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Growth factors contained in cultured medium of rat glioma C6 cells (C6 cells) were examined mainly for the activity of transforming growth factors (TGFs). Cultured medium of C6 cells was dialyzed against acetic acid, lyophilized and chromatographed by gel-permeation method, the fractions were assayed by soft agar colony formation, iodine 125 (125I)-epidermal growth factor (EGF)-binding competition and incorporation of tritium-thymidine. Two nontransformed cell lines, clonal NRK49F and BALB/3T3 A 31-1-1 (3T3) cells, were used as indicator cells for the soft agar colony assay. 3T3 cells were also used for the incorporation of tritium-thymidine. EGF receptor-rich A 431 cells were used for 125I-EGF-binding competition assay. The activity of alpha-type TGFs was examined by soft agar colony formation of NRK49F cells and inhibition of EGF-binding to A 431 cells since TGF alpha has sequence homology with EGF and binds to EGF receptors on the cell membrane, while the activity of beta-type TGFs was examined by soft agar colony formation of 3T3 cells and NRK 49 F cells with the addition of EGF. High level of activities of both TGF alpha and TGF beta were detected in 14,000 to 45,000 daltons, and also high level of the activity of DNA synthesis was detected at the same molecular weight. These results suggest that C6 cells produce TGF alpha and TGF beta as well as platelet-derived growth factors (PDGFs)-analogue. Since amplification of EGF receptor gene has been demonstrated in glioma, TGF alpha released by glioma may provide autocrine stimulation through the binding to the amplified EGF receptors. TGF beta is known to increase EGF receptors on the cell membrane. TGF beta has been demonstrated not only to stimulate but also inhibit cell proliferation under certain circumstances.(ABSTRACT TRUNCATED AT 250 WORDS)
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