Murine Appendectomy Model of Chronic Colitis Associated Colorectal Cancer by Precise Localization of Caecal Patch

Summary

The presented protocol describes a facile surgical removal of the appendix (caecal patch) in a mouse followed by the induction of inflammatory bowel disease-associated colorectal cancer. This murine appendectomy model enables investigation of the biological role of the appendix in the pathogenesis of human gastrointestinal disease.

Abstract

The human appendix has been recently implicated to play important biological roles in the pathogenesis of various complex diseases, such as colorectal cancer, inflammatory bowel disease, and Parkinson’s disease. To study the function of the appendix, a gut disease-associated murine appendectomy model has been established and its step-by-step protocol is described here. This report introduces a facile protocol for caecal patch removal in mice followed by the chemical induction of chronic colitis-associated colorectal cancer using a combination of dextran sulfate sodium (DSS) and azoxymethane (AOM). IgA specific cells and IgA concentration were significantly reduced upon removal of the caecal patch in male C57BL/6 mice compared to those in the sham group. Simultaneously administering 2% DSS and AOM resulted in nearly 80% mice survival in both sham and appendectomy groups without significant body weight loss. Histological results confirmed colonic inflammation and different degrees of adenocarcinoma. This model can be used for the study of the functional role of the appendix in maintaining gut microbiota homeostasis and pathogenesis of gut colitis and malignancies, as well as for the potential development of drug targeting therapies.

Introduction

The clinical appendectomy is a standard surgical procedure involving removal of the appendix mostly due to inflammation (e.g., appendicitis)^1,2,3. However, the biological function of the vermiform human appendix remains controversial^4,5,6. The appendix has been regarded as a vestigial remnant projecting from the cecum in the large bowel. Until recently, evolutionary, immunological, morphological, and microbiological studies have suggested that the appendix may possess distinct functi....

Protocol

All animal procedures were approved by the Institutional Animal Care and Use Committee of Xi’an Jiaotong University (No. XJTULAC2019-1023).

1. Mice appendectomy

1. House 8–10-week-old C57BL/6 male mice in a certified specific-pathogen free (SPF) environment for 1 week prior to surgery.
2. Prepare the following sterile surgical instruments: one pair of micro-scissors, one pair of micro-forceps, two sizes (4-0 and 8-0) of sterilized non-absorbable sutures, an electric coagulation pen with needle holder, 75% medical alcohol, iodine-based scrub (e.g., entoiodine), and a package of sterile cotton swabs.
3. Fill....

Results

Establishment of murine appendectomy model

This murine appendectomy model of chronic colitis associated colorectal cancer can be generated by following the sequential surgical and induction steps as illustrated in Figure 1. The most frequent positions of caecum are in the left and right iliac fossa followed by the middle line of the abdomen (Figure 2). The successful rate of pre-localization of caecum prior to the abd.......

Discussion

A murine appendectomy model of colitis-associated colorectal cancer was obtained using surgical steps with a high survival rate in mice. In most cases, since the caecum was positioned under the abdominal wall (Supplementary Table 1, Supplementary Table 2, and Figure 2), it was difficult to prejudge its location without laparotomy. In this surgical protocol, an easy step of touching the bump was introduced, and quantitative evaluation of the cecum location was also provided a.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Azoxymethane(AOM)	Sigma-Aldrich,Inc.	A5486
Dextran Sulfate Sodium Salt(DSS)	MP Biomedicals,Inc.	160110
Entoiodine	Shanghai likon high technology disinfection co. LTD	310102
digital caliper	Ningbo yuanneng trading co. LTD	4859263
4-0 Silk Sutures	Yuanlikang co. LTD	20172650032
8-0 Prolene Sutures	Yuanlikang co. LTD	20172650032
Electric coagulation pen	Chuang mei medical equipment co. LTD	28221777292
disposable syringe 1ml	Shengguang medical products co. LTD	3262-2014
disposable syringe 10ml	Shengguang medical products co. LTD	3262-2014
75% Medicinal alcohol	Shandong anjie high-tech disinfection technology co. LTD	371402AAJ008
Pentobarbital sodium salt	Sigma-Aldrich,Inc.	57-33-0
Physiological Saline	Shandong qidu pharmaceutical co. LTD	H37020766
Absorbent Cotton Swab	Henan ruike medical co., LTD	RK051
Surgical Instruments-Ophthalmic	Jinzhong Shanghai co.LTD	WA3050