Synergistic effects of tumor necrosis factor-α and insulin-like growth factor-I on survival of human trophoblast-derived BeWo cell line

Objective: Trophoblast survival is regulated by cytokines and growth factors. While the pharmacological levels (10-100 ng/mL) of tumor necrosis factor (TNF)- alpha affect trophoblasts survival in vitro, the effects of the physiological levels (1-10 pg/mL) of TNF-alpha remain unknown. We investigated the effects of the physiological levels of TNF-alpha on proliferation and apoptosis of human trophoblast cells by using BeWo cells. Insulin-like growth factor (IGF)-I is also a potent regulator of trophoblast survival and has been known to exert synergistic effects with other hormones. The interaction of IGF-I and TNF-alpha on BeWo cells survival was also examined. Methods: After incubating BeWo under the presence of TNF-alpha (10-10(5) pg/mL) and IGF-I (10(2) ng/mL), we assessed cell number by WST-1 assay and cell proliferation by BrdU uptake assay and immunocytochemistry with anti-Ki67 antibody. Apoptosis was evaluated by TUNEL assay and caspase-3, 8 activity assays. Results: Under the presence of IGF I, cell number, BrdU uptake, and Ki-67 expression of BeWo were dose dependently enhanced by low TNF-alpha (10-10(2) pg/mL), while no such effects were detected without IGF I. Higher levels of TNF-alpha (104-10(5) pg/mL) showed inhibiting effects on cell number and cell proliferation. The number of TUNEL positive cells were decreased and caspase activities were suppressed by lower levels (10-10(2) pg/mL) of TNF-alpha and IGF-I independently. Higher levels of TNF-alpha (104-10(5) pg/mL) showed promoting effects on apoptosis irrespective of IGF I. Conclusion: The physiological levels of TNF-alpha and IGF-I had synergetic effects on enhancing cell proliferation and also independently inhibited apoptosis of Bewo cells.