached a high resolution at single-cell level, which enabled us to identify specific clusters associated with mesenchymal stem-like cells.

Dataset Information

Single-Cell RNA Sequencing of Mouse Fibroblasts Grown as Spheroids Reveals Presence of Mesenchymal Stem-Like Cells

Dataset's files

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets