scholarly journals Endocrine Pancreas Development and Dysfunction Through the Lens of Single-Cell RNA-Sequencing

2021 ◽  
Vol 9 ◽  
Author(s):  
Wojciech J. Szlachcic ◽  
Natalia Ziojla ◽  
Dorota K. Kizewska ◽  
Marcelina Kempa ◽  
Malgorzata Borowiak
Keyword(s):  
Blood Glucose ◽  
Single Cell ◽  
Rna Sequencing ◽  
Endocrine Pancreas ◽  
Pancreas Development ◽  
Β Cells ◽  
Β Cell ◽  
Regulatory Pathways ◽  
Endocrine Differentiation ◽  
Single Cell Rna Sequencing

A chronic inability to maintain blood glucose homeostasis leads to diabetes, which can damage multiple organs. The pancreatic islets regulate blood glucose levels through the coordinated action of islet cell-secreted hormones, with the insulin released by β-cells playing a crucial role in this process. Diabetes is caused by insufficient insulin secretion due to β-cell loss, or a pancreatic dysfunction. The restoration of a functional β-cell mass might, therefore, offer a cure. To this end, major efforts are underway to generate human β-cells de novo, in vitro, or in vivo. The efficient generation of functional β-cells requires a comprehensive knowledge of pancreas development, including the mechanisms driving cell fate decisions or endocrine cell maturation. Rapid progress in single-cell RNA sequencing (scRNA-Seq) technologies has brought a new dimension to pancreas development research. These methods can capture the transcriptomes of thousands of individual cells, including rare cell types, subtypes, and transient states. With such massive datasets, it is possible to infer the developmental trajectories of cell transitions and gene regulatory pathways. Here, we summarize recent advances in our understanding of endocrine pancreas development and function from scRNA-Seq studies on developing and adult pancreas and human endocrine differentiation models. We also discuss recent scRNA-Seq findings for the pathological pancreas in diabetes, and their implications for better treatment.

scholarly journals Single-cell RNA sequencing reveals a role for reactive oxygen species and peroxiredoxins in fatty-acid-induced rat β-cell proliferation

2021 ◽  
Author(s):  
Alexis Vivoli ◽  
Julien Ghislain ◽  
Ali Filali-Mouhim ◽  
Zuraya Elisa Angeles ◽  
Anne-Laure Castell ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Cell Proliferation ◽  
Single Cell ◽  
Rna Sequencing ◽  
Molecular Mechanisms ◽  
Reactive Oxygen ◽  
Oxygen Species ◽  
Β Cells ◽  
Β Cell ◽  
Single Cell Rna Sequencing

The functional mass of insulin-secreting pancreatic β cells expands to maintain glucose homeostasis in the face of nutrient excess, in part via replication of existing β cells. To decipher the underlying molecular mechanisms, we assessed β-cell proliferation in isolated rat islets exposed to glucose and oleate or palmitate for 48 h and analyzed the transcriptional response by single-cell RNA sequencing. Unsupervised clustering of pooled βcells identified subpopulations, including proliferating β cells. β-cell proliferation increased in response to oleate but not palmitate. Both fatty acids enhanced the expression of genes involved energy metabolism and mitochondrial activity. Comparison of proliferating vs. non-proliferating β cells and pseudotime ordering suggested the involvement of reactive oxygen species (ROS) and peroxiredoxin signaling. Accordingly, the antioxidant N-acetyl cysteine and the peroxiredoxin inhibitor Conoidin A both blocked oleate-induced β-cell proliferation. Our data reveal a key role for ROS signaling in β-cell proliferation in response to nutrients.

scholarly journals Ins2 gene bursting activity defines a mature β-cell state

2019 ◽  
Author(s):  
Honey Modi ◽  
Søs Skovsø ◽  
Cara Ellis ◽  
Nicole A.J. Krentz ◽  
Yiwei Bernie Zhao ◽  
...  
Keyword(s):  
Single Cell ◽  
Rna Sequencing ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Gene Activity ◽  
Insulin Gene ◽  
Insulin Production ◽  
Β Cells ◽  
Β Cell ◽  
Single Cell Rna Sequencing

AbstractHeterogeneity within specific cell types is common and increasingly apparent with the advent of single-cell transcriptomics. Transcriptional and functional cellular specialization has been described for insulin-secreting β-cells of the endocrine pancreas, including so-called extreme β-cells exhibiting >2 fold higher insulin gene activity. However, it is not yet clear whether β-cell heterogeneity is stable or reflects dynamic cellular states. We investigated the temporal kinetics of endogenous insulin gene activity using live-cell imaging, with complementary experiments employing FACS and single-cell RNA sequencing, in β-cells from Ins2GFP knock-in mice. In vivo staining and FACS analysis of islets from Ins2GFP mice confirmed that at a given moment, ∼25% of β-cells exhibited significantly higher activity at the conserved insulin gene Ins2(GFP)HIGH. Live-cell imaging captured on and off ‘bursting’ behaviour in single β-cells that lasted hours to days. Single cell RNA sequencing determined that Ins2(GFP)HIGH β-cells were enriched for markers of β-cell maturity and had reduced expression of anti-oxidant genes. Ins2(GFP)HIGH β-cells were also significantly less viable at all glucose concentrations and in the context of ER stress. Collectively, our results demonstrate that the heterogeneity of extreme insulin production, observed in mouse and human β-cells, can be accounted for by dynamic states of insulin gene activity. Our observations define a previously uncharacterized form of β-cell plasticity. Understanding the dynamics of insulin production has relevance for understanding the pathobiology of diabetes and for regenerative therapy research.

scholarly journals NEUROD1 Is Required for the Early α and β Endocrine Differentiation in the Pancreas

2021 ◽  
Vol 22 (13) ◽  
pp. 6713
Author(s):  
Romana Bohuslavova ◽  
Ondrej Smolik ◽  
Jessica Malfatti ◽  
Zuzana Berkova ◽  
Zaneta Novakova ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Cell Fate ◽  
Cell Mass ◽  
Neonatal Diabetes ◽  
Diabetes Research ◽  
Pancreas Development ◽  
Β Cells ◽  
Β Cell ◽  
Pancreatic Islet Cell ◽  
Endocrine Differentiation

Diabetes is a metabolic disease that involves the death or dysfunction of the insulin-secreting β cells in the pancreas. Consequently, most diabetes research is aimed at understanding the molecular and cellular bases of pancreatic development, islet formation, β-cell survival, and insulin secretion. Complex interactions of signaling pathways and transcription factor networks regulate the specification, growth, and differentiation of cell types in the developing pancreas. Many of the same regulators continue to modulate gene expression and cell fate of the adult pancreas. The transcription factor NEUROD1 is essential for the maturation of β cells and the expansion of the pancreatic islet cell mass. Mutations of the Neurod1 gene cause diabetes in humans and mice. However, the different aspects of the requirement of NEUROD1 for pancreas development are not fully understood. In this study, we investigated the role of NEUROD1 during the primary and secondary transitions of mouse pancreas development. We determined that the elimination of Neurod1 impairs the expression of key transcription factors for α- and β-cell differentiation, β-cell proliferation, insulin production, and islets of Langerhans formation. These findings demonstrate that the Neurod1 deletion altered the properties of α and β endocrine cells, resulting in severe neonatal diabetes, and thus, NEUROD1 is required for proper activation of the transcriptional network and differentiation of functional α and β cells.

scholarly journals Maternal insulin resistance and transient hyperglycemia impact the metabolic and endocrine phenotypes of offspring

2014 ◽  
Vol 307 (10) ◽  
pp. E906-E918 ◽  
Author(s):  
Sevim Kahraman ◽  
Ercument Dirice ◽  
Dario F. De Jesus ◽  
Jiang Hu ◽  
Rohit N. Kulkarni
Keyword(s):  
Insulin Resistance ◽  
Cell Proliferation ◽  
Blood Glucose ◽  
Endocrine Pancreas ◽  
Cell Mass ◽  
Postnatal Life ◽  
Pancreas Development ◽  
Β Cell ◽  
Early Postnatal Life ◽  
The Impact

Studies in both humans and rodents suggest that maternal diabetes leads to a higher risk of the fetus developing impaired glucose tolerance and obesity during adulthood. However, the impact of hyperinsulinemia in the mother on glucose homeostasis in the offspring has not been fully explored. We aimed to determine the consequences of maternal insulin resistance on offspring metabolism and endocrine pancreas development using the LIRKO mouse model, which exhibits sustained hyperinsulinemia and transient increase in blood glucose concentrations during pregnancy. We examined control offspring born to either LIRKO or control mothers on embryonic days 13.5, 15.5, and 17.5 and postpartum days 0, 4, and 10. Control offspring born to LIRKO mothers displayed low birth weights and subsequently rapidly gained weight, and their blood glucose and plasma insulin concentrations were higher than offspring born to control mothers in early postnatal life. In addition, concentrations of plasma leptin, glucagon, and active GLP-1 were higher in control pups from LIRKO mothers. Analyses of the endocrine pancreas revealed significantly reduced β-cell area in control offspring of LIRKO mothers shortly after birth. β-Cell proliferation and total islet number were also lower in control offspring of LIRKO mothers during early postnatal days. Together, these data indicate that maternal hyperinsulinemia and the transient hyperglycemia impair endocrine pancreas development in the control offspring and induce multiple metabolic alterations in early postnatal life. The relatively smaller β-cell mass/area and β-cell proliferation in these control offspring suggest cell-autonomous epigenetic mechanisms in the regulation of islet growth and development.

scholarly journals Single-Cell RNA Sequencing Efficiently Predicts Transcription Factor Targets in Plants

2020 ◽  
Vol 11 ◽  
Author(s):  
Yunjie Xie ◽  
Shenfei Jiang ◽  
Lele Li ◽  
Xiangzhen Yu ◽  
Yupeng Wang ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Single Cell ◽  
Rna Sequencing ◽  
Expression Profiles ◽  
Expression System ◽  
Strong Correlations ◽  
Regulatory Pathways ◽  
Sequencing Method ◽  
Single Cell Rna Sequencing ◽  
Gene Overlapping

Discovering transcription factor (TF) targets is necessary for the study of regulatory pathways, but it is hampered in plants by the lack of highly efficient predictive technology. This study is the first to establish a simple system for predicting TF targets in rice (Oryza sativa) leaf cells based on 10 × Genomics’ single-cell RNA sequencing method. We effectively utilized the transient expression system to create the differential expression of a TF (OsNAC78) in each cell and sequenced all single cell transcriptomes. In total, 35 candidate targets having strong correlations with OsNAC78 expression were captured using expression profiles. Likewise, 78 potential differentially expressed genes were identified between clusters having the lowest and highest expression levels of OsNAC78. A gene overlapping analysis identified 19 genes as final candidate targets, and various assays indicated that Os01g0934800 and Os01g0949900 were OsNAC78 targets. Additionally, the cell profiles showed extremely similar expression trajectories between OsNAC78 and the two targets. The data presented here provide a high-resolution insight into predicting TF targets and offer a new application for single-cell RNA sequencing in plants.

scholarly journals Revealing Cellular Heterogeneity and In Vitro Differentiation Trajectory of Cultured Human Endometrial Mesenchymal-like Stem Cells Using Single-cell RNA Sequencing

2020 ◽  
Author(s):  
Dandan Cao ◽  
Rachel W.S. Chan ◽  
Ernest H.Y. Ng ◽  
Kristina Gemzell-Danielsson ◽  
William S.B. Yeung
Keyword(s):  
Stem Cells ◽  
Single Cell ◽  
Rna Sequencing ◽  
Adult Stem Cells ◽  
Cellular Heterogeneity ◽  
In Vitro Differentiation ◽  
Regulatory Pathways ◽  
Single Cell Rna Sequencing ◽  
Unbiased Manner

ABSTRACTEndometrial mesenchymal-like stem cells (eMSCs) are adult stem cells contributing to endometrial regeneration. One set of perivascular markers (CD140b+CD146+) have been widely used to enrich eMSCs. Although eMSCs are easily accessible for regenerative medicine and have long been studied, their cellular heterogeneity and molecular program controlling their expansion and differentiation in vitro remains largely unclear. In this study, we applied 10X genomics single-cell RNA sequencing to eMSCs cultured in vitro after microbeading from 7 donors to investigate cellular heterogeneity in an unbiased manner. Corresponding clonogenic progenies of eMSCs after culture for 14 days were also sequenced to construct the in vitro differentiation trajectory of eMSCs. Transcriptomic expression based clustering revealed several subpopulations in eMSCs. Each subpopulation manifested distinct functional characteristics associated with immunomodulation, proliferation, extracellular matrix organization and cell differentiation. Pseudotime trajectory analysis on eMSCs and their differentiated progenies identified in vitro differentiation hierarchy of eMSCs. Further ligand-receptor pair analysis found that WNT signaling, NOTCH signaling, TGF-beta signaling and FGF signaling were important regulatory pathways for eMSC self-renewal and differentiation. By comparing eMSCs to Wharton’s Jelly MSCs and adipose-derived MSCs, we found these 3 kinds of MSCs expressed largely overlapping differentiation (CD) genes and highly variable genes. In summary, we reveal for the first time high molecular and cellular heterogeneity in cultured eMSCs, and identify the key signaling pathways that may be important for eMSC differentiation.

scholarly journals Single-cell RNA sequencing of mouse islets exposed to proinflammatory cytokines

2021 ◽  
Vol 4 (6) ◽  
pp. e202000949
Author(s):  
Jennifer S Stancill ◽  
Moujtaba Y Kasmani ◽  
Achia Khatun ◽  
Weiguo Cui ◽  
John A Corbett
Keyword(s):  
Gene Expression ◽  
Single Cell ◽  
Rna Sequencing ◽  
Proinflammatory Cytokines ◽  
Cell Damage ◽  
Cytokine Signaling ◽  
Β Cells ◽  
Antiviral Genes ◽  
Single Cell Rna Sequencing ◽  
Mouse Islets

Exposure to proinflammatory cytokines is believed to contribute to pancreatic β-cell damage during diabetes development. Although some cytokine-mediated changes in islet gene expression are known, the heterogeneity of the response is not well-understood. After 6-h treatment with IL-1β and IFN-γ alone or together, mouse islets were subjected to single-cell RNA sequencing. Treatment with both cytokines together led to expression of inducible nitric oxide synthase mRNA (Nos2) and antiviral and immune-associated genes in a subset of β-cells. Interestingly, IL-1β alone activated antiviral genes. Subsets of δ- and α-cells expressed Nos2 and exhibited similar gene expression changes as β-cells, including increased expression of antiviral genes and repression of identity genes. Finally, cytokine responsiveness was inversely correlated with expression of genes encoding heat shock proteins. Our findings show that all islet endocrine cell types respond to cytokines, IL-1β induces the expression of protective genes, and cellular stress gene expression is associated with inhibition of cytokine signaling.

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