RESUMEN
ALS is a fatal neurodegenerative disease characterized by a progressive loss of motor neurons and atrophy of distal axon terminals in muscle, resulting in loss of motor function. Motor end plates denervated by axonal retraction of dying motor neurons are partially reinnervated by remaining viable motor neurons; however, this axonal sprouting is insufficient to compensate for motor neuron loss. Activating transcription factor 3 (ATF3) promotes neuronal survival and axonal growth. Here, we reveal that forced expression of ATF3 in motor neurons of transgenic SOD1(G93A) ALS mice delays neuromuscular junction denervation by inducing axonal sprouting and enhancing motor neuron viability. Maintenance of neuromuscular junction innervation during the course of the disease in ATF3/SOD1(G93A) mice is associated with a substantial delay in muscle atrophy and improved motor performance. Although disease onset and mortality are delayed, disease duration is not affected. This study shows that adaptive axonal growth-promoting mechanisms can substantially improve motor function in ALS and importantly, that augmenting viability of the motor neuron soma and maintaining functional neuromuscular junction connections are both essential elements in therapy for motor neuron disease in the SOD1(G93A) mice. Accordingly, effective protection of optimal motor neuron function requires restitution of multiple dysregulated cellular pathways.
Asunto(s)
Factor de Transcripción Activador 3/metabolismo , Esclerosis Amiotrófica Lateral/fisiopatología , Neuronas Motoras/patología , Músculo Esquelético/inervación , Esclerosis Amiotrófica Lateral/patología , Animales , Supervivencia Celular , Modelos Animales de Enfermedad , Ratones , Superóxido Dismutasa/genéticaRESUMEN
Peripheral axons of dorsal root ganglion (DRG) neurons, but not their central axons in the dorsal columns, regenerate after injury. However, if the neurons are conditioned by a peripheral nerve injury into an actively growing state, the rate of peripheral axonal growth is accelerated and the injured central axons begin to regenerate. The growth-promoting effects of conditioning injuries have two components, increased axonal growth and a reduced response to inhibitory myelin cues. We have examined which transcription factors activated by peripheral axonal injury may mediate the conditioning effect by regulating expression of effectors that increase the intrinsic growth state of the neurons. Activating transcription factor 3 (ATF3) is a prime candidate because it is induced in all injured DRG neurons after peripheral, but not central, axonal damage. To investigate if ATF3 promotes regeneration, we generated transgenic mice that constitutively express this transcription factor in non-injured adult DRG neurons. The rate of peripheral nerve regeneration was enhanced in the transgenic mice to an extent comparable to that produced by a preconditioning nerve injury. The expression of some growth-associated genes, such as SPRR1A, but not others like GAP-43, was increased in the non-injured neurons. ATF3 increased DRG neurite elongation when cultured on permissive substrates but did not overcome the inhibitory effects of myelin or promote central axonal regeneration in the spinal cord in vivo. We conclude that ATF3 contributes to nerve regeneration by increasing the intrinsic growth state of injured neurons.
Asunto(s)
Factor de Transcripción Activador 3/fisiología , Ganglios Espinales/crecimiento & desarrollo , Ganglios Espinales/metabolismo , Regeneración Nerviosa/fisiología , Neuronas/fisiología , Animales , Células Cultivadas , Femenino , Ganglios Espinales/citología , Humanos , Ratones , Ratones Transgénicos , Neuronas/metabolismo , Nervios Periféricos/crecimiento & desarrollo , Nervios Periféricos/metabolismoRESUMEN
Dorsal root ganglion (DRG) neurons regenerate after a peripheral nerve injury but not after injury to their axons in the spinal cord. A key question is which transcription factors drive the changes in gene expression that increase the intrinsic growth state of peripherally injured sensory neurons? A prime candidate is activating transcription factor-3 (ATF-3), a transcription factor that we find is induced in all DRG neurons after peripheral, but not central axonal injury. Moreover, we show in adult DRG neurons that a preconditioning peripheral, but not central axonal injury, increases their growth, correlating closely with the pattern of ATF-3 induction. Using viral vectors, we delivered ATF-3 to cultured adult DRG neurons and find that ATF-3 enhances neurite outgrowth. Furthermore, ATF-3 promotes long sparsely branched neurites. ATF-3 overexpression did not increase c-Jun expression. ATF-3 may contribute, therefore, to neurite outgrowth by orchestrating the gene expression responses in injured neurons.
Asunto(s)
Factor de Transcripción Activador 3/metabolismo , Ganglios Espinales/metabolismo , Regeneración Nerviosa/genética , Neuritas/metabolismo , Neuronas Aferentes/metabolismo , Nervios Periféricos/metabolismo , Factor de Transcripción Activador 3/genética , Animales , Células Cultivadas , Ganglios Espinales/citología , Ganglios Espinales/lesiones , Regulación de la Expresión Génica/fisiología , Vectores Genéticos/fisiología , Conos de Crecimiento/metabolismo , Conos de Crecimiento/ultraestructura , Neuritas/ultraestructura , Neuronas Aferentes/citología , Traumatismos de los Nervios Periféricos , Nervios Periféricos/citología , Proteínas Proto-Oncogénicas c-jun/metabolismo , Ratas , Ratas Sprague-Dawley , Rizotomía/efectos adversos , Raíces Nerviosas Espinales/citología , Raíces Nerviosas Espinales/lesiones , Raíces Nerviosas Espinales/metabolismo , Regulación hacia Arriba/fisiologíaRESUMEN
Delivery of genes into DRG neurons by viral vectors is a powerful tool for the study of axonal outgrowth. In order to achieve efficient transfer of growth-related genes and simultaneously label neuronal processes, we have utilized the HSV-based amplicon vector system. A bicistronic expression cassette encoding the growth associated protein-43 (GAP-43) and the axonal marker human placental alkaline phosphatase (hPLAP) reporter gene under translation control of an internal ribosomal entry site was cloned into the HGCX amplicon vector. This hPLAP reporter enabled efficient labeling of neurites in both dissociated adult DRG neurons and embryonic DRG explants. Using this reporter, the effect of GAP-43 on neurite outgrowth in transduced DRG neurons could be demonstrated. HSV-based amplicon vectors can contribute to the study of axonal growth and guidance in cultured neurons.