Analysis of NO synthase expression in neuronal, astroglial and fibroblast-like derivatives differentiating from PCC7-Mz1 embryonic carcinoma cells

We studied the expression of the NO synthase isoforms in an in vitro model of neural development using RT-PCR, Western blot and immunohistochemistry. Murine PCC7-Mz1 cells (Jostock et al,, Eur. J. Cell Biol, 76, 63-76, 1998) differentiate in the presence of all-trans retinoic acid and dibutyryl cAMP along the neural pathway into neuron-like, fibroblast-like and astroglia-l ike cells. Undifferentiated cells showed immunofluorescent staining for neu ronal-type NOS I and endothelial-type NOS III. This expression pattern was retained in those cells differentiating into neurofilament- and tau protein -positive neuronal cells. Thymocyte alloantigen (Thyl.2/CD 90.2)-positive f ibroblasts, appearing around day 3, and glial fibrillary acidic protein (GF AP)-positive astroglial cells, appearing after day 6 of differentiation, st ained negative for any NOS isoform, Starting at day 6 of differentiation, e xpression of inducible-type NOS II could be stimulated with cytokines in a subset of cells, which may represent activated astrocytes, NOS II was alway s undetectable in non-induced cultures. These data indicate that the abilit y of stem cells to express NOS I and NOS III is only retained when the cell s differentiate along the neuronal lineage, while a small subpopulation of cells acquires the ability to express NOS II in response to cytokines.