RESUMEN
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
Vision loss is a devastating consequence of systemic hypoxia, but the cellular mechanisms are unclear. We investigated the impact of acute hypoxia in the retina and optic nerve. We induced systemic hypoxia (10% O2) in 6-8w mice for 48 h and performed in vivo imaging using optical coherence tomography (OCT) at baseline and after 48 h to analyze structural changes in the retina and optic nerve. We analyzed glial cellular and molecular changes by histology and immunofluorescence and the impact of pretreatment with 4-phenylbutyric acid (4-PBA) in oligodendroglia survival. After 48 h hypoxia, we found no change in ganglion cell complex thickness and no loss of retinal ganglion cells. Despite this, there was significantly increased expression of CCAAT-enhancer-binding protein homologous protein (CHOP), a marker of endoplasmic reticulum stress, in the retina and optic nerve. In addition, hypoxia induced obvious increase of GFAP expression in the anterior optic nerve, where it co-localized with CHOP, and significant loss of Olig2+ oligodendrocytes. Pretreatment with 4-PBA, which has been shown to reduce endoplasmic reticulum stress, rescued total Olig2+ oligodendrocytes and increased the pool of mature (CC-1+) but not of immature (PDGFRa+) oligodendrocytes. Consistent with a selective vulnerability of the retina and optic nerve in hypoxia, the most striking changes in the 48 h murine model of hypoxia were in glial cells in the optic nerve, including increased CHOP expression in the astrocytes and loss of oligodendrocytes. Our data support a model where glial dysfunction is among the earliest events in systemic hypoxia - suggesting that glia may be a novel target in treatment of hypoxia.
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Hipoxia/complicaciones , Neuroglía/patología , Enfermedades del Nervio Óptico/diagnóstico , Nervio Óptico/patología , Animales , Supervivencia Celular , Modelos Animales de Enfermedad , Femenino , Hipoxia/diagnóstico , Etiquetado Corte-Fin in Situ , Ratones , Ratones Endogámicos C57BL , Enfermedades del Nervio Óptico/etiología , Células Ganglionares de la Retina/patología , Tomografía de Coherencia Óptica/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
Purpose: Increased endoplasmic reticulum (ER) stress is one of the earliest subcellular changes in neuro-ophthalmic diseases. In this study, we investigated the expression of key molecules in the ER stress pathways following nonarteritic anterior ischemic optic neuropathy (AION), the most common acute optic neuropathy in adults over 50, and assessed the impact of chemical chaperon 4-phenylbutyric acid (4-PBA) in vivo. Methods: We induced AION using photochemical thrombosis in adult mice and performed histologic analyses of key molecules in the ER stress pathway in the retina and optic nerve. We also assessed the effects of daily intraperitoneal injections of 4-PBA after AION. Results: In the retina at baseline, there was low proapoptotic transcriptional regulator C/EBP homologous protein (CHOP) and high prosurvival chaperon glucose-regulated protein 78 (GRP78) expression in retinal ganglion cells (RGCs). One day after AION, there was significantly increased CHOP and reduced GRP78 expressions in the ganglion cell layer. In the optic nerve at baseline, there was little CHOP and high GRP78 expression. One day after AION, there was significantly increased CHOP and no change in GRP78 expression. Treatment immediately after AION using daily intraperitoneal injection of chemical chaperone 4-PBA for 19 days significantly rescued Brn3A+ RGCs and Olig2+ optic nerve oligodendrocytes. Conclusions: We showed for the first time that acute AION resulted in increased ER stress and differential expression of ER stress markers CHOP and GRP78 in the retina and optic nerve. Rescue of RGCs and oligodendrocytes with 4-PBA provides support for ER stress reduction as possible treatment for AION.
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Estrés del Retículo Endoplásmico , Oligodendroglía/patología , Disco Óptico/patología , Neuropatía Óptica Isquémica/patología , Fenilbutiratos/farmacología , Células Ganglionares de la Retina/patología , Animales , Modelos Animales de Enfermedad , Chaperón BiP del Retículo Endoplásmico , Femenino , Regulación de la Expresión Génica , Proteínas de Choque Térmico/biosíntesis , Proteínas de Choque Térmico/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Chaperonas Moleculares , Oligodendroglía/metabolismo , Disco Óptico/metabolismo , Neuropatía Óptica Isquémica/tratamiento farmacológico , Neuropatía Óptica Isquémica/genética , Células Ganglionares de la Retina/metabolismo , Factor de Transcripción CHOP/biosíntesis , Factor de Transcripción CHOP/genéticaRESUMEN
Bone marrow-derived cells have been used in different animal models of neurological diseases. We investigated the therapeutic potential of mesenchymal stem cells (MSC) injected into the vitreous body in a model of optic nerve injury. Adult (3-5 months old) Lister Hooded rats underwent unilateral optic nerve crush followed by injection of MSC or the vehicle into the vitreous body. Before they were injected, MSC were labeled with a fluorescent dye or with superparamagnetic iron oxide nanoparticles, which allowed us to track the cells in vivo by magnetic resonance imaging. Sixteen and 28 days after injury, the survival of retinal ganglion cells was evaluated by assessing the number of Tuj1- or Brn3a-positive cells in flat-mounted retinas, and optic nerve regeneration was investigated after anterograde labeling of the optic axons with cholera toxin B conjugated to Alexa 488. Transplanted MSC remained in the vitreous body and were found in the eye for several weeks. Cell therapy significantly increased the number of Tuj1- and Brn3a-positive cells in the retina and the number of axons distal to the crush site at 16 and 28 days after optic nerve crush, although the RGC number decreased over time. MSC therapy was associated with an increase in the FGF-2 expression in the retinal ganglion cells layer, suggesting a beneficial outcome mediated by trophic factors. Interleukin-1ß expression was also increased by MSC transplantation. In summary, MSC protected RGC and stimulated axon regeneration after optic nerve crush. The long period when the transplanted cells remained in the eye may account for the effect observed. However, further studies are needed to overcome eventually undesirable consequences of MSC transplantation and to potentiate the beneficial ones in order to sustain the neuroprotective effect overtime.