1. Cryosectioning the intestinal crypt-villus axis: An ex vivo method to study the dynamics of epigenetic modifications from stem cells to differentiated cells
- Author
-
Vincent, Audrey, Kazmierczak, Catherine, Duchêne, Belinda, Jonckheere, Nicolas, Leteurtre, Emmanuelle, Van Seuningen, Isabelle, Laboratoire d'Études Rurales (LER), Université Lumière - Lyon 2 (UL2), Centre de Recherche Jean-Pierre AUBERT Neurosciences et Cancer - U837 (JPArc), Université Lille Nord de France (COMUE)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille, Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Mucines Epitheliales : du Gene a la Fonction, Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille, Droit et Santé, Université Lumière - Lyon 2 (UL2)-Isara, Centre de Recherche Jean-Pierre AUBERT Neurosciences et Cancer - U1172 Inserm - U837 (JPArc), and Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Lille Nord de France (COMUE)-Université de Lille
- Subjects
[SDV]Life Sciences [q-bio] ,Gene Expression ,[SDV.BC]Life Sciences [q-bio]/Cellular Biology ,digestive system ,Epigenesis, Genetic ,Receptors, G-Protein-Coupled ,Histones ,Mice ,Proto-Oncogene Proteins ,Basic Helix-Loop-Helix Transcription Factors ,Animals ,CDX2 Transcription Factor ,Intestinal Mucosa ,Promoter Regions, Genetic ,lcsh:QH301-705.5 ,Medicine(all) ,Homeodomain Proteins ,Polycomb Repressive Complex 1 ,Mucin-2 ,Stem Cells ,Cell Differentiation ,SOX9 Transcription Factor ,Cell Biology ,DNA Methylation ,Mice, Inbred C57BL ,lcsh:Biology (General) ,CpG Islands ,Chorionic Villi ,Developmental Biology ,Transcription Factors - Abstract
International audience; The intestinal epithelium is a particularly attractive biological adult model to study epigenetic mechanisms driving adult stem cell renewal and cell differentiation. Since epigenetic modifications are dynamic, we have developed an original ex vivo approach to study the expression and epigenetic profiles of key genes associated with either intestinal cell pluripotency or differentiation by isolating cryosections of the intestinal crypt-villus axis. Gene expression, DNA methylation and histone modifications were studied by qRT-PCR, methylation-specific PCR and micro-chromatin immunoprecipitation, respectively. Using this approach, it was possible to identify segment-specific methylation and chromatin profiles. We show that (i) expression of intestinal stem cell markers (Lgr5, Ascl2) exclusively in the crypt is associated with active histone marks, (ii) promoters of all pluripotency genes studied and transcription factors involved in intestinal cell fate (Cdx2) harbour a bivalent chromatin pattern in the crypts and (iii) expression of differentiation markers (Muc2, Sox9) along the crypt-villus axis is associated with DNA methylation. Hence, using an original model of cryosectioning along the crypt-villus axis that allows in situ detection of dynamic epigenetic modifications, we demonstrate that regulation of pluripotency and differentiation markers in healthy intestinal mucosa involves different and specific epigenetic mechanisms.
- Published
- 2015