A Protocol for Constructing a Rat Wound Model of Type 1 Diabetes

Many details of the Type 1 diabetes wound modeling process are prone to error. It is a difficult challenge for experienced researchers. As a result, it is necessary to utilize this protocol.

In the simplest way possible, this technique can effectively replicate some characteristics of chronic diabetic wounds such as no wound healing, delayed re-epithelialization, and reduced angiogenesis. The streptozotocin solution should be used immediately after preparation. When creating the wound, look that the size and location of the wound must be regulated.

Begin by dissolving the streptozotocin powder in an appropriate amount of 0.1 moles per liter sodium citrate buffer to achieve a concentration of 1%and shake it for 30 seconds using a drug oscillator before placing it in a nice box. For intraperitoneal injection of 1%streptozotocin solution, grasp the rat and expose the abdominal skin at the injection site. Disinfect the injection site twice using a cotton ball soaked in 75%alcohol and place the rat's head below the abdomen.

Insert the needle parallel to the abdominal midline at 45 degrees. Then reduce the needle angle to 30 degrees and insert two to three millimeters. Gently pull the needle plug, inject the streptozotocin solution, and pull out the needle before applying a cotton swab.

To measure the causal blood glucose levels at 9:00 AM on day 3 and day 7, find the location of the caudal vein. Disinfect the rat tail twice using 75%alcohol before punctioning the caudal vein to induce bleeding and measure the blood glucose using a glucometer. Once done, stop the bleeding with a cotton swab.

Shave the 5 by 5 centimeter area on the dorsum side of the rat with an electric razor one day before wound modeling. Wipe the shaved area with a warm normal saline cotton ball and allow it to dry for applying depilatory cream for five minutes. Clean the area with gauze and wash any residual depilatory cream with warm normal saline.

Then disinfect the dorsal skin two times using cotton balls soaked in iodine, and 75%alcohol in the alternative rounds. After drying, cut the skin with a 20 millimeter diameter circular biopsy punch. Tend the skin with forceps and then use surgical scissors to remove the full thickness of skin along the punch cut marks.

Once done, stop the bleeding with a normal saline cotton ball. Cover the wound using Vaseline gauze. Diabetes was induced in all the rats after three days of streptozotocin injection, with the blood glucose level higher than 16.7 millimoles per liter.

The glucose level was stabilized five weeks after the induction. The weight of the diabetic group increased gradually after the streptozotocin injection. Weight decreased in week 3 and slowly increased again from week 4.

In contrast, the weight of the normal group rat increased steadily. Their mean weight after the three days of diabetes induction was higher than the diabetic group. The macroscopic analysis performed on days 7 and 14 after the wounding revealed that the re-epithelialization was more pronounced in rats in the normal group than in the diabetic group.

The quantitative results of day 7 and day 14 revealed that the wound healing rate was significantly lower in the diabetic group than in the normal group. However, on day 14th, the wound healing rate was above 90%in the diabetic group. The diabetic wounds showed partial loss of the hair follicles and sebaceous glands with fewer visible capillaries.

In wound angiogenesis analyzed by CD31 immunostaining, the average optical density of CD31 expression showed that the angiogenesis at the wound site was significantly higher in the normal than in the diabetic group. When modeling the wound, ensure that the fore thickness skin is removed including the epidermis, dermis, and subcutaneous tissue.