RESUMEN
Gangliosides are glycosphingolipids abundantly expressed in the vertebrate nervous system, and are classified into a-, b-, or c-series according to the number of sialic acid residues. The enzyme GD3 synthase converts GM3 (an a-series ganglioside) into GD3, a b-series ganglioside highly expressed in the developing and adult retina. The present study evaluated the visual system of GD3 synthase knockout mice (GD3s-/- ), morphologically and functionally. The absence of b- series gangliosides in the retinas of knockout animals was confirmed by mass spectrometry imaging, which also indicated an accumulation of a-series gangliosides, such as GM3. Retinal ganglion cell (RGC) density was significantly reduced in GD3s-/- mice, with a similar reduction in the number of axons in the optic nerve. Knockout animals also showed a 15% reduction in the number of photoreceptor nuclei, but no difference in the bipolar cells. The area occupied by GFAP-positive glial cells was smaller in GD3s-/- retinas, but the number of microglial cells/macrophages did not change. In addition to the morphological alterations, a 30% reduction in light responsiveness was detected through quantification of pS6-expressing RGC, an indicator of neural activity. Furthermore, electroretinography (ERG) indicated a significant reduction in RGC and photoreceptor electrical activity in GD3s-/- mice, as indicated by scotopic ERG and pattern ERG (PERG) amplitudes. Finally, evaluation of the optomotor response demonstrated that GD3s-/- mice have reduced visual acuity and contrast sensitivity. These results suggest that b-series gangliosides play a critical role in regulating the structure and function of the mouse visual system.
Asunto(s)
Sensibilidad de Contraste/fisiología , Eliminación de Gen , Retina/enzimología , Sialiltransferasas/deficiencia , Sialiltransferasas/genética , Agudeza Visual/fisiología , Animales , Electrorretinografía/métodos , Femenino , Masculino , Ratones , Ratones de la Cepa 129 , Ratones Noqueados , Estimulación Luminosa/métodosRESUMEN
After an injury, axons in the central nervous system do not regenerate over large distances and permanently lose their connections to the brain. Two promising approaches to correct this condition are cell and gene therapies. In the present work, we evaluated the neuroprotective and neuroregenerative potential of pigment epithelium-derived factor (PEDF) gene therapy alone and combined with human mesenchymal stem cell (hMSC) therapy after optic nerve injury by analysis of retinal ganglion cell survival and axonal outgrowth. Overexpression of PEDF by intravitreal delivery of AAV2 vector significantly increased Tuj1-positive cells survival and modulated FGF-2, IL-1ß, Iba-1, and GFAP immunostaining in the ganglion cell layer (GCL) at 4 weeks after optic nerve crush, although it could not promote axonal outgrowth. The combination of AAV2.PEDF and hMSC therapy showed a higher number of Tuj1-positive cells and a pronounced axonal outgrowth than unimodal therapy after optic nerve crush. In summary, our results highlight a synergistic effect of combined gene and cell therapy relevant for future therapeutic interventions regarding optic nerve injury.
Asunto(s)
Proteínas del Ojo/farmacología , Factores de Crecimiento Nervioso/farmacología , Traumatismos del Nervio Óptico/terapia , Células Ganglionares de la Retina/efectos de los fármacos , Serpinas/farmacología , Animales , Axones/fisiología , Línea Celular Tumoral , Supervivencia Celular , Tratamiento Basado en Trasplante de Células y Tejidos/métodos , Modelos Animales de Enfermedad , Proteínas del Ojo/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica/genética , Humanos , Masculino , Trasplante de Células Madre Mesenquimatosas/métodos , Células Madre Mesenquimatosas/metabolismo , Compresión Nerviosa , Factores de Crecimiento Nervioso/metabolismo , Regeneración Nerviosa , Neuroprotección , Nervio Óptico , Ratas Wistar , Retina , Células Ganglionares de la Retina/metabolismo , Serpinas/metabolismoRESUMEN
BACKGROUND: Optic-nerve injury results in impaired transmission of visual signals to central targets and leads to the death of retinal ganglion cells (RGCs) and irreversible vision loss. Therapies with mesenchymal stem cells (MSCs) from different sources have been used experimentally to increase survival and regeneration of RGCs. METHODS: We investigated the efficacy of human umbilical Wharton's jelly-derived MSCs (hWJ-MSCs) and their extracellular vesicles (EVs) in a rat model of optic nerve crush. RESULTS: hWJ-MSCs had a sustained neuroprotective effect on RGCs for 14, 60, and 120 days after optic nerve crush. The same effect was obtained using serum-deprived hWJ-MSCs, whereas transplantation of EVs obtained from those cells was ineffective. Treatment with hWJ-MSCs also promoted axonal regeneration along the optic nerve and reinnervation of visual targets 120 days after crush. CONCLUSIONS: The observations showed that this treatment with human-derived MSCs promoted sustained neuroprotection and regeneration of RGCs after optic nerve injury. These findings highlight the possibility to use cell therapy to preserve neurons and to promote axon regeneration, using a reliable source of human MSCs.
Asunto(s)
Trasplante de Células Madre Mesenquimatosas , Células Ganglionares de la Retina , Animales , Axones , Supervivencia Celular , Humanos , Regeneración Nerviosa , Nervio Óptico , RatasRESUMEN
Mitochondrial dysfunctions are linked to a series of neurodegenerative human conditions, including Parkinson's disease, schizophrenia, optic neuropathies, and glaucoma. Recently, a series of studies have pointed mitotherapy - exogenous mitochondria transplant - as a promising way to attenuate the progression of neurologic disorders; however, the neuroprotective and pro-regenerative potentials of isolated mitochondria in vivo have not yet been elucidated. In this present work, we tested the effects of transplants of active (as well-coupled organelles were named), liver-isolated mitochondria on the survival of retinal ganglion cells and axonal outgrowth after optic nerve crush. Our data show that intravitreally transplanted, full active mitochondria incorporate into the retina, improve its oxidative metabolism and electrophysiological activity at 1 day after transplantation. Moreover, mitotherapy increases cell survival in the ganglion cell layer at 14 days, and leads to a higher number of axons extending beyond the injury site at 28 days; effects that are dependent on the organelles' structural integrity. Together, our findings support mitotherapy as a promising approach for future therapeutic interventions upon central nervous system damage.