RFP Lentivirus

Astrocytes are a highly heterogeneous population of glial cells that are important regulators of brain development and homeostasis. The heterogeneity of the astrocyte population underlies its functional diversity. In addition to the typical mammalian astrocyte architecture, the human cerebral cortex exhibits a radial distribution of interlaminar astrocytes in the supragranular layer. These primate-specific interlaminar astrocytes are located in the superficial layers and project long processes that traverse multiple layers of the cerebral cortex. However, their functional properties and their role in regulating neural circuits remain unclear due to the lack of available experimental models. Here, researchers modeled human interlaminar astrocytes in humanized glial chimeric mice by transplanting astrocytes differentiated from human induced pluripotent stem cells into the mouse cortex. This model provides a new platform for understanding neuron-glial interactions and their alterations in neurological diseases.

Here, researchers differentiated human induced pluripotent stem cells (hiPSCs) into neural progenitor cells (NPCs) and subsequently into astrocytes via a xeno-free, chemically defined approach (Figure 1a). To visualize hiPSC-astroglia in chimeric mouse brains, researchers generated RFP-expressing astrocytes by transducing NPCs with lentivirus expressing RFP in astrocyte differentiation medium (Figure 1a). Prior to implantation, RFP-expressing hiPSC-astroglia showed robust expression of canonical markers, including aldehyde dehydrogenase 1 family member L1 (ALDH1L1), glial fibrillary acidic protein (GFAP), and S100 calcium-binding protein B (S100B) (Figure 1b). RFP-expressing hiPSC-derived immature astrocytes were transplanted into the cortex of postnatal day 1 (P1) rag1−/− immunodeficient mouse brains. Two-site injection resulted in the widespread distribution of RFP-expressing hiPSC-astrocytes in the frontal cortex. Although thick bands of cell bodies were found in the superficial layers of the cortex at the site where the hiPSC-astroglia were likely engrafted, more distally, the cells were primarily confined to layer 1. Furthermore, although cells with interlaminar processes were found in the superficial layers, protoplasmic astrocytes were observed in the deeper layers of the putative injection site.

Figure 1. Generation of hiPSC-astrocytes. (Padmashri R, et al., 2021)