Intrarenal Injection of Escherichia coli in a Rat Model of Pyelonephritis

Summary

This manuscript describes a rat surgical model of pyelonephritis using direct intra-renal infection by Escherichia coli into the renal pelvis. The experimental procedure can be utilized to study the pathogenesis of pyelonephritis as well as the associated inflammation and fibrosis.

Abstract

Pyelonephritis is a bacterial infection of the kidney and is most commonly caused by Escherichia coli. Recurrent infections can cause significant renal inflammation and fibrosis ultimately resulting in declining kidney function. Before improved clinical management and prevention of pyelonephritis can be instituted, a reliable animal model must be established in order to study the mechanisms of progression, recurrence, and therapeutic efficacy. The transurethral infection model closely mimics human pyelonephritis but exhibits considerable variation due to its reliance on urethral reflux to transport the bacteria to the kidney. Herein, a detailed surgical protocol for performing bacterial injections into the rat renal pelvis is provided and confirmed by non-invasive Magnetic Resonance Imaging (MRI). Using this protocol, animals receive direct exposure to a desired concentration of E. coli bacteria and can fully recover from the surgical procedure with adequate post-operative care. This facilitates subsequent longitudinal MRI assessments of the experimental animal models for comparison with saline (sham) controls. Using this direct delivery approach, the severity of infection is controllable and applicable for mechanistic studies of progression as well as development of novel treatment strategies.

Introduction

Rodent models have been used to study numerous human disease manifestations, including pyelonephritis and urinary tract infections (UTI). UTIs are a global health problem, and can impact children, men, and women of all ages.^1,2,3 The initial manifestation of UTIs includes cystitis, and if the infection ascends along the ureter, a kidney infection (pyelonephritis) may follow. At the same time, the prevalence of diabetes is approaching 400 million people worldwide.^4,5 Importantly, UTI incidence may be up to 4 times high....

Protocol

All animal studies were performed according to approved Institutional Animal Care and Use Committee (IACUC) protocols at Case Western Reserve University. The duration of the surgical procedure described below is approximately 45-60 min. The MRI procedure itself is approximately 15 min for each time point.

1. Anesthesia

1. Anesthetize rat in isoflurane chamber set at 2% isoflurane mixed with oxygen to facilitate animal handling and restraint prior to administering injectable anesthesia intra-peritoneally.
2. After 3-5 min of exposure to isoflurane, check that the animal is anesthetized and exhibits no response to toe pinch.

Results

Medical imaging techniques offer the opportunity to non-invasively assess UTI and therapeutic efficacy. Therefore, MRI was utilized to validate induction of acute infection after injection of 1-2 x 10⁷ UTI89 E. coli, and to visualize the changes in the kidney before and after surgery. Figure 1a-b shows a progressively increasing region of kidney infection (yellow arrows). MRI images obtained for each animal at days 1 and 4 post-infection help character.......

Discussion

Ascending acute pyelonephritis in rodents (i.e. mice and rats) can be produced by transurethral catheterization.^16,17,18 This transurethral infection method is advantageous in that it is non-invasive and mimics the human pathophysiology of ascending infection.^17,18,19,20 However, this method c.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Absorbing Pad	Fisher	14-127-47
Sterile Cotton Gauze Pad	Fisher	22-415-469
Latex Surgical Gloves	Henry Schein Animal Health	21540
Curved Mayo Scissors	Fisher	S17341
Straight Blunt Foreceps	Fisher	08-895
Scalpel Handle	Fisher	08-913-5
Sterile Scalpel Blades	Fisher	53220
1 ml Luer-Lok Syringe	BD Biosciences	309628	For bacterial injections
20 ml Luer-Lok Syringe	BD Biosciences	301031	For saline wash
Hemostat	Seneca Medical	240267
23 G 3/4 in. Needle	BD Biosciences	305143
30 G 1 in. Needle	BD Biosciences	305128
U-100 Insulin Syringe	Exel International	25846	For medication injections
Isoflurane	Henry Schein Animal Health	050033
Xylazine	Henry Schein Animal Health	33197	Inject IP
Ketamine	Patterson Vetrinary	07-881-9413	Inject IP
Yohimbine (Atipamezole)	Patterson Vetrinary	07-867-7097	Inject IP after surgery
Bupivacaine (Marcaine)	Patterson Vetrinary	07-890-4584	Inject SQ at site of incision
4-0 Chromic Gut Suture	Ethicon Inc.	U203H
4-0 Braided Vicryl Suture	Ethicon Inc.	J304H
1 ml SubQ Syringe	BD Biosciences	309597
E. coli  UTI89 or CFT073	ATCC	700928
Surgicel Absorbable Hemostat	Ethicon Inc.	ETH1951CS
Biospec 9.4T MRI	Bruker	94/20 USR