OBJECTIVE: The object of this study was to determine the effect of epithelial growth factor (EGF), vascular endothelial growth factor (VEGF), and placental growth factor (PIGF) on the diferentiation of first-trimester and term cytotrophoblasts.
METHODS: The first-trimester trophoblasts were isolated from villous tissue obtained at suction termination (n = 5), and the term trophoblasts were isolated from placentas (n = 6) at elective cesarean. Cultured cells were stimulated with EGF, VEGF, or PIGF at 0.5, 5, and 50 ng/mL, in the presence or absence of NG-nitro-L-arginine methyl ester hydrochloride (10-4 M). Syncytialized trophoblasts were identified by immunostaining with antidesmosomal protein and anti-cytokeratin-7, whereas nuclei were counted in each syncytia using hematoxylin.
RESULTS: Without treatment, background levels of syncytialization were signficantly higher in term preparations than first-trimester cells. With VEGF and EGF, the number and size of syncytia increased signficantlyfor thefirst-trimester cytotrophoblasts (P < .05). Neither VEGF nor EGF had any effect on the syncytialization of cultured cells at term. Nitric oxide showed no involvement in syncytial induction, and PIGF had no effect on syncytialization of cytotrophoblasts, from either the first or third trimester.
CONCLUSION: Both EGF and VEGF appeared to enhance the in vitro syncytialization of first trimester cytotrophoblasts.
1. Athanassiades A, Lala PK. Role of placenta growth factor (PIGF) in human extravillous trophoblast proliferation, migration and invasiveness. Placenta1998;19:465-473.
2.
2. Athanassiades A, Hamilton GS, Lala PK. Vascular endothelial growth factor stimulates proliferation but not migration or invasiveness in human extravillous trophoblast. Biol Reprod1998; 59:643-654.
3.
3. Cartwright JE, Holden DP, Whitley GSJ. Hepatocyte growth factor regulates human trophoblast motility and invasion: A role for nitric oxide. Br J Pharmacol1999;128:181-189.
4.
4. Lash GE, Cartwright JE, Whitley GS, Trew AJ, Baker PN. The effects of angiogenic growth factors on extravillous trophoblast invasion and motility. Placenta1999;20:661-667.
5.
5. Clark DE, Smith SK, Licence D, Evans AL, Charnock-Jones DS. Comparison of expression patterns for placenta growth factor, vascular endothelial growth factor (VEGF), VEGF-B and VEGF-C in the human placenta throughout gestation. J Endocrinol1998;159:459-467.
6.
6. Shore VH, Wang TH, Wang CL, Torry RJ, Caudle MR, Torry DS. Vascular endothelial growth factor, placenta growth factor and their receptors in isolated human trophoblast. Placenta1997; 18:657-665.
7.
7. Matsuo H, Maruo T, Murata K, Mochizuki M. Epidermal growth factor regulates trophoblast proliferation and differentiation in an autocrine/paracrine manner. Nippon Sanka Fujinka Gakkai Zasshi1993;45:23-30.
8.
8. Maruo T, Matsuo H, Otani T, Mochizuki M. Role of epidermal growth factor (EGF) and its receptor in the development of the human placenta. Reprod Fertil Dev1995;7:1465-1470.
9.
9. Ahmed A, Dunk C, Kniss D, Wilkes M. Role of VEGF receptor-1 (Flt-1) in mediating calcium-dependent nitric oxide release and limiting DNA synthesis in human trophoblast cells. Lab Invest1997;76:779-791.
10.
10. Kliman HJ, Nestler JE, Sermasi E, Sanger JM, Strauss JF. Purification, characterization, and in vitro differentiation of cytotrophoblasts from human term placentae. Endocrinology1986;118: 1567-1582.
11.
11. Clover LM, Coghill E, Redman CWG, Sargent IL. A threecolour flow cytometry technique for measuring trophoblast intracellular antigens: The relative expression of TAP1 in human cytotrophoblast and decidual cells. Placenta2000;21:743-753.
12.
12. Ances IG. Serum concentrations of epidermal growth factor in human pregnancy. Am J Obstet Gynecol1973; 115:357-362.
13.
13. Hunter A, Aitkenhead M, Caldwell C, McCracken G, Wilson D, McClure N. Serum levels of vascular endothelial growth factor in preeclamptic and normotensive pregnancy. Hypertension2000; 36:965-969.
14.
14. Vuorela-Vepsalainen P, Alfthan H, Orpana A, Alitalo K, Stenman UH, Halmesmaki E. Vascular endothelial growth factor is bound in amniotic fluid and maternal serum. Hum Reprod1999;14:1346-1351.
15.
15. Blaschitz A, Weiss U, Dohr G, Desoye G. Antibody reaction patterns in first trimester placenta: Implications for trophoblast isolation and purity screening. Placenta2000;21:733-741.
16.
16. Douglas GC, King BF. Differentiation of human trophoblast cells in vitro as revealed by immunocytochemical staining of desmoplakin and nuclei. J Cell Sci1990;96:131-141.
17.
17. Vuorela P, Hatva E, Lymboussaki A, et al. Expression of vascular endothelial growth factor and placenta growth factor in human placenta. Biol Reprod1997;56:489-494.
18.
18. Hofmann GE, Scott RT, Bergh PA, Deligdisch L. Immunohistochemical localization of epidermal growth factor in human endometrium, decidua, and placenta. J Clin Endocrinol Metab1991;73:882-887.
19.
19. Charnock-Jones DS, Sharkey AM, Boocock CA, et al. Vascular endothelial growth factor receptor localization and activation in human trophoblast and choriocarcinoma cells. Biol Reprod1994;51:524-530.
20.
20. Shiraishi S, Nakagawa K, Kinukawa N, Nakano H, Sueishi K. Immunohistochemical locahzation of vascular endothelial growth factor in the human placenta. Placenta1996;17:111-121.
21.
21. Morrish DW, Dakour J, Li H, et al. In vitro cultured human term cytotrophoblast: A model for normal primary epithelial cells demonstrating a spontaneous differentiation programme that requires EGF for extensive development of syncytium. Placenta1997;18:577-585.
22.
22. Mayhew TM, Leach L, McGee R, Ismail WW, Myklebust R, Lammiman MJ. Proliferation, differentiation and apoptosis in villous trophoblast at 13-41 weeks of gestation (including observations on annulate lamellae and nuclear pore complexes). Placenta1999;20:407-422.
23.
23. Bulmer JN, Morrison L, Johnson PM. Expression of the proliferation markers Ki67 and transfemrn receptor by human trophoblast populations. J Reprod Immunol1988;14:291-302.
24.
24. Smith SC, Baker PN, Symonds EM. Placental apoptosis in normal human pregnancy. Am J Obstet Gynecol1997;177:57-65.
25.
25. Smith SC, Leung TN, To KF, Baker PN. Apoptosis is a rare event in first-tnrmester placental tissue. Am J Obstet Gynecol2000;183:697-699.
27. Gerber HP, McMurtrey A, Kowalski J, et al. Vascular endothelial growth factor regulates endothelial cell survival through the phosphatidylinositol 3'-kinase/Akt signal transduction pathway. Requirement for Flk-1/KDR activation. J Biol Chem1998;273: 30336-30343.
28.
28. Meekins JW, Pijnenborg R, Hanssens M, McFadyen IR, van Asshe A. A study of placental bed spiral arteries and trophoblast invasion in normal and severe pre-eclamptic pregnancies. Br J Obstet Gynaecol1994;101:669-674.
29.
29. Levy R, Smith SD, Chandler K, Sadovsky Y, Nelson DM. Apoptosis in human cultured trophoblasts is enhanced by hypoxia and diminished by epidermal growth factor. Am J Physiol Cell Physiol2000;278:982-988.
30.
30. Popovici RM, Irwin JC, Giaccia AJ, Giudice LC. Hypoxia and cAMP stimulate vascular endothelial growth factor (VEGF) in human endometrial stromal cells: Potential relevance to menstruation and endometrial regeneration. J Clin Endocrinol Metab1999;84:2245-2248.
