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Cross-species single-cell transcriptomic analysis reveals the landscape of animal gastric antrum [RNA-Seq]

ABSTRACT: Purpose: One goal of the bulk RNA-seq of organoids is to verify the differentially expressed genes between species identified by scRNA-seq .The other goal is to detected the differentially expressed genes when organoids from different species were treated with TNFα Methods: The total RNA from organoids was extracted by RNeasy Mini Kit (Qiagen). After total RNA extraction and then subjected to high-throughput sequencing on an Illumina Novaseq 6000 platform. Results: Most of the differentially expressed genes between species identified by bulk RNA-seq are consistent with the genes filtered by scRNA-seq. Conclusions: F3 and CLCA1 show higher expression in organoids of pig antral epithelium than in other three species. TNFα treatment upregulated the expression of immune response genes in all species

ORGANISM(S): Mus musculus Rattus norvegicus Sus scrofa Homo sapiens

PROVIDER: GSE225274 | GEO | 2023/09/11

REPOSITORIES: GEO

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Project description:Pancreatic cancer is a complex disease with a desmoplastic stroma, extreme hypoxia, and inherent resistance to therapy. Understanding the signaling and adaptive response of such an aggressive cancer is key to making advances in therapeutic efficacy and understanding disease progression. Redox factor-1 (Ref-1), a redox signaling protein, regulates the DNA binding activity of several transcription factors, including HIF-1. The conversion of HIF-1 from an oxidized to reduced state leads to enhancement of its DNA binding. In our previously published work, knockdown of Ref-1 under normoxia resulted in altered gene expression patterns on pathways including EIF2, protein kinase A, and mTOR. In this study, single cell RNA sequencing (scRNA-seq) and proteomics were used to explore the effects of Ref-1 on metabolic pathways under hypoxia.Results: We also integrated the scRNA data analysis with the proteomic analysis and found that the differentially expressed genes and pathways identified from the scRNA-seq data are highly consistent to the significant proteins observed in the proteomics data, especially for the upregulated cell cycle and transcription pathways and downregulated metabolic, apoptosis and signaling pathways under hypoxia. Conclusion: The scRNA-seq and proteomics data consistently demonstrated down-regulated central metabolism pathways in APE1/Ref-1 knockdown vs scrambled control under both normoxia and hypoxia conditions. Experimental Methods: scRNA-seq comparing pancreatic cancer cells expressing less than 20% of the Ref-1 protein was analyzed using left truncated mixture Gaussian model. Matched samples were also collected for bulk proteomic analysis of the four conditions. scRNA-seq data was validated using proteomics and qRT-PCR. Ref-1’s role in mitochondrial function was confirmed using mitochondrial function assays and qRT-PCR. Results: We also integrated the scRNA data analysis with the proteomic analysis and found that the differentially expressed genes and pathways identified from the scRNA-seq data are highly consistent to the significant proteins observed in the proteomics data, especially for the upregulated cell cycle and transcription pathways and downregulated metabolic, apoptosis and signaling pathways under hypoxia. Conclusion: The scRNA-seq and proteomics data consistently demonstrated down-regulated central metabolism pathways in APE1/Ref-1 knockdown vs scrambled control under both normoxia and hypoxia conditions.

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Cross-species single-cell transcriptomic analysis reveals the landscape of animal gastric antrum [RNA-Seq]

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