A novel human astrocyte cell line (A735) with astrocyte-specific neurotransmitter function

Studies of brain cell function and physiology are hampered by the limited a vailability of immortal human brain-derived cell lines, as a result of the technical difficulties encountered in establishing immortal human cells in culture. In this study, we demonstrate the application of recombinant DNA v ectors expressing SV40 T antigen for the development of immortal human cell cultures, with morphological, growth, and functional properties of astrocy tes. Primary human astrocytes were transfected with the SV40 T antigen expressio n vectors, pSV3neo or p735.6, and cultures were established with an extende d lifespan. One of these cultures gave rise to an immortal cell line, desig nated A735. All the human SV40-derived lines retained morphological feature s and growth properties of type 1 astrocytes. Immunohistochemical studies a nd Western blot analysis of the intermediate filament proteins and glutamin e synthetase demonstrated a differentiated but immature astrocyte phenotype . Transport of gamma-amino butyric acid and glutamate were examined and fou nd to be by a glial-specific mechanism, consistent with the cell lines' ret aining aspects of normal glial function. We conclude that methods based on the use of SV40 T antigen can successfull y immortalize human astrocytes, retaining key astrocyte functions, but T an tigen-induced proliferation appeared to interfere with expression of glial fibrillary acidic protein. We believe A735 is the first documented nontumor -derived human glial cell line which is immortal.