Uteroferrin induces lipid peroxidation in endometrial and conceptus microsomal membranes and is inhibited by apotransferrin, retinol binding protein, and the uteroferrin-associated proteins

Abstract

Iron containing proteins catalyze lipid peroxidation when combined with either H2O2 or ascorbic acid (ASC). Microsomal membranes were prepared from Day 13 endometrial and conceptus tissues (5 pigs) and from Day 30 endometrial, placental, fetal liver, and fetus minus fetal liver tissues (5 pigs). Microsomal membranes were subjected to the following in vitro treatments: 1) no treatment, 2) 50 μM ASC, 3) 100 μM uteroferrin (UF), 4) 50 μM ASC + 100 μM UF, 5) 50 μM ASC + 100 μM UF + 10 μM apotransferrin (transferrin with no iron bound; ATF), and 6) 50 μM ASC + 100 μM UF + 10 μM holotransferrin (transferrin saturated with iron; HTF). For treatments 7 through 10, membranes were preincubated (0°C, 3 h) with either 7) no treatment, 8) 50 μM fetuin, 9) 50 μM holoretinol binding protein (holoRBP: retinol binding protein [HoloRBP] with retinol bound), or 10) 50 μM apoRBP (RBP with no retinol bound) followed by incubation with 50 μM ASC + 100 μM UF. Lipid peroxidation was measured in the samples as thiobarbituric acid reactive substances (TBARS). Endogenous TBARS were greater (p < 0.05) in Day 13 conceptus than in Day 13 endometrium and were highest (p < 0.05) on Day 30 in fetal liver. Combined ASC and UE caused a large increase (p < 0.05) in TBARS in all membranes except Day 30 placental membranes. Addition of ATF, but not HTF, decreased TBARS production in all membrane preparations. HoloRBP, but not fetuin or apoRBP, decreased (p < 0.05) TBARS production in all but Day 30 endometrial membranes. In other experiments, when combined with ASC, UF/UF-associated protein complex induced less (p < 0.01) lipid peroxidation in fetal liver microsomal membranes than did free UF. Catalase and superoxide dismutase had no effect on UF-induced lipid peroxidation in fetal liver membranes. These results indicate that 1) UF combined with ASC induces lipid peroxidation in Day 13 endometrial and conceptus and Day 30 endometrial, fetal liver, and fetus minus liver microsomal membranes, and 2) ATF, holoRBP, and the UF-associated proteins, but not catalase or superoxide dismutase, inhibit this reaction.