Analysis of the C/EBP family of transcription factors in neuronal repair

Abstract

Neurons within the peripheral nervous system (PNS) have a remarkable ability to repair themselves after injury; however, neurons within the central nervous system (CNS) do not spontaneously regenerate. Therefore, understanding the molecular elements responsible for successful regenerative response in the PNS can help us to establish basic principles and strategies for promoting regeneration in CNS structures such as the spinal cord. Nerve repair in the PNS has been suggested to be in part due to the involvement of intrinsic molecules such as transcription factors. In this thesis, I am focusing on the C/EBP family of transcription factors and their potential role in axonal regeneration after PNS injury. I examined the expression of different C/EBP members in PNS after injury after, using the sciatic nerve crush injury model I found that C/EBPδ mRNA is upregulated 4, 24 and 72 hours in dorsal root ganglia (DRG) following injury, whereas C/EBPβ and C/EBPγ expression is transiently upregulated by 4 hours resuming background levels after 72 hours. Conversely, C/EBPα and C/EBPε did not show upregulation following injury. In order to determine the function of C/EBPs in axonal growth in an in vitro system I used the ND7/23 cell line where I found that upon neurite growth induced by cyclic adenosine monophosphate (cAMP), the mRNA levels of C/EBPβ and C/EBPδ were upregulated. Furthermore, the conditional expression of a C/EBP total inhibitor or a C/EBPδ antisense construct decreases neurite elongation in vitro. Additionally, I found that 24 hours after treatment of ND7/23 cells with trichostatin A (TSA) C/EBPδ expression is elevated. Subsequently, I found that in DRG cultures from C/EBPδ knock-out animals, the lack of C/EBPδ affects the intrinsic growth capacity of dorsal root ganglion neurons which show a 3 drastically reduced axonal growth in vitro. To address the role of C/EBPδ in vivo, peripheral nerve repair was assessed in transgenic C/EBPδ knock-out animals following sciatic nerve crush. C/EBPδ knock-outs show, by immunostaining, impaired nerve regeneration 3 days and 14 days after sciatic nerve injury. Furthermore, functional recovery and morphometric analysis indicate that nerve regeneration is delayed in C/EBPδ deficient animals. These data demonstrate that the C/EBPδ gene is involved in neuronal repair after peripheral nerve injury.