ラット脳腫瘍細胞の産生する増殖因子に関する研究—特にtransforming growth factorについて

GROWTH FACTORS PRODUCED BY RAT GLIOMA CELLS: ACTIVITIES OF TRANSFORMING GROWTH FACTORS

　抄録　ラットグリオーマ細胞（C6細胞）の培養上清中に含まれる増殖因子，とくにtransforminggrowth factor （TGF）について検討した。C6細胞の培養上清を酢酸にて透析，凍結乾燥後，ゲル濾過した分画について，NRK49F細胞およびBALB／3T3細胞を指標細胞とした軟寒天培地コロニー形成法，¹²⁵I-EGF結合阻害試験，BALB／3T3細胞の³H-thymidineの取り込みにより，検討した。分子量14,000から45,000の領域に高いTGFαおよびTGFβ活性を認め，また同領域に高いDNA合成促進活性を認めた。これらの結果より，C6細胞がTGFα，TGFβを産生し，また血小板由来成長因子（PDGF）を産生している可能性が示唆され，これらの増殖因子がグリオーマの増殖に深く関与しているものと考えられた。

Growth factors contained in cultured medium of rat glioma C 6 cells (C 6 cells) were examined mainly for the activity of tranforming growth fac-tors (TGFs). Cultured medium of C 6 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 incorpora-tion of tritium-thymidine. Two nontransformed cell lines, clonal NRK49F and BALB/3 T 3 A 31-1-1 (3 T 3) cells, were used as indicator cells for the soft agar colony assay. 3 T 3 cells were also used for the incorporation of tritium-thymi-dine. EGF receptor-rich A 431 cells were used for ¹²⁵I-EGF-binding competition assay. The activity of α-type TGFs was examined by soft agar colonyformation of NRK49F cells and , inhibition of EGF-binding to A 431 cells since TGF a has se-quence homology with EGF and binds to EGF re-ceptors on the cell membrane, while the activity of β-type TGFs was examined by soft agar colony formation of 3 T 3 cells and NRK 49 F cells with the addition of EGF. High level of activities of both TGF a and TGF β were detected in 14, 000 to 45, 000 daltons, and also high level of the activi-ty of DNA synthesis was detected at the same molecular weight. These results suggest that C 6 cells produce TGF a and TGF β as well as platelet-derived growth factors (PDGFs)-analogue. Since amplification of EGF receptor gene has been demon-strated in glioma, TGF a released by glioma may provide autocrine stimulation through the binding to the amplified EGF receptors. TGF β is known to increase EGF receptors on the cell membrane. TGF β has been demonstrated not only to stimu-late but also inhibit cell proliferation under certain circumstances. Though only α-type TGF activiy has been detected in urine from patients with malignant glioma, it is interesting to point out that conditioned medium of C 6 cells showed a high level of both α-type and β-type TGF activities. It has been demonstrated that a PDGF-analogue is produced by glioma. Thus, cellular products, such as TGFs, PDGF, and other unknown factors have influence on tumor cell proliferation through the mechanism of autocrine stimulation.