Cellular reprogramming is a new and rapidly emerging field in which somatic cells can be turned into pluripotent stem cells or other somatic cell types simply by the expression of specific combinations of genes. By viral expression of neural fate determinants, it is possible to directly reprogram mouse and human fibroblasts into functional neurons, also known as induced neurons. The resulting cells are nonproliferating and present an alternative to induced pluripotent stem cells for obtaining patient-and disease-specific neurons to be used for disease modeling and for development of cell therapy. In addition, because the cells do not pass a stem cell intermediate, direct neural conversion has the potential to be performed in vivo. In this study, we show that transplanted human fibroblasts and human astrocytes, which are engineered to express inducible forms of neural reprogramming genes, convert into neurons when reprogramming genes are activated after transplantation. Using a transgenic mouse model to specifically direct expression of reprogramming genes to parenchymal astrocytes residing in the striatum, we also show that endogenous mouse astrocytes can be directly converted into neural nuclei (NeuN)-expressing neurons in situ. Taken together, our data provide proof of principle that direct neural conversion can take place in the adult rodent brain when using transplanted human cells or endogenous mouse cells as a starting cell for neural conversion.T he ability to reprogram somatic cells to pluripotent stem cells or other somatic cell types by expressing key combinations of genes has opened up new possibilities for disease modeling and cell therapy (1, 2). Using this technique, it is possible to directly reprogram mouse and human fibroblasts into functional neurons, also known as induced neurons (iNs), using viral delivery of the three neural conversion factors achaete-scute complex-like 1 (Ascl1), brain-2 (Brn2a), and myelin transcription factor-like 1 (Myt1l) (ABM) (3, 4). A growing number of studies now show that by altering the combination of genes used for reprogramming, different subtypes of neurons are obtained (3,5,6). Importantly, the resulting cells are nonproliferating, which makes them an interesting alternative to induced pluripotent stem cells as a source of patient-specific neurons for cell replacement therapy, once efficient grafting strategies for these cells are developed.The adult brain has a very limited inherent capacity for repair, and new neurons are only formed in two discrete regions: the subventricular zone of the lateral ventricles, which generates neurons migrating to the olfactory bulb, and the hippocampus (7,8). Experimental studies have shown that these endogenous progenitors can also be recruited to generate new neurons in other regions as well in response to injury (9-11). However, the number of new neurons is very low, their migration is hard to control, and the therapeutic implications are unclear. Several cell types residing outside the neurogenic niche, such as parenchymal...

Age-associated neurological diseases represent a profound challenge in biomedical research as we are still struggling to understand the interface between the aging process and the manifestation of disease. Various pathologies in the elderly do not directly result from genetic mutations, toxins, or infectious agents but are primarily driven by the many manifestations of biological aging. Therefore, the generation of appropriate model systems to study human aging in the nervous system demands new concepts that lie beyond transgenic and drug-induced models. Although access to viable human brain specimens is limited and induced pluripotent stem cell models face limitations due to reprogramming-associated cellular rejuvenation, the direct conversion of somatic cells into induced neurons allows for the generation of human neurons that capture many aspects of aging. Here, we review advances in exploring age-associated neurodegenerative diseases using human cell reprogramming models, and we discuss general concepts, promises, and limitations of the field.

Direct conversion of human fibroblasts into mature and functional neurons, termed induced neurons (iNs), was achieved for the first time 6 years ago. This technology offers a promising shortcut for obtaining patient‐ and disease‐specific neurons for disease modeling, drug screening, and other biomedical applications. However, fibroblasts from adult donors do not reprogram as easily as fetal donors, and no current reprogramming approach is sufficiently efficient to allow the use of this technology using patient‐derived material for large‐scale applications. Here, we investigate the difference in reprogramming requirements between fetal and adult human fibroblasts and identify REST as a major reprogramming barrier in adult fibroblasts. Via functional experiments where we overexpress and knockdown the REST‐controlled neuron‐specific microRNAs miR‐9 and miR‐124, we show that the effect of REST inhibition is only partially mediated via microRNA up‐regulation. Transcriptional analysis confirmed that REST knockdown activates an overlapping subset of neuronal genes as microRNA overexpression and also a distinct set of neuronal genes that are not activated via microRNA overexpression. Based on this, we developed an optimized one‐step method to efficiently reprogram dermal fibroblasts from elderly individuals using a single‐vector system and demonstrate that it is possible to obtain iNs of high yield and purity from aged individuals with a range of familial and sporadic neurodegenerative disorders including Parkinson's, Huntington's, as well as Alzheimer's disease.

SummaryThe possibility of directly converting non-neuronal cells into neurons in situ in the brain would open therapeutic avenues aimed at repairing the brain after injury or degenerative disease. We have developed an adeno-associated virus (AAV)-based reporter system that allows selective GFP labeling of reprogrammed neurons. In this system, GFP is turned on only in reprogrammed neurons where it is stable and maintained for long time periods, allowing for histological and functional characterization of mature neurons. When combined with a modified rabies virus-based trans-synaptic tracing methodology, the system allows mapping of 3D circuitry integration into local and distal brain regions and shows that the newly reprogrammed neurons are integrated into host brain.

Neurons are the longest-lived cells in our bodies and lack DNA replication, which makes them reliant on a limited repertoire of DNA repair mechanisms to maintain genome fidelity. These repair mechanisms decline with age, but we have limited knowledge of how genome instability emerges and what strategies neurons and other long-lived cells may have evolved to protect their genomes over the human life span. A targeted sequencing approach in human embryonic stem cell–induced neurons shows that, in neurons, DNA repair is enriched at well-defined hotspots that protect essential genes. These hotspots are enriched with histone H2A isoforms and RNA binding proteins and are associated with evolutionarily conserved elements of the human genome. These findings provide a basis for understanding genome integrity as it relates to aging and disease in the nervous system.