Summary
Low density lipoproteins (LDL) were conjugated to colloidal gold to visualize the route for internalization of LDL in the cultured cells of human term placenta. Cells were obtained from placental villi (caesarian section) by a standard trypsin-DNase dispersion method followed in some cases by a Percoll gradient centrifugation step. Employing electron microscopy it was observed that after 3 days of culture, cells obtained by trypsin-DNAse dispersion were a mixture of macrophages, mononucleated cells and large multinucleated cells. When the cells were incubated for 3 days after the Percoll purification, essentially multinucleated cells identical to the syncytiotrophoblast were present. The number of LDL receptor was increased by preincubation in medium with lipoprotein depleted serum. In binding experiments cells incubated at 4° C for 2 h with medium containing gold LDL conjugates showed gold LDL attached to the plasma membrane without characteristic localization. After incubation with gold LDL at 37° C for various times, the three cellular types showed ligand internalization. Gold LDL endocytosis involved first coated pits but also uncoated plasmalemmal invaginations. Then gold LDL was further observed in coated and non coated vesicles, smooth walled endosomes, multivesicular bodies and tubular vesicles. Lastly free gold particles were observed in lysosome like dense bodies. These results prove the internalization of gold LDL conjugates by human cultured placental cells, particularly by syncytiotrophoblast like multinucleated cells. This accumulation of LDL (the major cholesterol carrying protein in humans) is recognised to be responsable for the exogenous cholesterol supply indispensable to the progesterone biosynthesis and cellular growth of the placenta.
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References
Alsat E, Bouali Y, Goldstein S, Malassiné A, Laudat MH, Cedard I (1982) Characterization of specific low-density lipoprotein binding sites in human term placental microvillous membranes. Mol Cell Endocrinol 28:439–453
Alsat E, Bouali Y, Goldstein S, Malassiné A, Berthelier M, Mondon F, Cedard L (1984) Low density lipoprotein binding sites in the microvillous membranes of human placenta at different stages of gestation. Mol Cell Endocrinol 38:197–203
Alsat E, Mondon F, Rebourcet R, Berthelier M, Erlich D, Cedard L, Goldstein S (1985) Identification of specific binding sites for acetylated low density lipoprotein in microvillous membranes from human placenta. Mol Cell Endocrinol 41:229–235
Anderson RGW, Goldstein JL, Brown MS (1976) Localization of low density lipoprotein receptors on plasma membrane of normal human fibroblasts and their absence in cells from familial hypercholesterolemic homozygote. Proc Natl Acad Sci USA 73:2434–2438
Branchaud CL, Goodyer CG, Lipowski LS (1983) Progesterone and estrogen production by placental monolayer cultures effects of dehydroepiandrosterone and luteinizing hormone-releasing hormone. J Clin Endocrinol 56:761–766
Brown M, Goldstein JL (1986) A receptor-mediated pathway for cholesterol homeostasis. Science 232:34–47
Brown MS, Basu SK, Falck JR, Ho YK, Goldstein JL (1980) The scavenger cell pathway for lipoprotein degradation: specificity of the binding site that mediates the uptake of negativelycharged LDL by macrophages. J Supramol Struct 13:67–81
Contractor SF, Sooranna SR (1985) Formation of multinucleated cells in human placental cell cultures. IRCS Med Sci 13:1137–1138
Cummings SW, Hatley W, Simpson ER, Ohashi M (1982) The binding of high and low density lipoproteins to human placental membrane fractions. J Clin Endocrinol Metab 54:903–908
Frens G (1973) Controlled nucleation for the regulation of the particle size in monodisperse gold suspensions. Nature (Phys Sci) 241:20–24
Geoghegan WD, Ackerman A (1977) Adsorption of horseradish peroxidase, ovomucoid and anti-immunoglobulin to colloidal gold for the indirect detection of concanavalin A, wheat germ agglutinin and goat antihuman immunoglobulin G on cell surfaces at the electron microscopic level: a new method, theory and application. J Histochem Cytochem 25:1187–1200
Gwynne JT, Strauss III JF (1982) The role of lipoproteins in steroidogenesis and cholesterol metabolism in steroidogenic glands. Endocrine Rev 3:299–329
Hall CStG, James TE, Goodyer C, Branchaud C, Guyda H, Giroud CJP (1977) Short term tissue culture of human midterm and term placenta: parameters of hormonogenesis. Steroids 30:569–580
Handley DA, Arbeeny CM, Witte LD, Chien S (1981) Colloidal gold-low density lipoprotein conjugates as membrane receptor probes. Proc Natl Acad Sci USA 78:368–371
Havel RJ, Eder HA, Bragdon JH (1955) The distribution and chemical composition of ultracentrifugally separated lipoproteins in human serum. J Clin Invest 34:1345–1353
Hellig H, Gattereau D, Lefebvre Y, Bolte E (1970) Steroid production from plasma cholesterol. I. Conversion of plasma cholesterol to placental progesterone in humans. J Clin Metab 30:624–631
King BF (1976) Localization of transferrin on the surface of the human placenta by electron microscopy immunocytochemistry. Anat Rec 186:151–159
Kliman HJ, Nestler JE, Sermasi E, Sanger JM, Strauss III JF (1986) Purification, characterization, and in vitro differentiation of cytotrophoblasts from human term placentae. Endocrinology 118:1567–1582
Lai WH, Guyda HJ, Bergeron JJM (1986) Binding and internalization of epidermal growth factor in human term placental cells in culture. Endocrinology 118:413–423
Lin CT (1980) Immunoelectron microscopic localization of immunoglobulin G in human placenta. J Histochem Cytochem 28:339–346
Lobo JO, Bellino FL, Bankert L (1985) Estrogen synthetase activity in human term placental cells in monolayer culture. Endocrinology 116:889–895
Loke YW (1983) Human trophoblast in culture. In: Loke YW, Whyte A (eds) Biology of trophoblast. Elsevier Science. Amsterdam, pp 663–761
Malassiné A, Goldstein S, Alsat E, Merger Ch, Cedard L (1984) Ultrastructural localization of low density lipoprotein binding sites on the surface of the syncytial microvillous membranes of the human placenta. IRCS Med Sci 12:166–167
Mommaas-Kienhuis AM, Nagelkerle JF, Vermeer BJ, Daems WT, Van Berkel TJC (1985a) Visualization of the interaction of native and modified low density lipoproteins with isolated rat liver cells. Eur J Cell Biol 38:42–50
Mommaas-Kienhuis AM, Van der Schroeff JG, Wijsman MC, Daems WT, Vermeer BJ (1985b) Conjugates of colloidal gold with native and acetylated low density lipoproteins for ultrastructural investigations on receptor-mediated endocytosis by cultured human monocyte-derived macrophage. Histochemistry 83:29–35
Morales C, Hermo L (1986) Intracellular pathways of endocytosed transferrin and non-specific tracers in epithelial cells lining the rete testis of the rat. Cell Tissue Res 245:323–330
Morrish DW, Siy O (1986) Critical factors in establishing monolayer cultures of normal human placental cells in serum-free medium. Endocrine Res 12:229–253
Nelson BM, Smith RM, Jarett LJ (1978) Non uniform distribution and grouping of insulin receptors on the surface of human placental syncytial trophoblast. Diabetes 27:530–538
Paavola LG, Strauss III JF, Boyd CO, Nestler JE (1985) Uptake of gold-and 3H-cholesterol linoleate-labeled human low density lipoprotein by cultured rat granulosa cells: cellular mechanisms involved in lipoprotein metabolism and their importance to steroidogenesis. J Cell Biol 100:1235–1247
Rebourcet R, Alsat E, Mondon F, Berthelier M, Pastier D, Cedard L, Goldstein S, Polonovski J (1986) Detection of distinct receptors for native and acetylated low density lipoproteins in human placental microvilli by ligand-immunoblotting. Biochimie 68:1135–1140
Robenek H, Rassat J, Hesz A, Grünwald J (1982) A correlative study on the topographical distribution of the receptors for low-density lipoprotein (LDL) conjugated to colloidal gold in cultured human skin fibroblasts employing thin section, freezefracture, deep-etching and surface replication techniques. Eur J Cell Biol 27:242–250
Robeneck H, Hesz A (1983) Dynamics of low-density lipoprotein receptors in the plasma membrane of cultured human skin fibroblasts as visualized by colloidal gold in conjunction with surface replicas. Eur J Cell Biol 31:275–282
Vasile E, Simionescu M, Simionescu N (1983) Localization of the binding, endocytosis, transcytosis of low density lipoprotein in arterial endothelium in situ. J Cell Biol 96:1677–1678
Vermeer BJ, Wijsman MC, Mommaas-Kinehuis AM, Ponec M, Havekes L (1985) Modulation of low density lipoprotein receptor activity in squamous carcinoma. Eur J Cell Biol 38:353–360
Willingham MC, Pastan I (1985) Morphologic methods in the study of endocytosis in cultured cells. In: Pastan I, Willingham MC (eds) Endocytosis. Plenum Press, New York, pp 281–321
Winkel CA, Snyder JM, McDonald PC, Simpson ER (1980a) Regulation of cholesterol and progesterone synthesis in human placental cells in culture by serum lipoproteins. Endocrinology 106:1054–1060
Winkel CA, Gilmore J, McDonald PC, Simpson E (1980b) Uptake and degradation of lipoproteins by human trophoblastic cells in primary culture. Endocrinology 107:1892–1898
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Malassiné, A., Besse, C., Roche, A. et al. Ultrastructural visualization of the internalization of low density lipoprotein by human placental cells. Histochemistry 87, 457–464 (1987). https://doi.org/10.1007/BF00496817
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DOI: https://doi.org/10.1007/BF00496817