Murine model of Hirschsprung-associated enterocolitis. I: Phenotypic characterization with development of a histopathologic grading system

Cheng, Zhi; Dhall, Deepti; Zhao, Lifu; Wang, Hanlin L.; Doherty, Terence M.; Bresee, Catherine; Frykman, Philip K.

doi:10.1016/j.jpedsurg.2009.06.009

« Previous Next »Journal of Pediatric Surgery
Volume 45, Issue 3 , Pages 475-482, March 2010

Murine model of Hirschsprung-associated enterocolitis. I: Phenotypic characterization with development of a histopathologic grading system

Zhi Cheng
- Division of Pediatric Surgery and The Pediatric Surgery Laboratory, Department of Surgery, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA
- Authors contributed equally to this work.
Deepti Dhall
- Department of Pathology, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA
- Authors contributed equally to this work.
Lifu Zhao
- Division of Pediatric Surgery and The Pediatric Surgery Laboratory, Department of Surgery, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA
Hanlin L. Wang
- Department of Pathology, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA
Terence M. Doherty
- Department of Biomedical Sciences, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA
Catherine Bresee
- The Samuel Oschin Comprehensive Cancer Institute, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA
Philip K. Frykman
- Division of Pediatric Surgery and The Pediatric Surgery Laboratory, Department of Surgery, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA
- Corresponding author.

Received 8 April 2009; received in revised form 6 June 2009; accepted 8 June 2009.

Abstract

Purpose

The aim of the study was to characterize enterocolitis in the Ednrb-null (Ednrb−/−) mouse with aganglionosis of the colon and to develop and validate a semiquantitative histopathologic grading system to assess enterocolitis.

Methods

We isolated colon and ileal specimens of Ednrb−/− and control mice (Ednrb+/+) and performed histochemical staining (H&E) on tissue sections. After establishing inflammation grading criteria, 2 blinded pathologists independently assessed the severity and depth of inflammation of proximal colon segments on 2 separate occasions. Interclass correlations (ICCs) and coefficient of variation (CV) were calculated to determine interrater and intrarater agreement. We then prospectively applied the enterocolitis grading system to Ednrb−/− mice that became clinically ill. A cohort of Ednrb−/− mice were observed until they developed clinical illness, at which time they were euthanized and had multiple organ homogenates cultured for bacteria, and colon and small bowel were histopathologically graded for enterocolitis. Spearman's rank correlations comparing enterocolitis scores with level of bacteremia were performed.

Results

Intra- and interrater ICCs of the histologic scoring system were satisfactory (0.61 and 0.94, respectively), as were intra- and interrater CVs (18% and 9%, respectively). Of the Ednrb−/− mice, 65% developed bacteremia. Those with bacteremia had significantly higher enterocolitis scores than those without bacteremia (P < .01). Ednrb−/− mice that developed bacteremia showed a strong positive correlation between total enterocolitis scores and number of bacterial colony forming units in peritoneal lavage, liver, kidney, and aerobic spleen.

Conclusions

The Ednrb−/− mouse with aganglionosis develops enterocolitis and has features similar to Hirschsprung-associated enterocolitis in humans. Our grading system is a reliable way to assess enterocolitis. By performing microsurgical pull-through, we can now perform controlled, hypothesis-driven, mechanistic studies to evaluate etiologic factors affecting enterocolitis in the Ednrb−/− mouse.

Key words: Hirschsprung's disease, Hirschsprung-associated enterocolitis, Aganglionic megacolon, Ednrb-B-null mice, Enteric nervous system, Enteric neuropathy, Histopathologic grading system