Mouse Ileal Loop Model: A Surgical Model to Study Intestinal Permeability in the Mouse

Under pathological conditions, the disintegration of the epithelial and endothelial barriers increases intestinal permeability, causing the intestinal contents to leak into the surrounding vasculature and abdominal cavity. The intestinal permeability can be simulated using ileal loop model.

To begin, prepare an anesthetized mouse in the supine position. Make a midline incision in the abdomen. Place a sterile cotton gauze moistened with a suitable buffer over the surgical area to keep the organs hydrated during surgery.

Exteriorize the ileum — the distal part of the intestine — and spread it over the gauze to expose the mesentery. Identify two ligation sites in the ileum that are spaced appropriately in the mesentery and are devoid of any critical blood vessels. Then, pierce the mesentery to add ligatures at the identified sites.

Next, dissect the ileum to generate an isolated ileal loop with an intact blood supply and mesenteric membrane. Thereafter, inject a suitable buffer into the loop to flush the waste content.

Now, ligate the open ends of the ileal loop. Use a syringe to inject chemokine—a chemotactic protein suspension—into the loop, causing it to inflate. Return the organs to the abdomen.

Suture the incision. Allow the mouse to recover. Eventually, the chemokine molecules pass through the ileal wall into the adjoining blood vessels, causing inflammation and leukocyte influx.

After achieving a surgical plane of anesthesia, begin by scrubbing the fur of the abdominal midline with alcohol swabs, or a gauze sponge soaked with 70% ethanol. Do not wet a wide area of fur with alcohol to prevent hypothermia. Using scissors, perform a midline laparotomy. Make a horizontal incision in the middle of the abdomen, and expose the peritoneum, taking care to not injure intra-abdominal organs.

Place pre-cut wet cotton gauze over the exposed intra-abdominal cavity. Use wet cotton swabs to mobilize and exteriorize the caecum, and carefully place it on wet cotton gauze. Then, mobilize and gently exteriorize the ileum. Deploy at least 6 centimeters of terminal ileum on the wet cotton gauze without disrupting the mesenteric vessels and the blood supply.

Keep the exposed tissues moist at all times with warm HBSS. Identify the major artery supplying the ileum in the mesentery close to the cecum. Then, locate two ligation sites in the mesentery that are free of critical blood vessels. Firmly grab the terminal ileum with blunt tissue forceps and use fine tip forceps to fenestrate the mesentery, avoiding blood vessels.

Place silk suture across the perforation and tie a surgical knot to create the first ligation. Use the ruler to measure 4 centimeters away from the first ligature, and create the second ligature. Carefully cut next to each ligation with fine scissors to isolate the 4-centimeter ileal loop, keeping the blood supply and mesenteric membrane intact.

Gently flush the content of the ileal loop segment with warm HBSS using a flexible yellow feeding tube attached to a 10-milliliter syringe. Make sure to flush the luminal contents out of the abdominal cavity to keep the surgical site clean. Ligate the two cut ends of the flushed ileal loop with silk suture.

Use a 1-milliliter syringe with a 30-gauge needle to slowly inject 250 microliters of reagent, such as FITC-dextrans or chemokine into the intestinal lumen. The ileal loop will inflate, causing a moderate distension of the mucosa.

Close the abdominal wall using a needle holder, anatomical forceps, and 3.0 non-absorbable silk sutures with a reverse cutting needle. After drying the animal to prevent hypothermia, place it in a temperature-regulated anesthesia chamber for the incubation period.