A Mouse 5/6th Nephrectomy Model That Induces Experimental Uremic Cardiomyopathy

This manuscript provides a detailed two-step surgical procedure to perform mouse 5/6^th 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.

Transcript

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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Journal (Biology)

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A Mouse 5/6th Nephrectomy Model That Induces Experimental Uremic Cardiomyopathy

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A Mouse 5/6^th Nephrectomy Model That Induces Experimental Uremic Cardiomyopathy