Establishment of Coloproctitis Cancer Model in Mice and Evaluation of Therapeutic Effect of Chinese Medicine

This protocol provides a mouse model of ulcerative coloproctitis-associated colorectal cancer induced by azomethane combined with dextran sulfate sodium. The model was used to evaluate the efficacy of traditional Chinese medicine compounds in the prevention and treatment of colorectal cancer.

This study was on the prevention and treatment of coloproctitis cancer with traditional Chinese medicine. We aim to establish a mouse model for coloproctitis cancer and evaluate traditional Chinese medicine fixed indicators. We confirmed that the Chinese medicine Liujunzi decoction is a effective treatment for coloproctitis cancer prevention.

In the future, we will work to identify more sensitive biomarkers for colon and rectal cancer and screen Liujunzi decoction for therapeutic targets. To begin, place Danshen, BaiZhu, Danggui, Chenpi, and Jiangbanxia into a specialized ceramic pot. Add 1000 milliliters of distilled water and let it soak at room temperature for one hour.

Using a grinder, grind 12 grams of Fuling into a fine powder. Then soak the powdered Fuling in 300 milliliters of distilled water in a separate container for one hour at room temperature. Mix the prepared Chinese herbal mixture in the specialized ceramic pot.

Boil the mixture and maintain it over medium heat until the decoction volume reduces to 300 milliliters. Combine the filtrate obtained from the decoction and continue boiling until the volume reduces to 150 milliliters. Then centrifuge the concentrated liquid at 10, 000G for five minutes.

Concentrate the obtained supernatant to 30 milliliters over medium heat. Transfer the final concentrate to a dish and dry it using an electric drying oven until only the solute remains as a powder. Now weigh the obtained powder and dissolve it in sterile distilled water to create the daily dosage for mice.

To prepare 5-aminosalicylic acid, weigh 64 milligrams of 5-aminosalicylic acid powder and dissolve it in 200 milliliters of sterile distilled water. To begin, gently position the mouse with its belly up and the head slightly down. Grasp the back skin to secure the abdominal area and insert a one milliliter syringe needle one centimeter to the right of the midline of the abdomen at the intersection of the line connecting both thighs.

Gradually insert the one milliliter syringe needle to a depth of three to five millimeters under the skin. Insert the needle approximately 0.3 to 0.5 millimeters into the abdominal cavity at a 45 degree angle. Once the needle tip passes through the abdominal muscle, note a sudden decrease in resistance.

Slowly inject 0.1 milliliters of the Azoxymethane injection solution per 10 grams of body weight into the mice. Subsequently retract the syringe gently. Then add 500 milliliters of sterile distilled water to 10 grams of DSS to prepare a 2%DSS solution.

Mix the solution with a vortex mixer and store at four degrees Celsius until use. Administer the prepared LJZD and 5-aminosalicylic acid solution daily at a consistent time via gastric gavage to the respective groups at week 7 and week 15. To begin, establish the UC-CRC mouse model and administer the prepared LJZD in 5-ASA solution to the respective groups.

After the drug treatment, monitor the weight of mice daily from the beginning of adaptive feeding until the end of drug treatment. Diligently observe and document the fecal consistency of each experimental mouse, categorizing it as normal, loose stools, or watery diarrhea. Carefully document any fecal bleeding observed in the experimental animals, categorizing it into no bleeding, minimal bleeding, or visible blood in the stool.

To separate the colorectal tissue, maintain the euthanized mice within a cryogenic anatomical environment. Secure the mice in a supine position. Using scissors, trim the lower abdominal hair and sterilize the area with ethanol.

Using eyelid forceps, gently grasp the intersection point between the two thigh roots and the abdominal midline. Then use scissors to create a 1 to 1.5 centimeters long transverse incision. Perform a longitudinal incision along the median line of the abdomen, starting from the midpoint of the transverse incision to the xiphoid process.

Carefully detach the pericolorectal tissues in the direction towards the anus, separating the colorectum from the surrounding tissue. Gently push aside the abdominal skin to fully expose the colorectum. Using eyelid forceps, extract the colorectum from the abdominal cavity and section it from the anus to the cecum.

Preserve the collected colorectal tissues in saline at four degrees Celsius. The UC-CRC model group had a significant weight loss compared to the control group which was alleviated by LJZD treatment. LJZD treatment improved the disease activity index compared with the UC-CRC model group.