Flexible Colonoscopy in Mice to Evaluate the Severity of Colitis and Colorectal Tumors Using a Validated Endoscopic Scoring System

Summary

Over the past few years, new generation endoscopes have emerged as important diagnostic research aids for evaluating murine colitis and colorectal tumors. We present herein a detailed protocol for endoscopic assessment of inflammation and colorectal tumors in mice, as well as a novel scoring system that uses decimal identifiers to document the endoscopic severity of colitis and colorectal tumors.

Abstract

The use of modern endoscopy for research purposes has greatly facilitated our understanding of gastrointestinal pathologies. In particular, experimental endoscopy has been highly useful for studies that require repeated assessments in a single laboratory animal, such as those evaluating mechanisms of chronic inflammatory bowel disease and the progression of colorectal cancer. However, the methods used across studies are highly variable. At least three endoscopic scoring systems have been published for murine colitis and published protocols for the assessment of colorectal tumors fail to address the presence of concomitant colonic inflammation. This study develops and validates a reproducible endoscopic scoring system that integrates evaluation of both inflammation and tumors simultaneously. This novel scoring system has three major components: 1) assessment of the extent and severity of colorectal inflammation (based on perianal findings, transparency of the wall, mucosal bleeding, and focal lesions), 2) quantitative recording of tumor lesions (grid map and bar graph), and 3) numerical sorting of clinical cases by their pathological and research relevance based on decimal units with assigned categories of observed lesions and endoscopic complications (decimal identifiers). The video and manuscript presented herein were prepared, following IACUC-approved protocols, to allow investigators to score their own experimental mice using a well-validated and highly reproducible endoscopic methodology, with the system option to differentiate distal from proximal endoscopic colitis (D-PECS).

Introduction

Murine endoscopy has been performed as a useful research tool for over a decade^1-3. To date, most studies employing murine endoscopy have used rigid endoscopes, although some have also used flexible sigmoidoscopy. Murine endoscopy provides immediate results and a more objective estimate of the extent of intestinal normalcy, severity of inflammation, and tumor progression compared to indirect measures, such as body weight loss, diarrhea, and histology (which is suboptimal when lesions are patchy), while offering insights on the overall health of the colon. Most notably, endoscopy allows for the repeated assessment of laboratory animal models over time, as op....

Protocol

1. Setting up the Endoscopy System

1. Set up the endoscopy system before anesthetizing the mice, following the manufacturer's instructions. Use a video ureteroscope designed for humans for this protocol (Figure 1). The outer diameter of the insertion tube is 9.9 Fr (3.3 mm), its distal end is 8.5 Fr (2.8 mm), and the inner diameter of the instrumentation channel is 6 Fr (1.2 mm).
2. Although the use of endoscopes have not been linked to disease transmission [9], sterilize the endoscope and lens with 70% ethanol¹⁰ or with appropriate techniques when working with spore-forming microorganisms. Prepare bacteriostatic st....

Results

The protocol and images presented in this study were prepared with an Olympus URF-V flexible endoscope¹⁴ and NBI^11,15. Video images were saved in MPEG-2 format and pictures in JPG, TIF, PNG, or DICOM format with maximum 1,280 x 1,024 pixels, transferable to standard USB flash portable memories. NBI enhances the visual contrast of hemoglobin-containing tissues. Normal light and NBI imaging are key features to illustrate the normal anatomy of the colorectal mucosa and the differences between normal an.......

Discussion

There are several considerations regarding modification and troubleshooting of the endoscopic technique in mice. Critical aspects of the procedure that need to be mastered during the use of flexible endoscopy include the adjustment of the air volume needed to prevent abdominal distension and respiratory compromise of the mouse, and minor technical aspects of view control with coordination of the angulation and torque controls. Torque control is important to minimize the risk of intestinal perforation. When handled proper.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Isoflurane, USP	Webster Veterinary
Surgical lubricant	Savage laboratories	surgilube, 0281-0205-45
Phosphate Buffered Saline	Thermo Scientific	SH30256
RNAlater	Ambion	AM7021
Methylene Blue 1% Aqueous Solution	Fisher Science Education	S96393
Flexible digital ureteroscope	Olympus America	URF-V
Video system center	Olympus America	VISERA Pro OTV-S7 Pro
Xenon light source	Olympus America	VISERA Pro CLV-S40 Pro
Video recorder	MediCapture, Inc.	MediCap USB200
Flexible biopsy cup forceps	Olympus America	FBC-3115
Anesthesia machine	Euthanex Corporation	EZ-7000 Classic System