News29.05.2018Publication of the article “Purification of replicating pancreatic β-cells for gene expression studies” |
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PhD. Esther Castaño, Responsible of the Citometry and Genomics Technology of the CCiTUB at Bellvitge Campus has published the article: “Purification of replicating pancreatic β-cells for gene expression studies” in the journal Scientific Reports published on December. The services and equipment of the Citometry and Genomics Technology of the CCiTUB has been used to obtain the results. The authors of this article(Noelia Téllez, Reyes Carballar and others) used the services of the CCiTUB basically in three applications: - Identification by citometry of the beta-pancreatic replicant cells and its purification (sorting). - Analysis of the integrity of the RNA of these cells - Gene expression analysis of the cells purified by RT-PCR The summary of the article is: "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." More information about the article in question is available at the following link:[+]. |
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