In Vivo and in Vitro Characterization of Novel Neuronal Plasticity Factors Identified following Spinal Cord Injury

Abstract

Following spinal cord injury, there are numerous changes in gene expression that appear to contribute to either neurodegeneration or reparative processes. We utilized high density oligonucleotide microarrays to examine temporal gene profile changes after spinal cord injury in rats with the goal of identifying novel factors involved in neural plasticity. By comparing mRNA changes that were coordinately regulated over time with genes previously implicated in nerve regeneration or plasticity, we found a gene cluster whose members are involved in cell adhesion processes, synaptic plasticity, and/or cytoskeleton remodeling. This group, which included the small GTPase Rab13 and actin-binding protein Coronin 1b, showed significantly increased mRNA expression from 7-28 days after trauma. Overexpression in vitro using PC-12, neuroblastoma, and DRG neurons demonstrated that these genes enhance neurite outgrowth. Moreover, RNAi gene silencing for Coronin 1b or Rab13 in NGF-treated PC-12 cells markedly reduced neurite outgrowth. Coronin 1b and Rab13 proteins were expressed in cultured DRG neurons at the cortical cytoskeleton, and at growth cones along with the pro-plasticity/regeneration protein GAP-43. Finally, Coronin 1b and Rab13 were induced in the injured spinal cord, where they were also co-expressed with GAP-43 in neurons and axons. Modulation of these proteins may provide novel targets for facilitating restorative processes after spinal cord injury.Traumatic injury to the spinal cord induces delayed biochemical responses that affect both cell loss and subsequent repair. Modulation of reparative processes includes factors involved in either facilitating or inhibiting neurite outgrowth, which are often regulated at the gene and protein levels after injury. Collectively, these factors likely determine in part the degree of anatomical and functional recovery after injury.Regeneration-associated proteins (RAGs) appear to play a role in plasticity and regeneration following SCI. 1 These include transcription factors (c-Jun), cytoskeletal components (T␣1), microtubule-associated proteins, growth-associated proteins (GAP-43, CAP-23), cell adhesion molecules (N-CAM, L1, TAG1), neurotrophic factors, cytokines, and extracellular matrix components (SNAP25, munc13, and cpg15/neuritin) (1-10). In some cases, these factors share common molecular pathways. GAP-43 and CAP-23 bind downstream to the cofactor PI(4,5)P(2), at plasmalemmal rafts, contributing to the regulation of actin and modulating neurite outgrowth in neuronallike cell lines (11,12). In other instances, a common downstream effector, such as neurotrophin-dependent intracellular cAMP, may serve to facilitate axonal regeneration by overcoming inhibition from factors such as myelin-associated glycoprotein (MAG) (13)(14)(15).Microarray technology provides a powerful tool for identifying molecular pathways involved in either endogenous neurotoxicity or regeneration/plasticity after SCI (16 -19). It allows concurrent analysis of thousands of genes and the identification of c...

Activation of PERK Elicits Memory Impairment through Inactivation of CREB and Downregulation of PSD95 After Traumatic Brain Injury

“…Transfection experiments were performed using the NeuroPORTER transfection kit (Sigma-Aldrich) following a previously published protocol with modifications (Di Giovanni et al, 2005;Uchida et al, 2006;Chi and Nicol, 2007;Andres et al, 2011). The cells were rinsed once with Opti-MEM medium and incubated at 37°C for ϳ30 min.…”

“…Mice were deeply anesthetized with isoflurane and perfused transcardially with 0.9% saline solution. The brains were removed and stained using the Dil method as described previously (Stranahan et al, 2007;Erion et al, 2014). Coronal sections of 200 m thickness from the cortex were obtained using a vibratome.…”

A p53-CBP/p300 transcription module is required for GAP-43 expression, axon outgrowth, and regeneration

“…26 Importantly, GAP-43 was observed to be co-expressed with Coronin 1b and Rab13 in cultured neurons during axon outgrowth and in vivo within sprouting axons after spinal cord injury. 26,27 As a result, we asked whether axon outgrowth and GAP-43 could be regulated by p53 within a specific transcriptional environment.…”

Análise dos mecanismos de neuroplasticidade na porção lombar da medula espinal do rato submetida à lesão isquêmica fototrombótica e tratada pela injeção local de PEDF

“…Proteínas associadas à regeneração representam um papel importante na plasticidade neuronal, entre elas destacam-se a proteína associada ao microtúbulo-2 (MAP-2) e a proteína 43 associada ao crescimento (GAP-43) (Di Giovanni et al, 2005).…”

“…A proteína GAP-43 é um componente importante dos axônios e terminais pré-sinápticos e o aumento de sua expressão é visto durante o desenvolvimento neural e regeneração axonal (Benowitz e Routtenberg, 1997;Di Giovanni et al, 2005). A lesão isquêmica aqui aplicada lesou o funículo posterior da medula espinal, inclusive o tracto córtico-espinal dorsal, que nos ratos corresponde ao tracto córtico-espinal lateral, onde se encontra a maior parte das fibras motoras.…”