RESUMEN
The translational gap to treatments based on gene therapy has been reduced in recent years because of improvements in gene editing tools, such as the CRISPR/Cas9 system and its variations. This has allowed the development of more precise therapies for neurodegenerative diseases, where access is privileged. As a result, engineering of complexes that can access the central nervous system (CNS) with the least potential inconvenience is fundamental. In this review article, we describe current alternatives to generate systems based on CRISPR/Cas9 that can cross the blood-brain barrier (BBB) and may be used further clinically to improve treatment for neurodegeneration in Parkinson's and Alzheimer's disease (AD).
RESUMEN
Cell therapy in animal models of Parkinson's disease (PD) is effective after intrastriatal grafting of dopamine (DA) neurons, whereas intranigral transplantation of dopaminergic cells does not cause consistent behavioral recovery. One strategy to promote axonal growth of dopaminergic neurons from the substantia nigra (SN) to the striatum is degradation of inhibitory components such as chondroitin sulphate proteoglycans (CSPG). An alternative is the guidance of DA axons by chemotropic agents. Semaphorins 3A and 3C enhance axonal growth of embryonic stem (ES) cell-derived dopaminergic neurons in vitro, while Semaphorin 3C also attracts them. We asked whether intranigral transplantation of DA neurons, combined with either degradation of CSPG or with grafts of Semaphorin 3-expressing cells, towards the striatum, is effective in establishing a new nigrostriatal dopaminergic pathway in rats with unilateral depletion of DA neurons. We found depolarization-induced DA release in dorsal striatum, DA axonal projections from SN to striatum, and concomitant behavioral improvement in Semaphorin 3-treated animals. These effects were absent in animals that received intranigral transplants combined with Chondroitinase ABC treatment, although partial degradation of CSPG was observed. These results are evidence that Semaphorin 3-directed long-distance axonal growth of dopaminergic neurons, resulting in behavioral improvement, is possible in adult diseased brains.
Asunto(s)
Axones/metabolismo , Tratamiento Basado en Trasplante de Células y Tejidos , Neuronas Dopaminérgicas/metabolismo , Neuronas Dopaminérgicas/trasplante , Trastornos Parkinsonianos/metabolismo , Trastornos Parkinsonianos/terapia , Semaforinas/metabolismo , Animales , Diferenciación Celular , Línea Celular , Cuerpo Estriado/metabolismo , Células Madre Embrionarias/citología , Células Madre Embrionarias/metabolismo , Femenino , Células HEK293/metabolismo , Células HEK293/trasplante , Humanos , Ratones , Oxidopamina/metabolismo , Trastornos Parkinsonianos/fisiopatología , Ratas , Prueba de Desempeño de Rotación con Aceleración Constante , Semaforinas/genética , Sustancia Negra , Transmisión Sináptica , TransfecciónRESUMEN
Class 3 Semaphorins are a subfamily of chemotropic molecules implicated in the projection of dopaminergic neurons from the ventral mesencephalon and in the formation of the nigrostriatal pathway (NSP) during embryonic development. In humans, loss of mesencephalic dopaminergic neurons leads to Parkinson's disease (PD). Cell replacement therapy with dopaminergic neurons generated from embryonic stem cells (ES-TH(+)) is being actively explored in models of PD. Among several requisites for this approach to work are adequate reconstruction of the NSP and correct innervation of normal striatal targets by dopaminergic axons. In this work, we characterized the response of ES-TH(+) neurons to semaphorins 3A, 3C, and 3F and compared it with that of tyrosine hidroxylase-positive neurons (TH(+)) obtained from embryonic ventral mesencephalon (VM-TH(+)). We observed that similar proportions of ES-TH(+) and VM-TH(+) neurons express semaphorin receptors neuropilins 1 and 2. Furthermore, the axons of both populations responded very similarly to semaphorin exposure: semaphorin 3A increased axon length, and semaphorin 3C attracted axons and increased their length. These effects were mediated by neuropilins, insofar as addition of blocking antibodies against these proteins reduced the effects on axonal growth and attraction, and only TH(+) axons expressing neuropilins responded to the semaphorins analyzed. The observations reported here show phenotypic similarities between VM-TH(+) and ES-TH(+) neurons and suggest that semaphorins 3A and 3C could be employed to guide axons of grafted ES-TH(+) in therapeutic protocols for PD.