Stab-Wound Mouse Model for Studying Hemorrhage and Inflammation in Traumatic Brain Injury

Simplified traumatic brain injury (TBI) models have facilitated the development of therapeutic approaches. This protocol outlines the creation of a stab-wound mouse cortex using needles, enabling the analysis of hemorrhage and inflammation. The stab-wound TBI mouse model offers the advantage of being performed without requiring specialized equipment.

We are interested in developing a method to stop hemorrhage and suppress inflammation in TVI, so we established a protocol to easily make a TVI mass model using only a needle. Our stab wound TVI model is beneficial because it lubricate hemorrhage and gray activation. Our protocol has three main advantages compared to other techniques.

No specific equipment or tools are required. It can be done easily and quickly, and the wound is so tiny that it doesn't affect the mouse's behavior, so anyone can perform this with high reproducibility. Our laboratory will focus on the communication of neurons, gray cells, and glial cells, and parasites during the repair of traumatic brain injury.

Communication plays a critical role in the repair. However, it is not fully understood so far. Our protocol is very useful for analyzing communication.

To begin, place the anesthetized mouse on its ventral side on a paper towel. Confirm that the mouse is deeply anesthetized with a toe pinch test. Disinfect the surgical site by alternating rounds of Betadine and 70%ethanol scrub three times each.

Now, grasp the occipital skin with blunt forceps and make a one to 1.5 millimeter wide incision to expose the occipital bone without damaging the skull or any organ. Then gently and slowly open the incision to observe the boundary between the cerebral cortex and cerebellum through the skull. To perform mouse craniotomy, create a small hole in the right hemisphere of the occipital bone with a needle.

Gently rotate the needle to make an insertion point for stab wound needling, taking care not to damage the brain parenchyma. For the stab wound, insert the appropriate needle from the insertion point and stab the cerebral cortex along the rostral caudal axis. Finally, suture the skin incision using a nylon 3-0 suture with a half-round shaped needle.

The wound after the procedure was confirmed by microscopy. IGG extravasation peaked one day after the injury, followed by progressive recovery, and a reduction in staining intensity at three, five, and seven days. Evan's blue dye leakage into the brain parenchyma occurred immediately after injury with its concentration peaking by one hour.

The dye concentration gradually decreased after one and three days. Peak accumulation of IBA1 positive microglia and GFAP positive astrocytes occurred at five and three days post-injury respectively with their presence decreasing by seven days. Inflammatory cytokines, including tumor necrosis factor alpha, interleukin six, and interleukin one beta showed peak mRNA expression at one day post-injury while transforming growth factor beta one peaked at three days post-injury.