Epigenetic Analysis of Endocrine Cell Subtypes From Human Pancreatic Islets

Methods Mol Biol. 2017;1507:95-111. doi: 10.1007/978-1-4939-6518-2_8.

Abstract

Epigenetic mechanisms have been proposed to contribute to multiple complex diseases, including diabetes mellitus. Dissecting the epigenomic landscape of normal and disease states has greatly expanded our understanding of the regulation of key players in disease pathogenesis and is thus opening doors to novel therapeutic avenues. The human pancreatic islet is a pivotal micro-organ that maintains global glucose homeostasis. The heterogeneity of the islet impedes meaningful in-depth epigenetic analysis using whole islet tissue, because important marks in one cell type are masked by signals from other cell types. We describe here a detailed protocol for the isolation of highly purified endocrine cell subtypes (beta, alpha, and delta cells) from human cadaveric islets, to perform cell-type-specific epigenomic analysis of histone modifications as well as global gene expression profiling.

Keywords: Alpha cell; Beta cell; ChIP-seq; Delta cell; Epigenetics; FACS; Human islets; RNA extraction.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, N.I.H., Extramural

MeSH terms

  • Biomarkers / metabolism
  • Cell Separation
  • Cells, Cultured
  • Chromatin / isolation & purification
  • Chromatin Immunoprecipitation
  • DNA / isolation & purification
  • Enteroendocrine Cells / metabolism
  • Epigenesis, Genetic*
  • Flow Cytometry
  • Gene Expression Profiling*
  • Histones / isolation & purification
  • Histones / metabolism
  • Humans
  • Islets of Langerhans / cytology
  • Islets of Langerhans / metabolism*
  • Protein Processing, Post-Translational
  • RNA / genetics
  • RNA / isolation & purification
  • RNA / metabolism
  • Real-Time Polymerase Chain Reaction
  • Sequence Analysis, RNA

Substances

  • Biomarkers
  • Chromatin
  • Histones
  • RNA
  • DNA