ISOLATION, CLONING AND CHARACTERIZATION OF A PUTATIVE TYPE-1 ASTROCYTE CELL-LINE

We have established a permanent cell line (1H91) of putative type-1 as trocyte precursor cells that were clonally derived from a single cell isolated from E16 mouse cerebellum. Epidermal growth factor (EGF) and transforming growth factor (TGF alpha) are strong mitogens for 1H91 ce lls (ED(50) of 9.02 + 1.74 ng/ml and 15.98 +/- 2.34 ng/ml, respectivel y), while basic fibroblast growth factor (bFGF) is only weakly mitogen ic and platelet derived growth factor (PDGF) has no mitogenic activity . In the proliferative state, the 1H91 cells are immunohistochemically positive for nestin and vimentin, and negative for A2B5, CNPase, neur ofilament (NF), and neuron specific enolase (NSE). The majority of EGF -treated 1H91 cells are not immunoreactive for glial acid fibrillary p rotein (GFAP). In the presence of 5 ng/ml bFGF, 1H91 cells become non- mitotic and develop a morphology consistent with a fibrous astrocyte. In contrast to the proliferating cultures, the bFGF treated cultures w ere strongly immunoreactive for GFAP, only mildly immunoreactive for n estin and vimentin, and negative for A2B5, CNPase, NF, and NSE. Type-1 astrocytes are known to proliferate in response to EGF, and are immun ohistochemically GFAP positive, A2B5 negative, and CNPase negative [38 ]. However, type-1 astrocytes only develop a fibrous morphology during the process of reactive gliosis [31]. Since EGF is a strong mitogen f or 1H91 cells, and these cells may be differentiated into GFAP positiv e, A2B5 negative, CNPase negative astrocytes, we conclude that 1H91 ce lls conform to a type-1 astrocyte precursor phenotype. In addition, th e fibrous morphology of the bFGF treated 1H91 cells suggests that thes e cells follow the process of reactive gliosis. Therefore, the 1H91 cl onal cell line may provide an in vitro model for studying the underlyi ng cellular mechanisms of the type-1 astrocyte in reactive gliosis.