A Modified Two Kidney One Clip Mouse Model of Renin Regulation in Renal Artery Stenosis

Summary

A modified 2 kidney 1 clip (2K1C) Goldblatt mouse model was developed using polyurethane tubing to initiate renal artery stenosis, inducing an increase in renin expression and kidney injury. Here, we describe a detailed procedure of preparing and placing the cuff onto the renal artery to generate a reproducible and consistent 2K1C mouse model.

Abstract

Renal artery stenosis is a common condition in patients with coronary or peripheral vascular disease where the renin angiotensin aldosterone system (RAAS) is overactivated. In this context, there is a narrowing of the renal arteries that stimulate an increase in the expression and release of renin, the rate-limiting protease in RAAS. The resulting rise in renin expression is a known driver of renovascular hypertension, frequently associated with kidney injury and end organ damage. Thus, there is a great interest in developing novel treatments for this condition. The molecular and cellular mechanism of renin control in renal artery stenosis is not fully understood and warrants further investigation. To induce renal artery stenosis in mice, a modified 2 kidney 1 clip (2K1C) Goldblatt mouse model was developed. The right kidney was stenosed in wild type mice and sham operated mice were used as control. After renal artery stenosis, we determined renin expression and kidney injury. Kidneys were harvested, and fresh cortices were used to determine protein and mRNA expression of renin. This animal model is reproducible and can be used to study pathophysiological responses, molecular and cellular pathways involved in renovascular hypertension and kidney injury.

Introduction

Renal artery stenosis (RAStenosis) is an intractable problem affecting about 6% of people over 65 and in up to 40% of people with coronary or peripheral vascular disease^1,2. Current treatments for the disease are limited; therefore, there is a critical need to develop new therapies to treat renovascular hypertension or resistant hypertension induced by RAStenosis. Renin angiotensin aldosterone system (RAAS) is the key pathway involved in the pathogenesis of RAStenosis induced hypertension or renovascular hypertension^3,4. Known therapies targeting RAAS,....

Protocol

Mice were housed and cared at the Vanderbilt University Medical Center (VUMC) Division of Animal Care following the National Institutes of Health (NIH) guidelines and the Guide for the Care and Use of Laboratory Animals, US Department of Health and Human Services. All animal procedures were approved by the VUMC Institutional Animal Care and Use Committee prior to starting the experiments.

1. Animal preparation and dissection

1. Turn on the germinator and water pump of the heating pad about 30 min before starting surgery.
2. Cut 0.5 mm length polyurethane tubing with a sharp scalpel. Remove 0.2 mm of the circumference by....

Results

Renal artery constriction increases renin expression in the stenosed kidney while repressing expression in the contralateral kidney. The two kidney one clip (2K1C) or Goldblatt model of stenosis induces increased renin expression and kidney injury. This is recognized as the best representative model of unilateral renal artery stenosis in humans.

Expression of renin and prorenin (precursor of renin) were measured using immunoblotting. The data show that renin and prorenin expression increa.......

Discussion

Renal artery stenosis is an important cause of secondary or resistant hypertension, and kidney injury^1,29. The two kidney one clip (2K1C) Goldblatt model has been employed to study RAStenosis induced renovascular hypertension^1,17,18,19. A number of previous studies using various animals models have shown that stenosis in the renal arter.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Diet Gel	Clear H2O	Diet-Gel 76A	Surgery recovery diet
EMC Heated Hard pad	Hallowell	000A2788B	Heating pads were used to keep mice warm
Ethilon Nylon Suture	Ethicon	662G	4-0 (1.5 metric), This suture was used to close the peritoneum, and skin
Ethilon Nylon Suture	Ethicon	2815 G	8-0 (0.4 metric), This suture was used to close cuff to tie and constrict the artery
Germinator 500	Braintree Scientific Inc.	GER 5287	Sterilize surgical tools between surgeries
Ketoprofen	Zoetis	Ketofen	Painkiller
Polyurethane	Braintree Scientific Inc.	MRE-025	This tube was used to initiate stenosis
Povidone-iodine antiseptic swabsticks	Medline	MDS093901	It was applied after hair removal and surgery on the skin
Reflex 7 Clip Applier	Roboz Surgical Instrument Co	204-1000	This clip applier was used to apply clip in case one or more sutures went off
Sterile towel drapes	Dynarex	4410	It was used as a bedsheet for mice during surgery
Triple antibiotic ointment	Medi-First	22312
Water pump	Stryker	T/pump Professionals	Used to warm and circulate water in the heating hard pad to keep mice warm during and post-surgery