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Even though astrocytes are critical for both normal brain functions and the development and progression of neuropathological states, including neuroinflammation associated with neurodegenerative diseases, the mechanisms controlling gene expression during astrocyte differentiation are poorly understood. Thus far, several signaling pathways were shown to regulate astrocyte differentiation, including JAK‐STAT, bone morphogenic protein‐2/Smads, and Notch. More recently, a family of nuclear factor‐1 (NFI‐A, ‐B, ‐C, and ‐X) was implicated in the regulation of vertebral neocortex development, with NFI‐A and ‐B controlling the onset of gliogenesis. Here, we developed an in vitro model of differentiation of stem cells towards neural progenitors (NP) and subsequently astrocytes. The transition from stem cells to progenitors was accompanied by an expected change in the expression profile of markers, including Sox‐2, Musashi‐1, and Oct4. Subsequently, generated astrocytes were characterized by proper morphology, increased glutamate uptake, and marker gene expression. We used this in vitro differentiation model to study the expression and functions of NFIs. Interestingly, stem cells expressed only background levels of NFIs, while differentiation to NP activated the expression of NFI‐A. More importantly, NFI‐X expression was induced during the later stages of differentiation towards astrocytes. In addition, NFI‐X and ‐C were required for the expression of glial fibrillary acidic protein and secreted protein acidic and rich in cystein‐like protein 1, which are the markers of astrocytes at the later stages of differentiation. We conclude that an expression program of NFIs is executed during the differentiation of astrocytes, with NFI‐X and ‐C controlling the expression of astrocytic markers at late stages of differentiation. S tem  C ells  2009;27:1173–1181

Nuclear Factor I Isoforms Regulate Gene Expression During the Differentiation of Human Neural Progenitors to Astrocytes