Single-cell RNA sequencing of ST2 mouse cell line
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ABSTRACT:
PROVIDER: PRJNA1008706 | ENA |
REPOSITORIES: ENA
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Project description:Colorectal cancer (CRC) was induced in Foxp3/eGFP reporter mice by the azoxymethane/dextran sulphate sodium salt (AOM/DSS) protocol. Mice were injected i.p. with the procarcinogen AOM (12.5 mg/kg of body weight). After 1 week, mice received drinking water supplemented with 2.5% DSS for 5 to 7 days, followed by 2 weeks of regular water. The DSS administration was repeated twice with 2% DSS. Mice were sacrificed at week 11 and lamina propia lymphocytes (LPLs) from the colon were isolated. CD4+FOXP3+ (eGFP+) ST2+ or ST2- Tregs were separated from colonic LPLs of CRC induced mice using a FACSAria II cell sorter. Microarray analysis was performed to analyze if ST2+ FOXP3+ Tregs from the colon of CRC mice present a distinct transcription pattern compared to ST2- FOXP3+ Tregs. By this, the role of ST2 for Treg function during intestinal tumorigenesis should be characterized.
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Project description:In diabetics, methylglyoxal (MG), a glucose-derived metabolite, plays a noxious role by inducing oxidative stress, which causes and exacerbates a series of complications. With the use of microarray analysis, we comprehensively screened the gene expression profiles of ST2 cells, derived from a multipotent bone marrow stromal cell line, in the presence or absence of oxidative stress induced by100 M-NM-<M methylglyoxal (MG) treatment to charactrize genes related to diabetic complications. Mouse bone marrow stromal cell-line ST2 (RIKEN, Tsukuba, Japan) was cultured in M-NM-1-MEM (Sigma, St. Louis, MO) supplemented with 10% FBS (Sigma), 100 M-NM-<g/ ml penicillin/ streptomycin (ICN Biomedicals, Inc., Aurora, OH) and 100 M-NM-<M methylglyoxal (MG), and maintained at 37M-BM-0C in a humidified atmosphere with 5% CO2. Total RNA was isolated from ST2 cells treated with or without 100 M-NM-<M methylglyoxal (MG) or 1 M-NM-<M of 5-aza-2M-bM-^@M-^Y-deoxycytidine (5-aza-dC) by standard methods with the use of an RNeasy Protect Mini kit (Qiagen KK, Tokyo, Japan) according to the manufacturerM-bM-^@M-^Ys instructions. Cy3, Cy5 differential expression asasy by Mouse 32K Filgen array
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Project description:Foxp3+ regulatory T cells (Tregs) are critical mediators of peripheral tolerance and immune homeostasis. Tregs that express the IL-33 receptor ST2 are enriched in peripheral nonlymphoid tissues and can exert a variety of tissue-specific functions from metabolic regulation within adipose tissue to skeletal muscle repair. However, the relationship between ST2+ and ST2- Tregs within and across different tissues remains unclear. To compare murine ST2- and ST2+ Tregs within and across tissues, we performed RNA sequencing (RNAseq) of ST2-CD44hi and ST2+CD44hi Tregs from blood, spleen, lungs, visceral adipose tissue (VAT), colon, and skin. RNAseq was also performed on ST2- CD44lo CD62L+ Tregs from the spleen and lungs. We found that the tissue microenvironment was the major factor shaping the transcriptome of Tregs across tissues. Across the tissues studied, Treg transcriptomes displayed an ordered hierarchy that may represent graded levels of activation or differentiation across tissues. We also identified a core signature that distinguished ST2+ Tregs from ST2- Tregs across tissues and a large number of differentially expressed genes between ST2- and ST2+ Tregs within individual tissues that could support the tissue-specific adaptation and function of ST2+ Tregs. In summary, our work highlights the unique, tissue-specific phenotype of ST2+ Tregs and reveals a core ST2+ Treg transcriptional signature shared across tissues.
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