Automated Impactor for Contusive Spinal Cord Injury Model in Mice

My research scope includes the pathogenesis and the treatment methods of spinal cord injury. Our recent research has found that various biomaterials can promote the repair of spinal cord injury in mice through antioxidant stress. The development of biomaterial sensors, technologies, and the gene modification technology, as well as the optimization of spinal cord injury models are advancing our research field.

Due to compress and incompletely understood mechanism of spinal cord injury, it is currently difficult to cure spinal cord injury using a single intervention. In the future, multiple intervention methods may be needed for research. We found that mental immediate compress can participate in the treatment and the repair of spinal cord injury in animals through antioxidant stress.

To begin, shave the hair on the back of an eight-week-old female anesthetized mouse. Disinfect the skin three times with alternating rounds of iodophor and alcohol. Now, use a scalpel to make a 2.5-centimeter long medial incision in the dorsal skin.

Cut through the muscles till the backbone is seen. Dab any blood with cotton. With tweezers, expose the spine at the T9-T11 level.

Using a spinal fixator, fix T10 facets bilaterally, ensuring the stable fixation of the spine. Then, use a micro-grinding drill to strip the paravertebral muscles. Now remove the spinous processes and the laminae to expose the spinal cord at the T10 segment.

Next, switch on the impactor instrument and wait for the device to return to its original state automatically. Place the spinal fixator with the anesthetized mouse into the G smart SCI system. Then, firmly secure it using screws.

Using the operation touchscreen, set the impact speed, impact depth, and dwell time. Move the platform to align the laser rangefinder to the center of the exposed spinal cord. Click the ready button on the touch screen.

The impact head will automatically adjust to a specific height based on the setting parameters, and the carrier table will automatically move the spinal cord impact site below the impact head. Press the impact head manually to further determine the exact impact site. Then, click the start button.

This will cause the impact head to strike the spinal cord. Remove the mouse from the device and place it under a stereo microscope at 20x magnification to observe the spinal cord injuries. Look for injury markers such as localized congestion, collapse and spinal membrane rupture to confirm the success of model development.

To begin, calculate the Basso Mouse Scale scores for mice starting from the first postoperative day. On day 30 after surgery, performed the behavioral tests including catwalk, foot fault, and rotarod tests. For the catwalk test, record the mouse walking a 45-centimeter distance with a maximum run duration of eight seconds.

To perform the foot fault test, record the mouse walking 60 steps. For the rotarod test, note the time it takes for the mouse to fall off the rotarod, spinning at a speed of 20 revolutions per minute. On the 31st day postoperation, carefully remove the spinal cord of the euthanized mouse.

Cut the excised cord five millimeters above and below the injury site to prepare for paraffin embedding. Prepare five-micrometer paraffin sections from the center of the inflicted spinal cord injury with a microtome and perform Hematoxylin and eosin staining. Conduct statistical analysis using commercial software.

After one month, mice in the 0.5 millimeter group showed four to six postoperative scores and recovered similarly to the sham group. Mice in the 0.8 millimeter and 1.1 millimeter groups had one to two postoperative scores. The 0.8 millimeter group recovered to four to six scores after a month while the 1.1 millimeter group barely recovered.

In the foot fault test, no significant differences in hind limb foot fault were observed between the 0.5 millimeter group and the sham group. The mice in the 1.1 millimeter group had 100%foot fault. The 0.8 millimeter and 1.1 millimeter groups showed significantly different results in the rotarod test.

However, the sham and 0.5 millimeter groups were similar. All groups responded in significantly different ways in the catwalk test. Spinal resection demonstrated varying degrees of damage in the spinal cord images as well as in the Hematoxylin and eosin stained sections.