Purification of pancreatic endocrine subsets from mouse islets
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE149488 | GEO | 2020/04/29
REPOSITORIES: GEO
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Project description:Purification of pancreatic endocrine subsets from mouse islets
| PRJNA628871 | ENA
Project description:This study aimed to evaluate the efficacy of a purification method developed for isolating alpha, beta, and delta cells from pancreatic islets of adult mice, extending its application to islets from newborn and aged mice. Furthermore, it sought to examine transcriptome dynamics in mouse pancreatic endocrine islet cells throughout postnatal development and to evaluate age-related alterations in intercellular communication within these cell populations. We leveraged the high surface expression of CD71 on beta cells and CD24 on delta cells to FACS-purify alpha, beta, and delta cells from newborn (1-week-old), adult (12-week-old), and old (18-month-old) mice. Bulk RNA sequencing was conducted on these purified cell populations, and subsequent bioinformatic analyses included differential gene expression, overrepresentation, intersection, and intercellular communication analysis. Alpha, beta, and delta cells from newborn and aged mice were successfully FACS-purified using the same method employed for adult mice. Our analysis of the age-related transcriptional changes in alpha, beta, and delta cell populations revealed a decrease in cell cycling and an increase in so-called “neurogenesis” processes during the transition from newborn to adult mice. Progressing from adult to old mice, we observed an increase in β-2 microglobulin and major histocompatibility complex (MHC) Class I expression. Computational modeling of cell interactions suggested shifts in TGFβ and BMP signaling underpinning these age-related changes. Our study demonstrates the effectiveness of our cell sorting technique in purifying endocrine subsets from mouse islets at different ages. We provide a valuable resource for better understanding endocrine pancreas aging and identified increased β-2 microglobulin and MHC Class I expression as a common hallmark of old alpha, beta, and delta cells, with potential implications for immune response regulation.
2024-06-27 | GSE269616 | GEO
Project description:The peptide hormone Urocortin3 (Ucn3) is abundantly expressed by mature beta cells, yet its physiological role is unknown. Transcriptome data from Ucn3 null islets compared to wild type controls reveals selective depletion for delta cell markers and supports a role for Ucn3 as a trigger for somatostatin-mediated negative feedback. Isolated islets from Ucn3 knockout animals compared to wild type littermate controls. Islets from three indivual males were used for each genotype, lysed in Trizol for RNA isolation and library construction.
2015-05-22 | E-GEOD-58286 | biostudies-arrayexpress
Project description:Pluripotent stem cell-derived islets (hPSC-islets) are a promising cell resource for diabetes treatment. Here, we demonstrate that transplantation of pluripotent stem cell-derived islets into diabetic nonprimates effectively restored endogenous insulin secretion and improved glycemic control. Single-cell RNA sequencing analysis of S6D2 clusters confirmed the existence of the three major pancreatic endocrine cell populations (β cells, α-like cells and δ-like cells) and their proportions, which altogether accounted for 80%. Importantly, hierarchical clustering of S6D2 hCiPSC-islets, 10 wpt kidney grafts and primary islets showed that the hCiPSC differentiated pancreatic endocrine cells shared similar global gene expression profiles to their native counterparts in primary islets. Single-cell RNA sequencing analysis on PBMCs revealed the potential immune response of recipient macaque to hCiPSC-islets.
2022-02-04 | GSE185036 | GEO
Project description:Human pluripotent stem cell-derived islets (hPSC-islets) are a promising cell resource for diabetes treatment. Here, we demonstrate that transplantation of human pluripotent stem cell-derived islets into diabetic nonhuman primates effectively restored endogenous insulin secretion and improved glycemic control. Single-cell RNA sequencing analysis of S6D2 clusters confirmed the existence of the three major pancreatic endocrine cell populations (β cells, α-like cells and δ-like cells) and their proportions, which altogether accounted for 80%. Importantly, hierarchical clustering of S6D2 hCiPSC-islets, 10 wpt kidney grafts and primary human islets showed that the hCiPSC differentiated pancreatic endocrine cells shared similar global gene expression profiles to their native counterparts in primary human islets. Single-cell RNA sequencing analysis on PBMCs revealed the potential immune response of recipient macaque to hCiPSC-islets.
2022-02-04 | GSE185035 | GEO
Project description:Human pluripotent stem cell-derived islets (hPSC-islets) are a promising cell resource for diabetes treatment. Here, we demonstrate that transplantation of human pluripotent stem cell-derived islets into diabetic nonhuman primates effectively restored endogenous insulin secretion and improved glycemic control. Single-cell RNA sequencing analysis of S6D2 clusters confirmed the existence of the three major pancreatic endocrine cell populations (β cells, α-like cells and δ-like cells) and their proportions, which altogether accounted for 80%. Importantly, hierarchical clustering of S6D2 hCiPSC-islets, 10 wpt kidney grafts and primary human islets showed that the hCiPSC differentiated pancreatic endocrine cells shared similar global gene expression profiles to their native counterparts in primary human islets. Single-cell RNA sequencing analysis on PBMCs revealed the potential immune response of recipient macaque to hCiPSC-islets.
2022-02-04 | GSE185037 | GEO
Project description:To define genetic pathways that regulate development of the endocrine pancreas, we generated transcriptional profiles of enriched cells isolated from four biologically significant stages of endocrine pancreas development: endoderm before pancreas specification, early pancreatic progenitor cells, endocrine progenitor cells and adult islets of Langerhans. These analyses implicate new signaling pathways in endocrine pancreas development, and identified sets of known and novel genes that are temporally regulated, as well as genes that spatially define developing endocrine cells from their neighbors. The differential expression of several genes from each time point was verified by RT-PCR and in situ hybridization. Moreover, we present preliminary functional evidence suggesting that one transcription factor encoding gene (Myt1), which was identified in our screen, is expressed in endocrine progenitors and may regulate alpha, beta and delta cell development. In addition to identifying new genes that regulate endocrine cell fate, this global gene expression analysis has uncovered informative biological trends that occur during endocrine differentiation.
2006-06-30 | E-CBIL-14 | biostudies-arrayexpress
Project description:Identification of protein complexes for the synaptotagmin 13 protein from MDCK cells by Strep affinity purificaiton and LFQ mass spectrometry. Epithelial cell egression is important for organ development and cell differentiation, but also drives cancer metastasis. The tightly connected pancreatic epithelial differentiation and morphogenesis generate islets of Langerhans. However, the morphogenetic drivers and molecular mechanisms are largely unresolved. Here we identify the uncharacterized Synaptotagmin 13 (Syt13) as a major regulator of endocrine cell egression and islet morphogenesis and differentiation. We detected upregulation of Syt13 in endocrine precursors that associates with increased expression of several unique cytoskeletal components. High-resolution imaging reveals a previously unidentified apical-basal to front-rear repolarization during endocrine cell egression. Strikingly, Syt13 directly interacts with acetylated tubulin and phosphoinositide phospholipids to be recruited to the leading edge of egressing cells. Knockout of Syt13 discloses the impairment in endocrine cell egression and skews the α-to-β-cell ratio. At mechanistic levels, Syt13 regulates protein endocytosis to remodel the basement membrane and modulate cell-matrix adhesion at the leading edge of egressing endocrine cells. Altogether, these findings implicate that Ca2+-independent atypical Syt13 vesicular protein functions in regulating cell polarity to orchestrate endocrine cell egression and tissue morphogenesis.
2023-12-28 | PXD026698 | Pride
Project description:Rodent models are widely used to study diabetes. Yet, significant gaps remain in our understanding of mouse islet physiology. We generated comprehensive transcriptomes of mouse delta, beta and alpha cells using two separate triple transgenic mouse models generated for this purpose. This enables systematic comparison across thousands of genes between the three major endocrine cell types of the islets of Langerhans whose principal hormones control nutrient homeostasis. FACS purified delta or alpha cells and beta cells from the same islets. Islets were isolated from triple transgenic offspring of a cross between mIns1-H2b-mCherry (Jax # 028589) and either Sst-Cre (delta) or Gcg-cre (alpha) cells and a floxed YFP allele to label delta or alpha cells, respectively. Islets from replicate groups of 10 to 12 triple transgenic animals for each group were pooled by sex to obtain sufficient material. Pooled islets were dissociated, sorted and collect in Trizol for RNA isolation and library construction.
2016-06-26 | E-GEOD-80673 | biostudies-arrayexpress
Project description:Current approaches to profile the single-cell transcriptomics of human pancreatic endocrine cells almost exclusively rely on freshly isolated islets. However, human islets are limited in availability. Furthermore, the extensive processing steps during islet isolation and subsequent single cell dissociation might alter gene expressions. In this work, we cross-compared five nuclei isolation protocols and selected the citric acid method as the best strategy to isolate nuclei with high RNA integrity and low cytoplasmic contamination from human pancreata. We innovated fluorescence-activated nuclei sorting (FANS) based on the positive signal of NKX2-2 antibody to enrich for nuclei of the endocrine population from the entire nuclei pool of the pancreas. Our sample preparation procedure generated high-quality single-nucleus gene-expression libraries while preserving the endocrine population diversity. We observed comparable endocrine cellular composition and cell type signature gene expression between our snRNA-seq libraries and conventional scRNA-seq libraries generated with live cells from freshly isolated human islets. Our work fills a technological gap and helps to unleash archival pancreatic tissue for molecular profiling targeting the endocrine population. We expect that our protocol can be used to enrich nuclei for transcriptomics study from various populations in the pancreas and in different organs/tissues.
2024-04-01 | GSE252614 | GEO