Isolation of mouse pancreatic alpha, beta, duct and acinar populations with cell surface markers

Oregon Health and Science University and Oregon Stem Cell Center, 3181 SW Sam Jackson Park Rd., Portland, OR 97239, USA.
Molecular and Cellular Endocrinology (Impact Factor: 4.41). 06/2011; 339(1-2):144-50. DOI: 10.1016/j.mce.2011.04.008
Source: PubMed


Tools permitting the isolation of live pancreatic cell subsets for culture and/or molecular analysis are limited. To address this, we developed a collection of monoclonal antibodies with selective surface labeling of endocrine and exocrine pancreatic cell types. Cell type labeling specificity and cell surface reactivity were validated on mouse pancreatic sections and by gene expression analysis of cells isolated using FACS. Five antibodies which marked populations of particular interest were used to isolate and study viable populations of purified pancreatic ducts, acinar cells, and subsets of acinar cells from whole pancreatic tissue or of alpha or beta cells from isolated mouse islets. Gene expression analysis showed the presence of known endocrine markers in alpha and beta cell populations and revealed that TTR and DPPIV are primarily expressed in alpha cells whereas DGKB and GPM6A have a beta cell specific expression profile.

Download full-text


Available from: Markus Grompe, Jun 15, 2015
  • Source
    • "Many other genes however have not been described before in the context of beta cells, including: NPTX2, neuronal pentraxin 2, found in neuronal cells and gliomas but also shown to be frequently downregulated in pancreatic cancers (Zhang et al. 2012); TSPAN1, tetraspanin 1, which can associate with alpha6.beta1 integrin and promote FAK phosphorylation (Huang et al. 2008) shown by us to be involved in insulin secretion (Rondas et al. 2011); GPM6A, neuronal membrane glycoprotein of unknown function but identified as a beta cell marker in sorted mouse islet cells (Dorrell et al. 2011a); BMP5, bone morphogenic protein 5, implicated in pancreas and fetal beta cell development (Jiang et al. 2002); and P2RY1, purinergic receptor through which ADP and ATP modulate insulin secretion (Fernandez-Alvarez et al. 2001). Alternative splicing (AS), a common feature of most (;94%) eukaryotic genes contributing to tissue specificity (Pan et al. 2008) is also significantly enriched in beta-cell–specific genes (N = 200 genes, 1.22-fold enrichment). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Elucidating the pathophysiology and molecular attributes of common disorders as well as developing targeted and effective treatments hinges on the study of the relevant cell type and tissues. Pancreatic beta cells within the islets of Langerhans are centrally involved in the pathogenesis of both type 1 and type 2 diabetes. Describing the differentiated state of the human beta cell has been hampered so far by technical (low resolution microarrays) and biological limitations (whole islet preparations rather than isolated beta cells). We circumvent these by deep RNA sequencing of purified beta cells from 11 individuals, presenting here the first characterization of the human beta cell transcriptome. We perform the first comparison of gene expression profiles between beta cells, whole islets and beta cell depleted islet preparations, revealing thus beta cell specific expression and splicing signatures. Further, we demonstrate that genes with consistent increased expression in beta cells have neuronal-like properties, a signal previously hypothesized. Finally, we find evidence for extensive allelic imbalance in expression and uncover genetic regulatory variants (eQTLs) active in beta cells. This first molecular blueprint of the human beta cell offers biological insight into its differentiated function including expression of key genes associated with both major types of diabetes.
    Full-text · Article · May 2013 · Genome Research
  • [Show abstract] [Hide abstract]
    ABSTRACT: The relentless nature and increasing prevalence of human pancreatic diseases, in particular, diabetes mellitus and adenocarcinoma, has motivated further understanding of pancreas organogenesis. The pancreas is a multifunctional organ whose epithelial cells govern a diversity of physiologically vital endocrine and exocrine functions. The mechanisms governing the birth, differentiation, morphogenesis, growth, maturation, and maintenance of the endocrine and exocrine components in the pancreas have been discovered recently with increasing tempo. This includes recent studies unveiling mechanisms permitting unexpected flexibility in the developmental potential of immature and mature pancreatic cell subsets, including the ability to interconvert fates. In this article, we describe how classical cell biology, genetic analysis, lineage tracing, and embryological investigations are being complemented by powerful modern methods including epigenetic analysis, time-lapse imaging, and flow cytometry-based cell purification to dissect fundamental processes of pancreas development.
    No preview · Article · May 2012 · Cold Spring Harbor perspectives in biology
  • [Show abstract] [Hide abstract]
    ABSTRACT: Pancreatic endocrine cells are produced from a dynamic epithelium in a process that, as in any developing organ, is driven by interacting programs of spatiotemporally regulated intercellular signals and autonomous gene regulatory networks. These algorithms work to push progenitors and their transitional intermediates through a series of railroad-station-like switching decisions to regulate flux along specific differentiation tracks. Extensive research on pancreas organogenesis over the last 20 years, greatly spurred by the potential to restore functional β-cell mass in diabetic patients by transplantation therapy, is advancing our knowledge of how endocrine lineage bias is established and allocation is promoted. The field is working towards the goal of generating a detailed blueprint of how heterogeneous cell populations interact and respond to each other, and other influences such as the extracellular matrix, to move into progressively refined and mature cell states. Here, we highlight how signaling codes and transcriptional networks might determine endocrine lineage within a complex and dynamic architecture, based largely on studies in the mouse. The process begins with the designation of multipotent progenitor cells (MPC) to pancreatic buds that subsequently move through a newly proposed period involving epithelial plexus formation-remodeling, and ends with formation of clustered endocrine islets connected to the vascular and peripheral nervous systems. Developing this knowledge base, and increasing the emphasis on direct comparisons between mouse and human, will yield a more complete and focused picture of pancreas development, and thereby inform β-cell-directed differentiation from human embryonic stem or induced pluripotent stem cells (hESC, iPSC). Additionally, a deeper understanding may provide surprising therapeutic angles by defining conditions that allow the controllable reprogramming of endodermal or pancreatic cell populations.
    No preview · Article · Jun 2012 · Seminars in Cell and Developmental Biology
Show more