Effects of the post-spinal cord injury microenvironment on the differentiation capacity of human neural stem cells derived from induced pluripotent stem cells

This work was supported by TERCEL and CIBERNED funds from the Instituto de Salud Carlos III of Spain, and FEDER funds from the EC.Spinal cord injury (SCI) causes loss of neural functions below the level of the lesion due to interruption of spinal pathways and secondary neurodegenerative processes. The transplant of neural stem cells (NSCs) is a promising approach for the repair of SCI. Reprogramming of adult somatic cells into induced pluripotent stem cells (iPSCs) is expected to provide an autologous source of iPSC-derived NSCs, avoiding the immune response as well as ethical issues. However, there is still limited information on the behavior and differentiation pattern of transplanted iPSC-derived NSCs within the damaged spinal cord. We transplanted iPSC-derived NSCs, obtained from adult human somatic cells, into rats at 0 or 7 days after SCI, and evaluated motor-evoked potentials and locomotion of the animals. We histologically analyzed engraftment, proliferation, and differentiation of the iPSC-derived NSCs and the spared tissue in the spinal cords at 7, 21, and 63 days posttransplant. Both transplanted groups showed a late decline in functional recovery compared to vehicle-injected groups. Histological analysis showed proliferation of transplanted cells within the tissue and that cells formed a mass. At the final time point, most grafted cells differentiated to neural and astroglial lineages, but not into oligodendrocytes, while some grafted cells remained undifferentiated and proliferative. The proinflammatory tissue microenviroment of the injured spinal cord induced proliferation of the grafted cells and, therefore, there are possible risks associated with iPSC-derived NSC transplantation. New approaches are needed to promote and guide cell differentiation, as well as reduce their tumorigenicity once the cells are transplanted at the lesion site

Aberrant perineuronal nets alter spinal circuits, impair motor function, and increase plasticity

Altres ajuts: acords transformatius de la UABPerineuronal nets (PNNs) are a specialized extracellular matrix that have been extensively studied in the brain. Cortical PNNs are implicated in synaptic stabilization, plasticity inhibition, neuroprotection, and ionic buffering. However, the role of spinal PNNs, mainly found around motoneurons, is still unclear. Thus, the goal of this study is to elucidate the role of spinal PNNs on motor function and plasticity in both intact and spinal cord injured mice. We used transgenic mice lacking the cartilage link protein 1 (Crtl1 KO mice), which is implicated in PNN assembly. Crtl1 KO mice showed disorganized PNNs with an altered proportion of their components in both motor cortex and spinal cord. Behavioral and electrophysiological tests revealed motor impairments and hyperexcitability of spinal reflexes in Crtl1 KO compared to WT mice. These functional outcomes were accompanied by an increase in excitatory synapses around spinal motoneurons. Moreover, following spinal lesions of the corticospinal tract, Crtl1 KO mice showed increased contralateral sprouting compared to WT mice. Altogether, the lack of Crtl1 generates aberrant PNNs that alter excitatory synapses and change the physiological properties of motoneurons, overall altering spinal circuits and producing motor impairment. This disorganization generates a permissive scenario for contralateral axons to sprout after injury

Terapia celular para lesiones que afectan a la médula espinal

La médula espinal puede verse dañada por patologías no traumáticas como tumores, infecciones, enfermedades autoinmunes y enfermedades degenerativas, y por lesiones traumáticas, tanto por acción directa como indirecta. Las lesiones traumáticas que dañan la medula espinal constituyen unas de las mayores causa de discapacidad física persistente al largo de toda la vida del paciente. A día de hoy, no hay tratamiento eficaz para este tipo de dolencias y las aplicaciones terapéuticas se basan en sacar el máximo potencial a las funciones conservadas. En los esfuerzos por encontrar tratamientos para éstas lesiones, el trasplante de diferentes tipos celulares constituye uno de los pilares de la investigación experimental. Dentro de esta estrategia dos de las poblaciones de células más usadas son las células de la glia envolvente que se encuentran en el bulbo olfatorio (OEC) y las células estromales mesenquimales (MSC). Ambos tipos celulares han demostrado mejorar la recuperación funcional y proteger parte del tejido dañado después de ser trasplantadas en modelos animales de lesiones de médula espinal. A pesar de ello, los mecanismos que subyacen a su acción han sido poco estudiados. Por este motivo, los trabajos incluidos en la presente tesis buscan la respuesta a un aspecto desconocido de estas terapias, como es la comparación del potencial terapéutico de las OEC y las MSC para las lesiones de médula espinal por contusión, tanto en su efecto como en su mecanismo. Los datos mostraron como, a pesar de que ambas células promovían una protección de tejido, las mejoras funcionales encontradas fueron limitadas y tan solo el trasplante de MSC en tiempos inmediatos a la lesión resultó en una mejora significativa. La poca supervivencia de las células dentro de la médula espinal lesionada podría ser uno de los motivos que explicaría la falta de un efecto beneficioso a nivel funcional. Un análisis de los cambios génicos inducidos por el trasplante reveló que la muerte de las células trasplantadas podría ser consecuencia de un rechazo inmunológico. La administración de un fármaco inmunosupresor a lo largo del seguimiento de los animales confirmó esta hipótesis, extendiendo la supervivencia de las células, pero con mejoras funcionales aun escasas, sugiriendo que la poca supervivencia de las células no es la única causa de la limitada mejora funcional encontrada. Por otro lado, el estudio génico también mostró como ambas células inducen la protección del tejido potenciando vías de reparación tisular. Mientras que las MSC modulaban estos procesos de reparación, aumentándolos en los tiempos tempranos después de la lesión y normalizándolos más tarde, las OEC actuaban tan solo en los tiempos tempranos, con reducidos efectos en los tiempos tardíos después de la lesión. Además, se ha indagado en la utilidad de las MSC como coadyuvante para el tratamiento de las lesiones de médula espinal por avulsión de raíz ventral. Este trabajo demostró como la presencia de las MSC en la médula espinal después de una avulsión mejoraba la supervivencia de motoneuronas y, en combinación con la reparación quirúrgica, aumentaba la velocidad de regeneración axonal y de reinervación muscular. En conclusión, la terapia celular con OEC o con MSC para lesiones de médula espinal puede ayudar al tratamiento de estas patologías, aunque por si solas las mejoras obtenidas son insatisfactorias. Una mejor comprensión de los mecanismos por los cuales las células trasplantadas ejercen su acción protectora permitiría optimizar su uso y establecer tratamientos combinados que respondan a varios objetivos terapéuticos.The spinal cord can be affected by non traumatic pathologies such as, tumors, infections, auto-immune diseases and degenerative diseases, and by both direct and indirect traumatic injuries. Traumatic spinal cord injuries leads to a physical disabilities persistent during the patient's lifetime. Nowadays, there are not efficient treatment for spinal cord injuries (SCI). One of the most studied strategies to treat SCI diseases is based in cell transplantation. Several cell types have been evaluated for SCI treatment; among them adult Mesenchymal Stromal Cells (MSC) derived from the bone marrow and Olfactory Ensheathing Cells (OEC) from the olfactory bulb have received considerable attention, considering the positive results of several experimental studies regarding to tissue protection and functional outcome improvement. Although these beneficial effects of OEC and MSC transplantation after SCI, the mechanisms under the action of both type of cells are still little known. In the present work we wanted to address the comparison between OEC and MSC transplantation as a treatment to SCI contusion in their effects and in their mechanisms. Our results shown that both OEC and MSC transplant provided tissue protection, but only the MSC treatment in the acute time after SCI induced, limited but significantly, better functional recovery. One of the possible reason for the poor functional results could be due to the limited survival of the engrafted cells into injured spinal cord. The analysis of the gene expression after cell transplantation reveled a high up-regulation of genes associated to immunological reaction. The administration of an immunosuppressant drug during the follow up of the animals after transplantation allowed long-time survival of the graft, however the functional outcomes remained poor, suggesting that the short survival of the cells into damaged tissue was not the unique reason for the limited recovery. On the other hand, the gene expression study also shown how both type of cells cause the tissue protection by increasing the action of the tissue repair process. While MSC modulated the tissue repair by potentiating it in the early phase and resolving it during late times, the OEC acted only in the early time after transplantation. Moreover, we also investigated the MSC transplantation as a co-treatment to ventral root avulsion injuries. Our data shown that MSC can rescue the motoneurons from the death induced by the injury, and the combination of MSC transplant with surgical repair improved the axonal regeneration speed and muscle reinnervation. In conclusion, cell therapies using OEC or MSC to treat SCI can provided beneficial effects. A better understanding of the mechanisms by which these cells exert their protective action would optimize their use and would allow set up a combinatory therapies to respond to multiple therapeutic targets