A Mouse 5/6th Nephrectomy Model That Induces Experimental Uremic Cardiomyopathy
November 7th, 2017
This manuscript provides a detailed two-step surgical procedure to perform mouse 5/6th partial nephrectomy (PNx) with pole ligation. Four weeks after surgery, in comparison with sham-operated mice, the PNx mice developed impaired renal function, anemia, cardiac hypertrophy, cardiac fibrosis, and decreased heart systolic and diastolic function.
The overall goal of this procedure is to set up a simple mouse five-sixth partial nephrectomy model with pole ligation with a better survival rate than traditional models. This method can help answer key questions in the chronic kidney disease field about the development and potential therapeutic treatment of uremic cardiomyopathy. The main advantage of this technique is that it is an easy and consistently reproducible animal model that mimics the pathology of chronic kidney disease in human patients.
Generally, individuals new to this method will struggle because of the much smaller kidneys, blood vessels, and ureters in the mouse. Begin by confirming the appropriate level of sedation by a lack of response to toe pinch and applying lubricant to the mouse's eyes. Place the mouse on a warming pad and insert a rectal thermometer to maintain the body temperature around 37 degrees Celsius during the procedure.
Then, place the animal onto its right side and shave the hair over the surgical area. Remove any remaining hair with depillatory cream and disinfect the exposed skin with polyhydroxy dine solution and 70%ethanol. Align an autoclave surgical linen over the surgery site.
And make an approximately one-centimeter lumbar incision through the skin and muscle tissue. Using the scalpel and forceps, separate the muscle and fascia to expose the left kidney. Then, use small blunt forceps to gently pull on the perirenal fat to extract the kidney from the abdominal cavity.
Gently tease the connective tissue and adrenal gland from the superior end of the kidney towards the middle pylorus of the organ where the blood vessels enter and exit the renal tissue. Isolate the ureter around the inferior half of the kidney within the connective tissue, and gently separate the connective tissue from the kidney to ensure that the ureter is not ligated. It is essential to separate the connective tissue carefully so that the ureter is not damaged and does not bleed.
Isolate the ureter around the inferior half of the kidney within the connective tissue. Then, use three zero silk suture strings to ligate the kidney approximately 0.4 centimeters from the superior portion, toward the middle of the pylorus and 0.4 centimeters from the inferior portion towards the pylorus. When tightening the sutures, apply just enough tension to compromise the blood supply to the distal portion of the kidney without causing bleeding on either side.
If no bleeding is observed after two minutes, gently return the kidney to its original anatomic location, and use four zero and three zero sutures to close the muscle and skin layers respectively. Then, apply antibiotic ointment to the surgical area and house the mouse individually with a heat lamp and monitoring until sternal recumbency. Seven days after the polar ligation, expose the right kidney as just demonstrated.
Use blunt forceps to clear the surrounding fat and connective tissue, and loosely place two three zero silk sutures around the renal artery and vein. Gently tease the connective tissue and adrenal gland from the superior end of the kidney towards the middle pylorus. Gently displacing the ureter from the inferior end of the tissue, untie all of the vessels together without causing bleeding.
Next, move one loose ligature knot toward the abdominal aorta side, and the second loose ligature toward the kidney side. And tie off the abdominal aorta ligature with double knots. A solid knot will change the color of the right kidney.
Tie off the second ligature knot with double knots, leaving spaces between the knots so the vessels can be cut, and cut the renal vessels. After removing the kidney, close the muscle and skin layers as just demonstrated. Then, dry the kidney with sterile gauze and weigh the harvested renal tissue.
At four weeks post-surgery, the partial nephrectomy mice demonstrate significantly increased levels of cyctatin C, creatinine, and blood urea nitrogen in the plasma compared to sham surgery animals. Partial nephrectomy stimulated anemia is also apparent, as evidenced by the significantly lower hematocrit levels in the polar ligated animals. Cardiac hypertrophy and diastolic dysfunction in the partially nephrectomized mice are indicated by an increased heart-to-body-weight ratio and by transthoracic echocardiography analysis which reveals significant increases in the posterior and anterior wall thicknesses, the myocardial performance index, and the left ventricle mass index.
Partial nephrectomy mediated cardiac fibrosis is also demonstrated by increases in type 1 collagen expression in left ventricle homogenates and left remnant kidney homogenates. Further, oxidant stress places an important role in the experimental uremic cardiomyopathy, significantly stimulating direct protein carbonylation modification of multiple proteins and proto-oncogene tyrosine-protein kinase activation in the left ventricle homogenates. Once mastered, this technique can be completed in 40 minutes if it is performed properly.
While attempting this procedure, it's important to remember to check the respiration rate of the animal and to adjust the isofluranes as appropriate. Following this procedure, other experiments like comparing the effects of specific drug treatments on partial nephrectomy recovery can be performed to answer traditional questions about specific therapeutic treatments of interest. After its development, this technique paved the way for researchers in the field of chronic kidney disease to explore the pathology of and potential drug screening in uremic cardiomyopathy.
After watching this video, you should have a good understanding of how to set up a partial nephrectomy model in the mouse.
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Overview
This manuscript describes a two-step surgical procedure for performing mouse 5/6th partial nephrectomy (PNx) with pole ligation. This model aims to improve survival rates compared to traditional methods and helps investigate chronic kidney disease and uremic cardiomyopathy.
Key Study Components
Area of Science
- Nephrology
- Cardiology
- Surgical Techniques
Background
- Chronic kidney disease is a significant health issue.
- Uremic cardiomyopathy is a common complication of kidney disease.
- Existing animal models have limitations in survival rates.
- This study aims to establish a more effective model for research.
Purpose of Study
- To develop a reliable mouse model for 5/6th PNx.
- To investigate the effects of chronic kidney disease on cardiac function.
- To provide insights into potential therapeutic approaches.
Methods Used
- Mouse anesthesia and sedation confirmation.
- Application of lubricant to protect the eyes.
- Use of a warming pad to maintain body temperature.
- Monitoring of vital signs during the surgical procedure.
Main Results
- PNx mice exhibited impaired renal function compared to sham-operated mice.
- Development of anemia and cardiac hypertrophy was observed.
- Cardiac fibrosis and decreased heart function were noted.
- The model effectively mimics human chronic kidney disease pathology.
Conclusions
- The two-step PNx model is reproducible and effective.
- This model can facilitate research into uremic cardiomyopathy.
- Improved survival rates enhance its utility for future studies.
Frequently Asked Questions
Executive Industry Relevance
This model provides a reproducible system for studying the progression from chronic kidney disease to uremic cardiomyopathy, a key comorbidity in drug safety and efficacy assessments. It enables mechanistic de-risking of cardiovascular liability in renal impairment populations, supporting early target validation and predictive confidence in lead optimization. The model's utility in quantifying cardiac dysfunction and fibrosis aligns with preclinical screening needs for cardioprotective candidates in CKD populations.
Strategic Applications in Biopharma R&D
Early Discovery & Target Validation
- Scientific Value: Enables interrogation of therapeutic hypotheses linking renal dysfunction to cardiac pathology through measurable endpoints like cystatin C, creatinine, and BUN.
- Scientific Value: Supports functional target validation by demonstrating cardiac hypertrophy, fibrosis, and diastolic dysfunction in response to reduced renal mass.
- Scientific Value: Facilitates pathway clarification via observed oxidant stress and protein carbonylation modifications in left ventricle homogenates.
Screening & Assay Development
- Scientific Value: Provides a standardized biological system with quantifiable renal and cardiac readouts for assay standardization and reproducibility testing.
- Operational Value: Enables reliable compound evaluation through consistent phenotyping of anemia, cardiac hypertrophy, and fibrosis at four weeks post-surgery.
- Operational Value: Supports platform reuse due to the model's simplicity and improved survival rate compared to traditional methods.
Translational & Preclinical Research
- Scientific Value: Demonstrates disease relevance by mimicking human uremic cardiomyopathy pathology, including cardiac fibrosis and systolic/diastolic dysfunction.
- Scientific Value: Supports translational biomarker alignment through measurable shifts in cystatin C, creatinine, BUN, hematocrit, and heart-to-body-weight ratio.
- Scientific Value: Enables risk-adjusted advancement decisions by quantifying cardiac functional decline via echocardiography-derived indices like myocardial performance and left ventricle mass index.
Pipeline & Workflow Integration
The model fits within the discovery continuum from target validation through lead identification to preclinical safety assessment, particularly for compounds where renal impairment is a comorbidity concern.
- Discovery Biology: Supports hypothesis testing of cardiorenal axis mechanisms through inducible uremic cardiomyopathy phenotypes.
- Screening: Delivers quantitative outputs such as plasma biomarker levels and echocardiographic parameters for compound effect comparison.
- Analytics: Enables statistical analysis of cardiac structural and functional changes, including wall thickness ratios and diastolic function indices.
- Translational Research: Connects discovery findings to preclinical validation through shared pathological features like collagen upregulation and oxidant stress.
- Enterprise Reuse: Represents a scalable, reproducible surgical model suitable for multi-program screening campaigns in cardiorenal drug discovery.
Operational & Enterprise Impact
- Scientific Value: Increases predictive confidence by reducing mechanistic ambiguity in cardiorenal pathway modulation.
- Operational Value: Enhances standardization and reproducibility through a streamlined two-step surgical procedure with defined postoperative timelines.
- Strategic Value: Improves go/no-go decisions by enabling early detection of cardiovascular liability in CKD-relevant models.
- Portfolio Impact: Informs risk-adjusted prioritization by identifying compounds that mitigate or exacerbate uremic cardiomyopathy phenotypes.
Implementation Considerations
- Requires expertise in microsurgical techniques and rodent anesthesia management.
- Dependent on sterile surgical equipment, suture materials, and postoperative monitoring infrastructure.
- Necessitates standardization across teams for consistent pole ligation and nephrectomy timing.
- Involves adaptation considerations when translating to other strain or age models due to variability in renal reserve.
- Limited by the inability to precisely titrate remnant renal mass compared to artery ligation models, affecting dose-response control.
