Un modèle murin d'irréversible et réversible unilatérale obstruction urétérale

Summary

The murine model of irreversible unilateral ureteric obstruction (UUO) is presented together with the model of reversible UUO in which the ureteric obstruction is relieved by anastomosis of the severed ureter into the bladder. These models enable the study of renal inflammation and scarring as well as tissue remodeling.

Abstract

Obstruction of the kidney may affect native or transplanted kidneys and results in kidney injury and scarring. Presented here is a model of obstructive nephropathy induced by unilateral ureteric obstruction (UUO), which can either be irreversible (UUO) or reversible (R-UUO). In the irreversible UUO model, the ureter may be obstructed for variable periods of time in order to induce increasingly severe renal inflammation and interstitial fibrotic scarring. In the reversible R-UUO model the ureter is obstructed to induce hydronephrosis, tubular dilation and inflammation. After a suitable period of time the ureteric obstruction is then surgically reversed by anastomosis of the severed previously obstructed ureter to the bladder in order to allow complete decompression of the kidney and restoration of urinary flow to the bladder. The irreversible UUO model has been used to investigate various aspects of renal inflammation and scarring including the pathogenesis of disease and the testing of potential anti-inflammatory or anti-fibrotic therapies. The more challenging model of R-UUO has been used by some investigators and does offer significant research potential as it allows the study of inflammatory and immune processes and tissue remodeling in an injured and scarred kidney following the removal of the injurious stimulus. As a result, the R-UUO model offers investigators the opportunity to explore the resolution of kidney inflammation together with key aspects of tissue repair. These experimental models are of relevance to human disease as patients often present with obstruction of the renal tract that requires decompression and are commonly left with significant residual kidney impairment that has no current treatment options and may lead to eventual end stage kidney failure.

Introduction

The overall aim of the experimental model described here is to induce obstructive nephropathy by unilateral ureteric obstruction (UUO), which can either be irreversible (UUO) or reversible (R-UUO). A simple irreversible model of UUO is presented in which the left ureter is permanently obstructed by ligation with a suture or by the application of a ligating clip. This results in marked dilatation of the ureter together with reduced renal blood flow and glomerular filtration. Renal histology demonstrates tubular dilatation and increasingly severe interstitial renal inflammation and fibrosis. Irreversible UUO is a useful model and has been adopted by many researchers in the study of both renal inflammation and fibrosis.^1-4 Although the irreversible model of UUO requires some surgical expertise it is relatively straightforward and is often used to seek insights into the pathogenesis of interstitial renal injury and the ensuing fibrosis. Also presented is a less frequently used R-UUO model using a modification of the method originally described by Tapmeier et al.⁵ The R-UUO model has much future potential for the study of inflammatory and immune processes, cellular and tissue regeneration as well as the subsequent tissue remodeling following the removal of an injurious stimulus.

The more challenging R-UUO model has been used by a limited number of investigators with some groups employing a significantly different surgical technique to that described here^6,7 though with interesting results. In the R-UUO model presented, the ureter is ligated to induce complete ureteric obstruction for a period of time sufficient to induce the level of injury and fibrosis desired: 7 days of UUO was chosen in the method described here. The ureteric obstruction is reversed and the kidney is allowed to decompress for a period of time determined by the investigator before the mice are culled and the kidney removed for analysis: 7 days of decompression was chosen in the method described here though a longer period would be chosen if the resolution of inflammation and fibrosis was being studied. Although the method described here requires significant surgical expertise, it offers several advantages over other R-UUO models. The application of soft walled plastic tubing to the obstructed ureter prevents excessive ureteric dilation and this facilitates the subsequent manipulation and anastomosis of the ureter. Furthermore, in the described R-UUO model the ureter is divided thereby allowing the removal of any residual urinary sediment and debris. This confirms that the remaining ureter lumen is de-obstructed and patent prior to anastomosis to the bladder.

Experiments incorporating both the irreversible and reversible UUO models can provide researchers with a powerful insight into the molecular and cellular mechanisms of both injury and subsequent resolution and regeneration. Thus, the model of R-UUO described here would be highly relevant to researchers interested in post-inflammatory tissue remodeling and how this can be modulated.

Protocol

Directives générales: expériences sur les animaux ont été effectuées en conformité avec les directives et règlements imposés par les animaux (Scientific Procedures) de Loi 1986. Ce protocole et le protocole de vidéo qui l'accompagne sont à la fois un UUO standard et un R-UUO, qui peut être effectuée sur plusieurs souches de souris. Dans la vidéo d'accompagnement, les deux procédures sont effectuées sur des souris C57BL / 6 mâles âgées de 8 semaines les souris. Les données présentées dans la…

Representative Results

L'apparition des modifications rénales nettement suivante obstruction urétérale et il devient plus pâle et tendu à la palpation avec le temps (Figure 1). Il est de plus en plus la dilatation de l'urètre proximal à l'obstruction et le bassinet du rein. Le rein devient de plus en plus atrophique que la durée de néphropathie obstructive augmente avec obstruction prolongée entraîne un amincissement notable de la moelle et le cortex rénal. Après le renversement de l'obstruction d…

Discussion

Obstruction of the kidney may affect native or transplanted kidneys and results in kidney injury and scarring. The R-UUO model is of relevance to human disease as patients often present with obstruction of the renal tract secondary to prostatic hypertrophy, posterior urethral valves etc. that requires decompression with patients commonly left with significant residual kidney impairment that has no current treatment options and may eventually lead to end stage kidney disease.^8,9 The models detailed here enable …

Materials

Blunt Dissecting Scissors	Fine Science Tools	14072-10
Spring Scissors – straight	Fine Science Tools	15000-10
Toothed forceps	Fine Science Tools	11021-12
Angled forceps x 2	Fine Science Tools	00649-11
Straight forceps	Fine Science Tools	00632-11
Colibri 3cm wire retractor	Fine Science Tools	17000-03
Castroviejo needle holder with lock	Fine Science Tools	12565-14
Wound clip applicator	Fine Science Tools	12031-07
7mm wound clips	Fine Science Tools	12032-07
Castroviejo micro needle holder with lock	Fine Science Tools	12060-01
OPMI pico microscope	Carl Zeiss	S100
Heat electronic pad	Cozee Comfort	n/a
6/O silk braided suture	Harvard Apparatus	72-3287
9/O Dafilon (polyamide) suture	B-Braun	G1111434
5/O braided silk suture	Harvard Apparatus	51-7680
Regular bevel needle, 1 inch, 21G	Becton, Dickinson and Company	305175
1 ml syringe slip tip	Becton, Dickinson and Company	300184
Wypall paper swabs	Kimberley-Clark	L40	Sterilised (Autoclave)
Cotton wool buds	Johnson and Johnson	n/a	Sterilised (Autoclave)
Plain drapes	Guardian	CB03	Sterilised (Autoclave)
Soft wall silicone rubber tubing			Silicone tubing – internal diameter 1.0 mm; external diameter 2.0 mm; wall thickness 0.6 mm
(Lacri-Lube) White soft paraffin 57.3%, mineral oil 42.5% and lanolin alcohols 0.2%	Allergan Ltd	21956GB10X
(Videne) Povidone-iodine 10%	Ecolab Ltd	PL 04509/0041
(Vetalar V) Ketamine hydrochloride	Pfizer Animal Health	Vm 42058/4165	100mg/ml solution
(Domitor) Medetomidine hydrochloride	Orion Pharma	Vm 06043/4003	1mg/ml
(Vetergesic) Bupernorphine hydrochloride	Alsto Animal Health	Vm 00063/4002	0.3mg/ml
(Antisedan) Atipamezole hydrochoride	Orion Pharma	Vm 06043/4004	5mg/ml
(Adept) 4% Icodextrin	Baxter		Adhesion reduction solution
NaCl 0.9%	Baxter	FKE1323