Immunosuppressive activity in human in vitro fertilization (IVF) culture supernatants and prediction of the outcome of embryo transfer: A multicenter trial

Abstract

The supernatants from cultured human oocytes fertilized in vitro contain low molecular weight factors that can suppress or stimulate the proliferative response of lymphocytes in vitro. The inhibitory and stimulatory effects are nonspecific and may be detected using cultured human or murine tumor cell lines. Using such a bioassay, we previously tested fetal cord serum-supplemented culture supernatant and found that an absence of suppression was correlated with an absence of subsequent pregnancy. To test this association further, additional samples were obtained from four different in vitro fertilization (IVF) units and studied blindly without knowledge of the pregnancy outcome. In this series, samples were obtained after the first 12–24 hr of sperm-oocyte incubation and all of the supernatants were from individual embryo cultures. The average number of preembryos transferred to those achieving pregnancy did not differ significantly from the number transferred to those not achieving pregnancy but the level of suppression was greater (8.7±1.9%) in those becoming pregnant compared to those not achieving pregnancy (0.8±1.5%). Twenty-two of 61 patients who received at least one embryo with a suppressive supernatant achieved pregnancy, whereas 0 of 19 patients received embryos lacking suppressive supernatants became pregnant. Two patients who received a single embryo from cultures with suppression became pregnant. Several problems with the bioassay method were defined. The culture medium in this series was always supplemented with adult serum, usually from the patient herself, and this serum could be suppressive. Also, cultures containing sperm were suppressive to a greater extent than could be explained by the serum. High-performance liquid chromatographic (HPLC) analysis indicated that the suppressive effect of the serum was related to molecules greater than 100,000 kd and that sperm supernatants contained additional activity in this size range. Pooled supernatants from patients with suppression achieving pregnancy confirmed the presence of two low molecular weight species of approximately 3700 and 1200, and these moieties were absent in those patients with suppressive supernatants who failed to become pregnant. Similar low molecular weight molecules were also present in supernatants from sperm alone, suggesting sperm as a possible origin. The data indicate that the presence of certain types of molecules in human IVF supernatants may predict an increased likelihood of implantation and subsequent pregnancy, whereas a deficiency of these molecules predicts failure. A more rapid assay to detect specifically the presence or absence of these molecules in supernatants from the first 24 hr of human IVF culture could be useful in sparing patients the transfer of embryos unlikely to succeed and in selecting embryos with an optimal chance of success.