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Title: Purification of replicating pancreatic β-cells for gene expression studies
Author: Carballar, Reyes
Canyelles, Maria de Lluc
Fernández, Claudia
Martí, Yasmina
Bonnin, Sarah
Castaño Boldú, Esther
Montanya Mias, Eduard
Téllez i Besolí, Noèlia
Keywords: Expressió gènica
Malalties del pàncrees
Gene expression
Pancréas diseases
Issue Date: 13-Dec-2017
Publisher: Nature Publishing Group
Abstract: β-cell proliferation is a rare event in adult pancreatic islets. To study the replication-related β-cell biology we designed a replicating β-cells sorting system for gene expression experiments. Replicating β-cells were identifed by EdU incorporation and purifed by fow cytometry. For β-cell separation islet cells were sorted by size, granularity and Newport Green fuorescence emission that was combined with emitted fuorescence for EdU-labelled replicating cells sorting. The purity of the resulting sorted populations was evaluated by insulin staining and EdU for β-cell identifcation and for replicating cells, respectively. Total RNA was isolated from purifed cell-sorted populations for gene expression analysis. Cell sorting of dispersed islet cells resulted in 96.2% purity for insulin positivity in the collected β-cell fraction and 100% efciency of the EdU-based cell separation. RNA integrity was similar between FACSsorted replicating and quiescent β-cells. Global transcriptome analysis of replicating vs quiescent β-cells showed the expected enrichment of categories related to cell division and DNA replication. Indeed, key genes in the spindle check-point were the most upregulated genes in replicating β-cells. This work provides a method that allows for the isolation of replicating β-cells, a very scarce population in adult pancreatic islets.
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It is part of: Scientific Reports, 2017, vol. 7, p. 17515
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ISSN: 2045-2322
Appears in Collections:Articles publicats en revistes (Ciències Fisiològiques)
Articles publicats en revistes (Ciències Clíniques)
Articles publicats en revistes (Institut d'lnvestigació Biomèdica de Bellvitge (IDIBELL))
Articles publicats en revistes (Centres Científics i Tecnològics de la Universitat de Barcelona (CCiTUB))

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