Targeting PTEN with lentiviral shRNA promotes neurite outgrowth of cortical neurons and improves functional recovery in a rat spinal cord contusion model

Abstract

Objective To observe the effects of PTEN silencing on axon regeneration and functional recovery after spinal cord injury (SCI). Methods Four groups were designed: DMEM、lenti-control、lenti-scramble、lenti-shRNA. Cortical neurons were seeded on or adjacent to chondroitin sulfate proteoglycans (CSPGs). Three days after lentivirus infection, the expression of PTEN were assessed by Western blot, and immunocytochemistry method were applied to examine the length, number and crossing behavior of neurites; Lentivirus was locally injected into the conical motor area of the rats with spinal cord injury. 1 week later, the expression of PTEN was assessed by Western blot. 6 weeks later, motor function of hind limbs was evaluated by Basso Beattie Bresnahan (BBB) score, while the expressions of green fluorescence in injured spinal cord were observed by fluorescence microscope, and immunohistochemical method was used to observe the expression of synaptophysin changes in spinal tissues. Results Less expression of PTEN was detected in lentivirus-shRNA, and neurites with PTEN silencing exhibited significant enhancements in elongation, initiation and crossing ability when they encountered CSPGs in vitro. Better hindlimb functional recovery, less expression of PTEN and more synaptophysin positive response area were detected in rats injected with lentivirus-shRNA compared with other groups. Moreover, green fluorescence was found in the caudal region of injured spinal cord of rats injected with lentivirus-shRNA. Conclusion Lentiviral-mediated PTEN-shRNA can significantly improve the axon regeneration and neurological functional recovery after spinal cord injury. Key words: Spinal cord injuries; RNA interference; Lentivirus; Axons; Regeneration