Neuropathic Pain Model of Peripheral Neuropathies Mediated by Mutations of Glycyl-tRNA Synthetase

Abstract

Charcot-Marie-Tooth disease (CMT) is the most common inherited motor and sensory neuropathy. Previous studies have found that, according to CMT patients, neuropathic pain is an occasional symptom of CMT. However, neuropathic pain is not considered to be a significant symptom associated with CMT and, as a result, no studies have investigated the pathophysiology underlying neuropathic pain in this disorder. Thus, the first animal model of neuropathic pain was developed by our laboratory using an adenovirus vector system to study neuropathic pain in CMT. To this end, glycyl-tRNA synthetase (GARS) fusion proteins with a FLAG-tag (wild type [WT], L129P and G240R mutants) were expressed in spinal cord and dorsal root ganglion (DRG) neurons using adenovirus vectors. It is known that GARS mutants induce GARS axonopathies, including CMT type 2D (CMT2D) and distal spinal muscular atrophy type V (dSMA-V). Additionally, the morphological phenotypes of neuropathic pain in this animal model of GARS-induced pain were assessed using several possible markers of pain (Iba1, pERK1/2) or a marker of injured neurons (ATF3). These results suggest that this animal model of CMT using an adenovirus may provide information regarding CMT as well as a useful strategy for the treatment of neuropathic pain.Graphical Abstract

Novel animal models of GARS-associated neuropathy by overexpression of mutant GARS using an adenoviral vector

“…Animal models enable manifestation of morphological and functional phenotypes (Lee et al 2014;Seo et al 2014aSeo et al , 2014bSeo et al , 2014c. The promoter of the mouse choline acetyltransferase (ChAT) gene, which includes a neuron-restrictive silencer element and cholinergic-specific enhancer, key regulatory elements for neuron-specific expression, has been used in these models (Misawa et al 1992;Lönner-berg et al 1996).…”

“…The promoter of the mouse choline acetyltransferase (ChAT) gene, which includes a neuron-restrictive silencer element and cholinergic-specific enhancer, key regulatory elements for neuron-specific expression, has been used in these models (Misawa et al 1992;Lönner-berg et al 1996). Neuron-specific recombinant adenoviruses are injected into the sciatic nerve for anterograde and retrograde transduction (Lee et al 2014;Seo et al 2014aSeo et al , 2014bSeo et al , 2014c.…”

“…Because the expressed GFP is translated from a distinct transcript driven by a CMV promoter, Schwann cells and axons surrounding the long peripheral axon cell bodies infected by the virus vectors exhibit GFP-positive signals (Lee et al 2014;Seo et al 2014aSeo et al , 2014b. However, specific expression of WT or mutant GARS in peripheral neurons can be identified though detection of the flag-tag linked to the GARS gene terminus (Lee et al 2014;Seo et al 2014aSeo et al , 2014b. Thus, in vivo application of adenoviruses is highly efficient for the induction of neuron-specific expression of flag-tagged hGARS fusion proteins in long peripheral axons.…”

“…By evaluating several markers for pain induction and nerve damage, including p-ERK, activating transcription factor 3 (ATF3) and Iba1 (Ma & Quirion 2002;Ivanavicius et al 2007;Bráz & Basbaum 2010;Maeda et al 2010), this model was used to demonstrate the anatomical and functional phenotypes of neuropathic pain (Lee et al 2014).…”

“…The ability of adenoviral vectors to deliver GARS mutants specifically into peripheral nerve axons has allowed experimental analysis of the pathological processes involved in the abnormal localization of GARS mutants (Seo et al 2014a(Seo et al , 2014b. In addition, the use of adenoviral vectors has led to the generation of GARS axonopathy animal models of neuropathic pain and motor impairment, which are symptoms in human patients (Lee et al 2014;Seo et al 2014c). Therefore, highly efficient adenoviral vectors for in vivo application in peripheral nerves offer an approach to overcome the limitations of using genetically modified transgenic mice, which lack specificity to long peripheral axons.…”