An Immature Murine Model of Reversible Unilateral Ureteral Obstruction

This study establishes a reversible UU model in immature rats to investigate kidney injury recovery and assess its reliability and the application in studying post-obstruction renal repair. A key consideration in this approach is minimizing ureteral damage during surgery, as ureteral injury can induce inflammation, potentially leading to adhesion, closure, and recanalization failure. We will focus on the dynamic recovery of renal function and kidney injury following obstruction release, particularly in the context of immature RUU kidney.

[Narrator] To begin, use sterilized scissors to cut a silicone tube into approximately one-centimeter segments. Make a longitudinal incision along one side of the tube wall for subsequent use. Next, confirm adequate anesthesia of the rat by checking for the absence of reflex responses, such as the pedal withdrawal reflex upon toe pinch. Apply veterinary ophthalmic ointment to the eyes to prevent corneal drying. Depilate the rat's abdomen from the xiphoid process to the pubic symphysis, extending bilaterally to the midline. Then position the rat in a supine position on a heated surgical pad and secure its limbs with rubber ropes. Then, drape a sterile fenestrated sheet to maintain a sterile field. Now make a midline skin incision along the abdomen, extending from the sub-xiphoid region to just below the umbilicus to expose the kidneys and upper ureters. With a pair of surgical scissors, incise the subcutaneous tissues and fascia along the midline. Dissect the skin and underlying tissues meticulously layer by layer. Fully expose the retroperitoneal space using tissue forceps. For reversible unilateral ureteral obstruction, first retract the bowel to the right side of the abdominal cavity using a sterile swab. Cover the ureter with saline-soaked gauze to prevent desiccation. With microscopic forceps, dissect and mobilize the left ureter, freeing approximately 1.5 centimeters from the surrounding tissues. Then place a one-centimeter-long silicone tube beneath the freed ureter. Use forceps to ensure complete encasement within the tube. Use 3-0 silk thread to ligate the silicone tube and the middle portion of the ureter, and induce ureteral obstruction. Gradually pull the silicone tube along the ureter's longitudinal axis to ensure secure but non-slipping ligation. Reposition the bowel within the peritoneal cavity carefully, ensuring proper alignment without tension or obstruction. Suture the abdominal muscle and fascial layers using a 3-0 non-absorbable suture with a curved cutting needle in a continuous manner. Then, close the skin with a running 4-0 non-absorbable suture, ensuring anatomical alignment and even tension. For the relief surgery, first dissect the knot of the silicone tube carefully using a scalpel blade. Remove the silicone tube and the silk sutures, and irrigate the abdominal cavity with normal saline to minimize adhesion and infection risk. Reposition the intestine and suture the abdominal wall incision in layers using 4-0 non-absorbable sutures. Then suture the skin before transferring the rat into the recovery chamber for seven days. On postoperative day 14, section the kidneys into two halves to collect kidney samples. Store one half in 4% paraformaldehyde for histopathological examination, and rapidly freeze the other half in liquid nitrogen for storage at minus 80 degrees Celsius for subsequent molecular analysis. Perform the onscreen assessment for surgical follow-up. The morphology of the kidneys changed significantly following unilateral ureteral obstruction, with the UUO kidney appearing enlarged and swollen compared to the native kidney. After release of the obstruction, the kidney size reduced but remained larger than the native kidney. The ureteral obstruction was confirmed using methylene blue injection, which failed to pass through the blocked ureter. After the obstruction was released, the ureter at the ligation site exhibited a ruddy color and showed peristalsis. Histological analysis showed intact renal structures in the native kidney, whereas the UUO kidney exhibited tubular atrophy, vacuolation, and cell debris within the lumens. The RUUO kidney demonstrated partial recovery, with fewer damaged tubules than the UUO kidney. Masson's trichrome staining revealed severe interstitial fibrosis in the UUO kidney, indicated by increased blue-stained collagen fibers. The R-UUO kidney displayed reduced fibrosis compared to the UUO kidney. The renal damage score was significantly increased in the UUO group compared to the native group. The RUUO group had a lower renal injury score than the UUO group but remained significantly higher than the native kidney. The UUO group exhibited the largest kidney weight and volume compared to all other groups. The RUUO group showed a reduction in kidney weight and volume but did not fully return to control levels.