title: Cryosectioning the intestinal crypt-villus axis: an ex vivo method to study the dynamics of epigenetic modifications from stem cells to differentiated cells.
abstract: 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.
authors.affiliation: Inserm, UMR837, Jean Pierre Aubert Research Center (JPARC), Team 5 "Mucins, Epithelial Differentiation and Carcinogenesis," rue Polonovski, Lille, France; Université Lille 2 Droit et Santé, Lille, France; Centre Hospitalier Régional et Universitaire de Lille, Lille, France. Electronic address: audrey.vincent@inserm.fr.
authors.forename: Audrey
authors.initials: A
authors.lastname: Vincent
authors.affiliation: Inserm, UMR837, Jean Pierre Aubert Research Center (JPARC), Team 5 "Mucins, Epithelial Differentiation and Carcinogenesis," rue Polonovski, Lille, France; Université Lille 2 Droit et Santé, Lille, France; Centre Hospitalier Régional et Universitaire de Lille, Lille, France; Centre de Biologie Pathologie, CHRU Lille, Lille, France.
authors.forename: Catherine
authors.initials: C
authors.lastname: Kazmierczak
authors.affiliation: Inserm, UMR837, Jean Pierre Aubert Research Center (JPARC), Team 5 "Mucins, Epithelial Differentiation and Carcinogenesis," rue Polonovski, Lille, France; Université Lille 2 Droit et Santé, Lille, France; Centre Hospitalier Régional et Universitaire de Lille, Lille, France.
authors.forename: Belinda
authors.initials: B
authors.lastname: Duchêne
authors.affiliation: Inserm, UMR837, Jean Pierre Aubert Research Center (JPARC), Team 5 "Mucins, Epithelial Differentiation and Carcinogenesis," rue Polonovski, Lille, France; Université Lille 2 Droit et Santé, Lille, France; Centre Hospitalier Régional et Universitaire de Lille, Lille, France.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Jonckheere
authors.affiliation: Inserm, UMR837, Jean Pierre Aubert Research Center (JPARC), Team 5 "Mucins, Epithelial Differentiation and Carcinogenesis," rue Polonovski, Lille, France; Université Lille 2 Droit et Santé, Lille, France; Centre Hospitalier Régional et Universitaire de Lille, Lille, France; Centre de Biologie Pathologie, CHRU Lille, Lille, France.
authors.forename: Emmanuelle
authors.initials: E
authors.lastname: Leteurtre
authors.affiliation: Inserm, UMR837, Jean Pierre Aubert Research Center (JPARC), Team 5 "Mucins, Epithelial Differentiation and Carcinogenesis," rue Polonovski, Lille, France; Université Lille 2 Droit et Santé, Lille, France; Centre Hospitalier Régional et Universitaire de Lille, Lille, France.
authors.forename: Isabelle
authors.initials: I
authors.lastname: Van Seuningen
mesh_terms: D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics / Q000378:metabolism; D000071616:CDX2 Transcription Factor; D002454:Cell Differentiation; D002824:Chorionic Villi / Q000378:metabolism* / Q000473:pathology; D018899:CpG Islands; D019175:DNA Methylation; D044127:Epigenesis, Genetic; D015870:Gene Expression; D006657:Histones / Q000378:metabolism; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D007413:Intestinal Mucosa / Q000166:cytology / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D055262:Mucin-2 / Q000235:genetics / Q000378:metabolism; D063150:Polycomb Repressive Complex 1 / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism; D043562:Receptors, G-Protein-Coupled / Q000235:genetics / Q000378:metabolism; D055758:SOX9 Transcription Factor / Q000235:genetics; D013234:Stem Cells / Q000166:cytology* / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
chemical_list: D051792:Basic Helix-Loop-Helix Proteins; D000071616:CDX2 Transcription Factor; D006657:Histones; D018398:Homeodomain Proteins; D055262:Mucin-2; D063150:Polycomb Repressive Complex 1; D011518:Proto-Oncogene Proteins; D043562:Receptors, G-Protein-Coupled; D055758:SOX9 Transcription Factor; D014157:Transcription Factors; C489311:Ascl2 protein, mouse; C489421:Bmi1 protein, mouse; C487514:Cdx2 protein, mouse; C493597:Lgr5 protein, mouse; C524626:Muc2 protein, mouse