Reduction of annexin-V (placental anticoagulant protein-I) on placental villi of women with antiphospholipid antibodies and recurrent spontaneous abortion Jacob H. Rand, MD, Xiao-Xuan Wu, MD, Seth Guller, PhD, Joan Gil, MD, Arabinda Guha, PhD, Jonathan Scher, MD, and Charles J. Lockwood, MD New York, New York OBJECTIVE: The mechanism by which anti phospholipid antibodies are associated with pregnancy loss and thromboembolic conditions has yet to be elucidated. Annexin-V, an anticoagulant phospholipid-binding protein, is normally present in syncytiotrophoblasts lining the placental villi, where it may playa role in the maintenance of intervillous blood fluidity. We therefore investigated the distribution of annexin-V in placentas of patients with anti phospholipid antibodies in situ and then used short-term villous cultures to study the direct effect of antiphospholipid antibodies on the immunolocation of annexin-V. STUDY DESIGN: We performed a blinded study by means of computerized morphometric analysis of placental tissues that were stained for annexin-V with affinity-purified polyclonal antibody in an avidin-biotin peroxidase system. The distribution of villous surface annexin-V on cross sections of placentas of patients with antiphospholipid antibodies was compared with that of placentas from patients with uncomplicated pregnancies, elective abortions, and pregnancy losses not associated with anti phospholipid antibodies (n = 8 for each group). We quantitated villous surface annexin-V in cultured placental villi that were incubated with antiphospholipid antibodies immunoglobulin G compared with normal immunoglobulin G and measured annexin-V levels by enzyme-linked immunosorbent assay in conditioned media and in the villi. RESULTS: The mean villous surface annexin-V of the group with antiphospholipid antibodies was 26.2% ± 17% (SO) versus 93.9% ± 5.7% in the normal control group (p < 0.0001). Villi from patients undergoing elective abortions and with pregnancy losses that were not attributed to anti phospholipid antibodies also showed higher mean villous surface annexin-V levels (86.9% ± 10.6% and 83.5% ± 11.3%, respectively, p < 0.0001). Organ culture of normal placental villi with affinity-purified immunoglobulin G from patients with antiphospholipid antibodies showed a dose-dependent decrease of villous surface annexin-V over a concentration range of 1.5 IJ.g/ml to 1.5 mg/ml. Annexin-V concentrations in conditioned media were significantly lower in the presence of anti phospholipid antibodies immunoglobulin G compared with normal immunoglobulin G (49.4 ± 8.9 ng/gm wet weight vs 57.2 ± 11.5 ng/gm, respectively, p < 0.05). In contrast, the mean level of annexin-V in placental villi incubated with anti phospholipid antibodies immunoglobulin G was greater than in villi incubated with normal immunoglobulin G, 1328 ± 130 ng/gm wet weight versus 1183 ± 165 ng/gm (p < 0.02). CONCLUSIONS: Patients with anti phospholipid antibodies and a history of previous pregnancy losses have a significant reduction in annexin-V immunostaining on placental villous surfaces, and antiphospholipid antibodies immunoglobulin G can directly decrease levels of villous surface annexin-V on cultured placental villi. Assays of annexin-V in the conditioned media and cell pellets of cultured placental villi suggest that the mechanism for antiphospholipid antibodies-mediated reduced annexin-V surface staining is an inhibition of annexin-V transport to the villous surface rather than displacement by anti phospholipid antibodies from the surface. This anti phospholipid antibodies-induced deficiency of placental surface annexin-V may contribute to the placental thrombosis observed in these patients. (AM J OBSTET GYNECOL 1994;171 :1566-72.) Key words: Annexin-V, placental anticoagulant protein-I, placental villi, antiphospholipid antibodies, recurrent spontaneous abortion, thrombosis, syncytiotrophoblast From the Departments of Medicine; Obstetrics, Gynecology, and Repro- ductive Science; and Pathology; Mount Sinai School of Medicine. Presented in part at the Fourteenth Congress of the International Society of Thrombosis and Haemostasis, July 7, 1993, and at the Thirty-fifth Annual Meeting of the American Society of Hematology, December 6, 1993. Received for publication January 27, 1994; revised March 30, 1994; accepted July 13, 1994. 1566 Reprint requests: Jacob H. Rand, MD, Hematology Division, Box 1079, Mount Sinai School of Medicine, One Gustave Levy Place, New York, NY 10029. Copyright © 1994 by Mosby-Year Book, Inc. 0002-9378194 $3.00 + 0 6/1/59103