Dual Effects of TNF on Synthesis of Complement Components by a Gastric Cancer–Derived Cell Line, KATO–III

Abstract

Background: Complement components are synthesized extrahepatically, although hepatocytes are the major source of plasma complement. It is now clear that local production of complement is important in homeostasis and immune defense in tissue. Methods: The secretion of complement components was studied in vitro with a gastric cancer–derived cell line, KATO–III (signet–ring cell carcinoma). Complement components C2 and C3 were estimated by functional assay and/or ELISA in culture medium obtained after incubation of KATO–III cells for 3 days in protein–free culture medium, with or without addition of tumor necrosis factor (TNF), in a humidified atmosphere of 5% CO₂/95% air at 37°C. Results: (1) While a higher amount of C3 was detected in medium when KATO–III cells were cultured in the presence of TNF than in medium lacking TNF, higher C2 activity was detected when cultured in medium lacking TNF than in TNF–supplemented medium. (2) TNF suppressed C2 secretion and enhanced C3 secretion in a dose–dependent fashion. (3) C3 secretion remained less than 20 ng/10⁶ cells/24 h but increased from the first day of TNF (10U/ml) addition (concentrations approached 108.6– 115.6 ng/10⁶ cells/24 h on the 3rd day) and decreased on the 1st day without TNF. In contrast, C2 activity, detected when cultured in the absence of TNF, was decreased on the 2nd day of TNF addition and increased again on the 1st day without TNF. The daily secretion of C2 in the absence of TNF was 3.75–6.30×10⁷ effective molecules/10⁶ cells. (4) Reversible inhibition of C2 and C3 secretion was observed when the cell line was cultured in the presence of cycloheximide, indicating that both components were synthesized de novo. Conclusion: It appears that TNF enhances C3 secretion and suppresses C2 secretion by KATO–III.