To characterize the properties of adult neural stem cells (NSCs), we generated and analyzed Sox2-GFP transgenic mice. Sox2-GFP cells in the subgranular zone (SGZ) express markers specific for progenitors, but they represent two morphologically distinct populations that differ in proliferation levels. Lentivirus- and retrovirus-mediated fate-tracing studies showed that Sox2+ cells in the SGZ have potential to give rise to neurons and astrocytes, revealing their multipotency at the population as well as at a single-cell level. A subpopulation of Sox2+ cells gives rise to cells that retain Sox2, highlighting Sox2+ cells as a primary source for adult NSCs. In response to mitotic signals, increased proliferation of Sox2+ cells is coupled with the generation of Sox2+ NSCs as well as neuronal precursors. An asymmetric contribution of Sox2+ NSCs may play an important role in maintaining the constant size of the NSC pool and producing newly born neurons during adult neurogenesis.

The dentate gyrus of the hippocampus is one of the few areas of the adult brain that undergoes neurogenesis. In the present study, cells capable of proliferation and neurogenesis were isolated and cultured from the adult rat hippocampus. In defined medium containing basic fibroblast growth factor (FGF-2), cells can survive, proliferate, and express neuronal and glial markers. Cells have been maintained in culture for 1 year through multiple passages. These cultured adult cells were labeled in vitro with bromodeoxyuridine and adenovirus expressing ,i-galactosidase and were transplanted to the adult rat hippocampus. Surviving cells were evident through 3 months postimplantation with no evidence of tumor formation. Within 2 months postgrafting, labeled cells were found in the dentate gyrus, where they differentiated into neurons only in the intact region of the granule cell layer. Our results indicate that FGF-2 responsive progenitors can be isolated from the adult hippocampus and that these cells retain the capacity to generate mature neurons when grafted into the adult rat brain.Most neurons in the adult central nervous system are terminally differentiated, exist through the life of the organism, and are not replaced when they die. However, evidence exists that small populations of neurons continue to be born in the adult ventricular zone (1-3), olfactory system (4-6), and hippocampus (7-10). In the adult hippocampus, newly born neurons originate from putative stem cells that exist in the subgranular zone of the dentate gyrus. Progeny of these putative stem cells differentiate into neurons in the granule cell layer within a month of the cells' birth, and this late neurogenesis continues throughout the adult life of the rodent (11-15). Paralleling these in vivo findings, in vitro studies have shown that the precursor cells isolated from adult mouse ventricular zone and forebrain have the capacity for in vitro neurogenesis when stimulated with epidermal growth factor (16) or basic fibroblast growth factor (FGF-2) (17), respectively. FGF-2 is a potent mitogen for fetal cells isolated from different areas of the brain (18)(19)(20)(21)(22). This proliferative property of FGF-2 has allowed the isolation and culturing of fetal hippocampal cells for over a year through multiple passages (21), some of which expressed neuronal phenotypes in vitro. These studies have raised the question whether the FGF-2 responsive cells can be isolated and cultured from the adult brain, particularly in the hippocampus, where neurogenesis occurs even in adulthood. In the present study we report that a population of FGF-2 responsive progenitor cells can be isolated and cultured from the adult rat hippocampus. Cells in culture express precursor, glial, and neuronal cell markers. Upon implantation into the adult rat hippocampus, cells migrate and differentiate into mature neurons or glia, depending on the terminal site of migration. Such a progenitor population will be valuable forThe publication costs of this article were defrayed in ...

The finding of neurogenesis in the adult brain led to the discovery of adult neural stem cells. TLX was initially identified as an orphan nuclear receptor expressed in vertebrate forebrains and is highly expressed in the adult brain. The brains of TLX-null mice have been reported to have no obvious defects during embryogenesis; however, mature mice suffer from retinopathies, severe limbic defects, aggressiveness, reduced copulation and progressively violent behaviour. Here we show that TLX maintains adult neural stem cells in an undifferentiated, proliferative state. We show that TLX-expressing cells isolated by fluorescence-activated cell sorting (FACS) from adult brains can proliferate, self-renew and differentiate into all neural cell types in vitro. By contrast, TLX-null cells isolated from adult mutant brains fail to proliferate. Reintroducing TLX into FACS-sorted TLX-null cells rescues their ability to proliferate and to self-renew. In vivo, TLX mutant mice show a loss of cell proliferation and reduced labelling of nestin in neurogenic areas in the adult brain. TLX can silence glia-specific expression of the astrocyte marker GFAP in neural stem cells, suggesting that transcriptional repression may be crucial in maintaining the undifferentiated state of these cells.

The adult rat spinal cord contains cells that can proliferate and differentiate into astrocytes and oligodendroglia in situ. Using clonal and subclonal analyses we demonstrate that, in contrast to progenitors isolated from the adult mouse spinal cord with a combination of growth factors, progenitors isolated from the adult rat spinal cord using basic fibroblast growth factor alone display stem cell properties as defined by their multipotentiality and self-renewal. Clonal cultures derived from single founder cells generate neurons, astrocytes, and oligodendrocytes, confirming the multipotent nature of the parent cell. Subcloning analysis showed that after serial passaging, recloning, and expansion, these cells retained multipotentiality, indicating that they are self-renewing. Transplantation of an in vitro-expanded clonal population of cells into the adult rat spinal cord resulted in their differentiation into glial cells only. However, after heterotopic transplantation into the hippocampus, transplanted cells that integrated in the granular cell layer differentiated into cells characteristic of this region, whereas engraftment into other hippocampal regions resulted in the differentiation of cells with astroglial and oligodendroglial phenotypes. The data indicate that clonally expanded, multipotent adult progenitor cells from a non-neurogenic region are not lineage-restricted to their developmental origin but can generate region-specific neurons in vivo when exposed to the appropriate environmental cues.Key words : spinal cord; stem cells; FGF; transplantation; neuroplasticity; adult Most neurogenesis in the mammalian CNS is believed to end in the period just after birth (Nornes and Das, 1974;Altman and Bayer, 1984). However, neurogenesis continues in different regions of the brain of various adult mammalian species (Kaplan and Hinds, 1980;Bayer et al., 1982; Alvarez-Buylla, 1993, 1994;Luskin, 1993). The spinal cord, like most structures of the mammalian brain, belongs to the class of nonrenewable epithelium (Rakic, 1985). However, a small number of cells that line the central canal (Adrian and Walker, 1962;Johansson et al., 1999) remain mitotic. We have demonstrated recently that the adult rat spinal cord contains large numbers of dividing cells in vivo that give rise to glia but not neurons (Horner et al., 2000).Multipotent stem cells that respond to epidermal growth factor (EGF) or basic fibroblast growth factor (FGF-2) have been isolated from both neurogenic (Morshead et al., 1994;Palmer et al., 1997) and non-neurogenic regions of the adult mammalian CNS (Temple and Alvarez-Buylla, 1999). Recent studies indicate that embryonic day 14 mouse striatum or adult subventricular zone (SVZ) contains multipotent stem-like cells that are controlled by FGF-2 or EGF in a regulatable manner (Ciccolini and Svendsen, 1998;Gritti et al., 1999). However, this is not true of adult CNS stem cells from all regions, because a combination of EGF and FGF-2 was necessary to isolate stem cells from the adult mouse spinal cord ,...

No abstract