GA alleviates axon and myelin loss and decreases lesion volume after SCI
To examine the correlation between behavioral data and histological outcomes such as axon loss, myelin loss, and lesion volume, we performed histological analysis of the spinal cord tissues from the animals used for behavioral experiments. First, we performed immunostaining with NF200 and 5-HT antibodies to detect remaining axons and the density of preserved axons. In the sham control, NF200-positive axons in both ventral and dorsolateral funiculi were dense, and axonal packing was uniform (Fig. 9A and B, Sham). However, the axon density was markedly decreased in the injured tissue (Fig. 9A and B, Veh). The number of NF200 positive axons in both ventral and dorsolateral funiculi was significantly higher in the GA-treated group compared to that in the vehicle-treated group (Fig. 9A and B). Moreover, the density of 5-HT serotonergic axons in the ventral horn was higher in the GA-treated group compared to that in the vehicle-treated group (Fig. 9C). These results suggest that GA treatment alleviates axon loss after SCI. Next, we measured the extent of myelin loss after injury by Luxol fast blue staining. As shown in Fig. 9D, extensive myelin loss near the lesion area was evident in the vehicle-treated group at 35 d after injury compared with that of the sham control (Fig. 9D, Veh), whereas GA treatment apparently alleviated myelin loss (Fig. 9D, GA).
To evaluate tissue loss after SCI, serial longitudinal sections were stained with Cresyl violet, and the lesion volume was measured. As shown in Fig. 9E, the total lesion volume was significantly reduced in the GA-treated group at 35 d after injury compared to that of the vehicle-treated group (Fig. 9E).