The effects of activated Akt/mTOR/p70S6K signaling pathway on the neural regeneration after spinal cord injury in rats

English Keywords:Spinal cord injury  mTOR  Rapamycin  Protein kinase signaling cascades  Neural regeneration  Rat

English Abstract:

  【Abstract】 Objectives: To examine the impacts of the protein kinase B(Akt)/mammalian target of rapamycin (mTOR)/p70 ribosomal S6 protein kinase(p70S6K) signaling transduction pathway on neural regeneration and functional recovery after spinal cord injury(SCI) in adult rats, and to provide the molecular mechanism for promoting the SCI repair． Methods: A total of 72 female Sprague-Dawley rats of minimal injury model was randomly divided into the following groups: activation group(Act group, SCI plus ATP), control group(Con group, SCI plus saline) and interruption group(Int group, SCI plus ATP and rapamycin). The sham-operated group(Sham group) including 24 rats undergoing laminectomy without SCI. The BBB locomotor rating scale was used to evaluate the animal′s locomotor performance after different treatments were administered at 1, 3, 7, and 14 days following SCI. The changes of Akt, p-Akt, mTOR, p-mTOR, p70S6K, p-p70S6K, Nestin and NeuN expressions in spinal cord tissues of all groups were detected by using Western blot analysis at all experimental time points after different disposals were administered following surgery, and the immunostaining was further performed for detecting the Nestin and NeuN in all groups after surgery. Results: Locomotor function assessment showed the BBB scores were 3-5 in the animals with SCI at 1d after injury, which gradually exhibited partial recovery of locomotor function after 1d following injury. No significant differences in BBB scores were observed among the three SCI groups at 1d and 3d after injury(P＞0.05). At 7d and 14d after injury, the rats in the Act group exhibited significantly higher BBB score compared with the rats in the Con and Int group(P＜0.05). Western blot showed prominently elevated phosphorylation level of Akt, mTOR and p70S6K in spinal cord tissues of all SCI groups at all experimental time points after surgery compared with the sham-operated group(P＜0.05). The phosphorylation level of this signaling pathway in the Act group was significantly elevated by exogenous administration of ATP compared with the Con and Int group at all experimental time points after surgery(P＜0.05), and the elevated level was particularly evident at 7d after SCI(P＜0.01). Prominently elevated levels of nestin were detected in the injured spinal cords of all SCI groups compared with the Sham group at 1, 3, 7 and 14 days after surgery(P＜0.05). A marked upregulation of nestin was noted in the Act group following SCI compared with the Con and Int group(P＜0.05), and the number of Nestin-positive cells in the Act group was more than that of the Con and Int group(P＜0.05). Significantly decreased expression of NeuN in the SCI groups was noted compared with the Sham group after surgery(P＜0.01), and no significant differences in the expression were observed among the three SCI groups at 1d and 3d after injury(P＞0.05). The expression of NeuN in the SCI groups gradually increased after 7d following SCI. The injured spinal cords in the Act group showed faster increase of NeuN and significantly higher level of NeuN expression compared with those from rats in the Con and Int group at 7d and 14d after SCI(P＜0.05), and the number of NeuN-positive cells in the Act group was more than that of the Con and Int group respectively(P＜0.05). Conclusions: Activated Akt/mTOR/p70S6K signaling pathway can significantly increase the expression of Nestin and NeuN in the injured spinal cords after SCI and posses the potential of promoting neural progenitor cells(NPCs) proliferation and neural regeneration, which improves motor function and promotes neural repair following SCI, intervention to this signaling pathway activity should be considered as a potential therapeutic strategy.