Transcription factor combinations that define human astrocyte identity encode significant variation of maturity and function.

Increasing evidence indicates that cellular identity can be reduced to the distinct gene regulatory networks controlled by transcription factors (TFs). However, redundancy exists in these states as different combinations of TFs can induce broadly similar cell types. We previously demonstrated that by overcoming gene silencing, it is possible to deterministically reprogram human pluripotent stem cells directly into cell types of various lineages. In the present study we leverage the consistency and precision of our approach to explore four different TF combinations encoding astrocyte identity, based on previously published reports. Analysis of the resulting induced astrocytes (iAs) demonstrated that all four cassettes generate cells with the typical morphology of in vitro astrocytes, which expressed astrocyte-specific markers. The transcriptional profiles of all four iAs clustered tightly together and displayed similarities with mature human astrocytes, although maturity levels differed between cells. Importantly, we found that the TF cassettes induced iAs with distinct differences with regards to their cytokine response and calcium signaling. In vivo transplantation of selected iAs into immunocompromised rat brains demonstrated long term stability and integration. In conclusion, all four TF combinations were able to induce stable astrocyte-like cells that were morphologically similar but showed subtle differences with respect to their transcriptome. These subtle differences translated into distinct differences with regards to cell function, that could be related to maturation state and/or regional identity of the resulting cells. This insight opens an opportunity to precision-engineer cells to meet functional requirements, for example, in the context of therapeutic cell transplantation.

Electrophysiological In Vitro Study of Long-Range Signal Transmission by Astrocytic Networks.

Astrocytes are diverse brain cells that form large networks communicating via gap junctions and chemical transmitters. Despite recent advances, the functions of astrocytic networks in information processing in the brain are not fully understood. In culture, brain slices, and in vivo, astrocytes, and neurons grow in tight association, making it challenging to establish whether signals that spread within astrocytic networks communicate with neuronal groups at distant sites, or whether astrocytes solely respond to their local environments. A multi-electrode array (MEA)-based device called AstroMEA is designed to separate neuronal and astrocytic networks, thus allowing to study the transfer of chemical and/or electrical signals transmitted via astrocytic networks capable of changing neuronal electrical behavior. AstroMEA demonstrates that cortical astrocytic networks can induce a significant upregulation in the firing frequency of neurons in response to a theta-burst charge-balanced biphasic current stimulation (5 pulses of 100 Hz × 10 with 200 ms intervals, 2 s total duration) of a separate neuronal-astrocytic group in the absence of direct neuronal contact. This result corroborates the view of astrocytic networks as a parallel mechanism of signal transmission in the brain that is separate from the neuronal connectome. Translationally, it highlights the importance of astrocytic network protection as a treatment target.

Therapeutic Potential of Astrocyte Transplantation.

Cell transplantation is an attractive treatment strategy for a variety of brain disorders, as it promises to replenish lost functions and rejuvenate the brain. In particular, transplantation of astrocytes has come into light recently as a therapy for amyotrophic lateral sclerosis (ALS); moreover, grafting of astrocytes also showed positive results in models of other conditions ranging from neurodegenerative diseases of older age to traumatic injury and stroke. Despite clear differences in etiology, disorders such as ALS, Parkinson's, Alzheimer's, and Huntington's diseases, as well as traumatic injury and stroke, converge on a number of underlying astrocytic abnormalities, which include inflammatory changes, mitochondrial damage, calcium signaling disturbance, hemichannel opening, and loss of glutamate transporters. In this review, we examine these convergent pathways leading to astrocyte dysfunction, and explore the existing evidence for a therapeutic potential of transplantation of healthy astrocytes in various models. Existing literature presents a wide variety of methods to generate astrocytes, or relevant precursor cells, for subsequent transplantation, while described outcomes of this type of treatment also differ between studies. We take technical differences between methodologies into account to understand the variability of therapeutic benefits, or lack thereof, at a deeper level. We conclude by discussing some key requirements of an astrocyte graft that would be most suitable for clinical applications.