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©The Author(s) 2016. Published by Baishideng Publishing Group Inc. All rights reserved.
Characterisation of colonic dysplasia-like epithelial atypia in murine colitis
Sarron Randall-Demllo, Ruchira Fernando, Terry Brain, Sukhwinder Singh Sohal, Anthony L Cook, Nuri Guven, Dale Kunde, Kevin Spring, Rajaraman Eri
Sarron Randall-Demllo, Sukhwinder Singh Sohal, Dale Kunde, Rajaraman Eri, School of Health Sciences, University of Tasmania, Launceston, Launceston TAS 7250, Australia
Ruchira Fernando, Terry Brain, Department of Pathology, Launceston General Hospital, Launceston, Launceston TAS 7250, Australia
Anthony L Cook, Wicking Dementia Research and Education Centre, University of Tasmania, Hobart TAS 7005, Australia
Nuri Guven, Division of Pharmacy, School of Medicine, University of Tasmania, Hobart TAS 7005, Australia
Sukhwinder Singh Sohal, Breathe Well Centre of Research Excellence for Chronic Respiratory Disease and Lung Ageing, School of Medicine, University of Tasmania, Hobart TAS 7005, Australia
Kevin Spring, Medical Oncology, Ingham Institute for Applied Medical Research, Liverpool, Liverpool NSW 2170, Australia
Kevin Spring, Liverpool Clinical School, Western Sydney University, Richmond NSW 2753, Australia
Kevin Spring, South West Sydney Clinical School, University of New South Wales, Sydney NSW 2052, Australia
Author contributions: Randall-Demllo S and Eri R conceived and designed the experiments; Randall-Demllo S performed experiments and collected data; Randall-Demllo S, Fernando R and Brain T analysed the data; Fernando R, Brain T, Sohal SS, Cook AL, Guven N, Kunde D and Eri R contributed reagents and materials; Randall-Demllo S, Fernando R, Brain T, Sohal SS, Cook AL, Guven N, Kunde D, Spring K and Eri R contributed to writing the manuscript.
Supported by a Clifford Craig Medical Research Trust project grant and Cancer Council Tasmania (to Kunde D and Eri R); a Bowel Cancer Funding Partners PhD scholarship generously funded by Rotary District 9830, Australian Rotary Health and the University of Tasmania (to Randall-Demllo S).
Institutional review board statement: The study was reviewed and approved by the University of Tasmania Institutional Review Board.
Institutional animal care and use committee statement: All procedures involving animals were reviewed and approved by the Institutional Animal Care and Use Committee of the University of Tasmania animal Ethics Committee. Ethics Committee Number: A13329.
Conflict-of-interest statement: No conflict of interest exists.
Data sharing statement: All data relevant to this study are included in the paper and its supporting information.
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
Correspondence to: Rajaraman Eri, DVM, PhD, School of Health Sciences, University of Tasmania, Launceston, Churchill Avenue, Launceston TAS 7250, Australia.
rderi@utas.edu.au
Telephone: +61-3-62262999
Received: June 27, 2016
Peer-review started: June 28, 2016
First decision: July 29, 2016
Revised: August 15, 2016
Accepted: September 6, 2016
Article in press: September 6, 2016
Published online: October 7, 2016
Processing time: 95 Days and 3.3 Hours
AIM
To determine if exacerbation of pre-existing chronic colitis in Winnie (Muc2 mutant) mice induces colonic dysplasia.
METHODS
Winnie mice and C57BL6 as a genotype control, were administered 1% w/v dextran sulphate sodium (DSS) orally, followed by drinking water alone in week-long cycles for a total of three cycles. After the third cycle, mice were killed and colonic tissue collected for histological and immunohistochemical evaluation. Inflammation and severity of dysplasia in the colonic mucosa were assessed in H&E sections of the colon. Epithelial cell proliferation was assessed using Ki67 and aberrant β-catenin signalling assessed with enzyme-based immunohistochemistry. Extracted RNA from colonic segments was used for the analysis of gene expression using real-time quantitative PCR. Finally, the distribution of Cxcl5 was visualised using immunohistochemistry.
RESULTS
Compared to controls, Winnie mice exposed to three cycles of DSS displayed inflammation mostly confined to the distal-mid colon with extensive mucosal hyperplasia and regenerative atypia resembling epithelial dysplasia. Dysplasia-like changes were observed in 100% of Winnie mice exposed to DSS, with 55% of these animals displaying changes similar to high-grade dysplasia, whereas high-grade changes were absent in wild-type mice. Occasional penetration of the muscularis mucosae by atypical crypts was observed in 27% of Winnie mice after DSS. Atypical crypts however displayed no evidence of oncogenic nuclear β-catenin accumulation, regardless of histological severity. Expression of Cav1, Trp53 was differentially regulated in the distal colon of Winnie relative to wild-type mice. Expression of Myc and Ccl5 was increased by DSS treatment in Winnie only. Furthermore, increased Ccl5 expression correlated with increased complexity in abnormal crypts. While no overall difference in Cxcl5 mucosal expression was observed between treatment groups, epithelial Cxcl5 protein appeared to be diminished in the atypical epithelium.
CONCLUSION
Alterations to the expression of Cav1, Ccl5, Myc and Trp53 in the chronically inflamed Winnie colon may influence the transition to dysplasia.
Core tip: Patients with ulcerative colitis (UC) are at increased risk of developing colonic cancer. Understanding progression to early dysplastic change in the UC-associated inflammation in the colon required a suitable animal model. Winnie mice develop a UC-like chronic colitis and endoplasmic reticulum stress due to a Muc2 mutation encoding a misfolded mucin-2. We hypothesised that exacerbation of pre-existing chronic inflammation using colitogenic dextran sulphate sodium in a model of spontaneous colitis would induce colorectal tumourigenesis. This study demonstrated that exacerbation of colitis resulted in epithelial hyperplasia in the distal colon and crypt abnormalities resembling dysplasia. Altered expression of genes known to modify tumour growth, specifically Cav1, Ccl5, Myc and Trp53, in Muc2 mutants may predispose to early neoplastic change in the inflamed colon.
