A Clinically Relevant Murine Model of Peritoneal Fibrosis by Dialysate and Catheters

Summary

Patients with severe peritoneal fibrosis have high morbidity and mortality. The mechanism of peritoneal fibrosis is unclear. In this study, we describe a simple experimental murine model of peritoneal fibrosis induced by the injection of peritoneal dialysis fluid.

Abstract

Peritoneal fibrosis can occur in patients undergoing peritoneal dialysis (PD), and patients with severe peritoneal fibrosis have high morbidity and mortality. Peritonitis, high-glucose peritoneal dialysis fluid, and a long period of PD precipitate the onset of peritoneal fibrosis. An animal study of peritoneal fibrosis is needed due to the limitations of human and in vitro studies. However, most animal models do not mimic clinical conditions. To study peritoneal fibrosis, we developed a clinically relevant murine model by implanting a peritoneal catheter and injecting 2.5% high-glucose PD fluid plus 20 mM methylglyoxal (MGO) into the peritoneal cavity daily for 21 days. Implantation of the peritoneal catheter avoids peritoneal injury by needles and mimics clinical PD patients. Immunofluorescence staining showed that myofibroblasts accumulated in the fibrotic peritoneum. The experimental group had lower ultrafiltration volume and peritoneal membrane transport function (peritoneal equilibration test). In this article, we provide a detailed protocol of the model.

Introduction

Peritoneal dialysis (PD) is one kind of kidney replacement therapy received by about 11% of end-stage renal disease (ESRD) patients¹. Studies have reported that peritoneal fibrosis develops in patients receiving PD². Encapsulating peritoneal sclerosis (EPS), a severe form of peritoneal fibrosis, results in high morbidity and mortality³. The risk factors of peritoneal fibrosis include peritonitis, high-glucose peritoneal dialysis fluid, a long period of PD, and genetic factors⁴. The mechanism of peritoneal fibrosis involves complex changes in the perito....

Protocol

The experiments were approved and conducted according to the National Taiwan University College of Medicine and College of Public Health Institutional Animal Care and Use Committee (IACUC). All mice were housed under standard care.

1. Animal experiment

1. Use male and female C57BL/6 wild type (WT) mice more than 11 weeks of age.
2. Prepare the surgical materials, including a mouse 4-french silicone port, gloves, drapes, catheters, sutures, and needles.

Discussion

The peritoneum has a visceral and parietal peritoneum, covering the abdominal organs and abdominal walls. It is composed of a monolayer of mesothelial cells, a submesothelial layer, serosa, fibroblasts, lymphocytes, and secreted proteins¹⁴. Peritoneal fibrosis is the loss and denudation of mesothelial cells, submesothelial thickening, and the accumulation of many inflammatory cells, such as myofibroblasts and macrophages^4,14.

Materials

Name	Company	Catalog Number	Comments
anti-wide spectrum Cytokeratin antibody	abcam	ab9377
Beckman Coulter	Beckman Coulter	AU5800	Biochemical analyzer
DAPI, 4′,6-diamidino-2-phenylindole antibody	Sigma-Aldrich	98718-90-3
Drapes			any
FITC goat anti-rabbit	Jackson ImmunoResearch Secondary Antibody	111-095-144
Gloves			any
GraphPad Prizm	GraphPad Software	GraphPad Software 9.0
Kellys			any
Methylglyoxal (MGO)	Sigma-Aldrich
Mini-UTE Mouse Port	Access Technologies	MMP-4S 061108B	Mouse 4-French silicone port
Monoclonal anti-actin, α-smooth muscle-Cy3 antibody	Sigma-Aldrich	C6198
Needles			any
Reflex clips 7 mm			any
Suture 4-0 Nylon			any