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Transcriptional and epigenetic program changes of induced beta cells over time [RRBS-seq]

ABSTRACT: Direct lineage conversion is a promising approach to generate therapeutically important cell types for disease modeling and tissue repair. However, it is often unclear whether lineage-reprogrammed cells remain stable long-term and whether the properties of the reprogrammed cells evolve over time. Here, using an improved method of converting pancreatic acinar cells to beta-cells, we show that induced beta-cells persist in the adult pancreas for up to 14 months and form islet-like structures. Detailed analyses of induced cells over 7 months reveal that global DNA methylation changes occur rapidly whereas transcription network remodeling evolves over two months to resemble that of endogenous beta-cells and then stabilizes thereafter. Progressive gain of beta-cell function by converted cells during the 7 month period coincides with both transcriptional changes and the formation of islet-like structures. These studies demonstrate the ability of lineage-reprogrammed cells to achieve a stable state and identify key cellular and molecular milestones during their long-term evolution.

ORGANISM(S): Mus musculus

PROVIDER: GSE59615 | GEO | 2014/11/15

SECONDARY ACCESSION(S): PRJNA255762

REPOSITORIES: GEO

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Project description:Objectives The streptozotocin (STZ) model is widely used in diabetes research. However, the cellular and molecular states of pancreatic endocrine cells in this model remain unclear. This study explored the molecular characteristics of islet cells treated with STZ and re-evaluated β-cell dysfunction and regeneration in the STZ model. Methods We performed single-cell RNA sequencing of pancreatic endocrine cells from STZ-treated mice. High-quality sequencing data from 2,999 cells were used to identify clusters via Louvain clustering analysis. Principal component analysis (PCA), t-distributed stochastic neighbor embedding (t-SNE), uniform manifold approximation and projection (UMAP), force-directed layout (FDL), and differential expression analysis were used to define the heterogeneity and transcriptomic changes in islet cells. In addition, qPCR and immunofluorescence staining were used to confirm findings from the sequencing data. Results Untreated β-cells were divided into two populations at the transcriptomic level, a large high-Glut2 expression (Glut2high) population and a small low-Glut2 expression (Glut2low) population. At the transcriptomic level, Glut2low β-cells in adult mice did not represent a developmentally immature state, although a fraction of genes associated with β-cell maturation and function were downregulated in Glut2low cells. After a single high-dose STZ treatment, most Glut2high cells were killed, but Glut2low cells survived and over time changed to a distinct cell state. We did not observe conversion of Glut2low to Glut2high β-cells up to 9 months after STZ treatment. In addition, we did not detect transcriptomic changes in the non-β endocrine cells or a direct trans-differentiation pathway from the α-cell lineage to the β-cell lineage in the STZ model. Conclusions We identified the heterogeneity of β-cells in both physiological and pathological conditions. However, we did not observe conversion of Glut2low to Glut2high β-cells, transcriptomic changes in the non-β endocrine cells, or direct trans-differentiation from the α-cell lineage to the β-cell lineage in the STZ model. Our results clearly define the states of islet cells treated with STZ and allow us to re-evaluate the STZ model widely used in diabetes studies.

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Transcriptional and epigenetic program changes of induced beta cells over time [RRBS-seq]

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