Tumor necrosis factor α receptors in microvascular endothelial cells from bovine corpus luteum

There is sufficient evidence to prove that tumor necrosis factor α (TNFα) modulates bovine corpus luteum (CL) function. Our previous study demonstrated that functional TNFα receptors are present on luteal cells in bovine CL throughout the estrous cycle. The purpose of the present study was to identify the presence of functional TNFα receptors on the microvascular endothelial cells derived from developing bovine CL. TNFα receptors were analyzed by a radioreceptor assay using ¹²⁵I-labeled TNFα on two types of cultured endothelial cells. One has a cobblestone appearance (CS cells), and the other has a tube-like structure (TS cells). ¹²⁵-Labeled TNFα binding was maximal after incubation for 30 h at 37°C, and the specificity of binding was confirmed. A Scatchard analysis showed the presence of two binding sites (high- and low-affinity) for TNFα receptors on both CS and TS cells. The dissociation constant (K(d)) values and concentrations of the high-affinity binding sites for TNF receptors were similar for CS and TS cells. However, K(d) values and concentrations of the low-affinity binding sites in CS cells were significantly higher than those in TS cells (P < 0.05 or lower). The expression of TNF receptor type 1 (TNF-RI) mRNA was determined in both cell types. Furthermore, TNFα significantly stimulated prostaglandin E₂ and endothelin-1 secretion by both CS and TS cells (P < 0.05 or lower). These results indicate the presence of two types of TNF receptors and the expression of TNF-RI mRNA in the endothelial cells derived from bovine CL, and suggest that TNFα plays two or more roles in regulating the secretory function of the endothelial cells.