This article describes the implantation of renal cell carcinoma (RCC) cells into the kidneys of immunocompromised mice to create an orthotopic RCC model. The process involves surgical techniques to ensure proper cell delivery and monitoring of tumor growth.
The implantation of renal cell carcinoma, or RCC, cells from a donor mouse to an immunocompromised recipient mouse's kidney leads to the development of an orthotopic RCC model.
To develop an orthotopic RCC model, begin by placing an anesthetized immunodeficient mouse in the prone position on a dissection board. Now, use a scalpel to make a surgical cut on the left flank of the mouse, keeping the underlying peritoneum intact.
Palpate the mouse by applying gentle pressure from the bottom. This step helps visualize the kidney through the translucent peritoneum. Next, take a syringe containing RCC cell suspension in an appropriate medium. Using a needle, inject the cell suspension into the kidney without damaging the peritoneum.
Retract the needle and seal the incision site using a tissue adhesive. Allow the mouse to recover and monitor the tumor growth at desired intervals. Initially, the injected renal cancer cells get implanted into the kidney. These cells then proliferate to form tumors.
Subsequently, a few cells may detach from the primary tumor and disseminate within the peritoneal cavity. These migrating tumor cells can invade the lung tissue and metastasize to form secondary tumors.
To prepare the Renca tumor cells, first, attach the cell culture with 0.25% trypsin in HBSS after rinsing with PBS. Neutralize the reaction after 5 minutes with complete RPMI medium. Transfer the cells to a conical tube for centrifugation and resuspend the pellet in fresh HBSS.
After counting, adjust the volume to a 2 times 10 to the sixth cells per milliliter concentration in HBSS, and draw the cells into a 1-milliliter syringe equipped with an 18-gauge needle. Next, place a sterile surgical drape over a heating pad and confirm the appropriate level of sedation by toe pinch. Remove the hair from the left flank of the mouse and apply vet ointment to the eyes.
Then, swab the implantation site with 5% povidone-iodine antiseptic, and use a sterile scalpel to make a single 1 to 1.5-centimeter incision on the left flank, taking care not to penetrate the peritoneum. Using scissors, separate the dermis from the peritoneum, dabbing the incision site with sterile cotton gauze to remove any blood. Then, grasp the head with the left hand and support the flank with the right hand to locate the spleen through the peritoneum.
Using one finger from the right hand, palpate the mouse from underneath. The kidneys should become visible. Keeping a firm hold on the animal to provide tension across the peritoneum and kidney, have an assistant slowly deliver 0.1 milliliters of the Renca cells through the intact peritoneum into the center of the kidney. Hold the needle in place for 5 to 10 seconds to reduce the backflow of the cells. Then, close the incision with a thin layer of cyanoacrylate tissue adhesive.
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