Membrane topography and organization of cortical cytoskeletal elements and organeUes during early embryogenesis of the mouse have been studied by transmission and scanning electron microscopy with improved cellular preservation. At the four-and early eight-cell stages, blastomeres are round, and scanning electron microscopy shows a uniform distribution of microvilli over the cell surface. At the onset of morphogenesis, a reorganization of the blastomere surface is observed in which microvilli become restricted to an apical region and the basal zone of intercellular contact. As the blastomeres spread on each other during compaction, many microvilli remain in the basal region of imminent cell-cell contact, but few are present where the cells have completed spreading on each other. Microvilli on the surface of these embryos contain linear arrays of microfilaments with lateral cross bridges.Microtubules and mitochondria become localized beneath the apposed cell membranes during compaction. Arrays of cortical microtubules are aligned parallel to regions of apposed membranes. During cytokinesis, microtubules become redistributed in the region of the mitotic spindle, arid fewer microvilli are present on most of the cell surface. The cell surface and cortical changes initiated during compaction are the first manifestations of cell polarity in embryogenesis. These and previous findings are interpreted as evidence that cell surface changes associated with trophoblast development appear as early as the eight-cell stage. Our observations suggest that morphogenesis involves the activation of a developmental program which coordinately controls cortical cytoplasmic and cell surface organization.During the development of the preimplantation mouse blastocyst, a major period of membrane differentiation takes place at approx, the eight-cell stage. At this time, changes occur in membrane transport systems (4, 16), surface glycoproteins (33), surface antigens (2, 30, 50), and intercellular junctions (12).At the two-and four-cell stages, cell shape changes are limited to blastomere cleavage, whereas at the eight-cell stage morphogenetic al-THE JOURNAL OF CELL BIOLOt;V 9 VOLUME 74, 1977 9 pages 153-167 153 on

The topographical distributions of concanavalin A-binding sites on the surfaces of 3T3, proteasetreated 3T3, and simian virus 40-transformed 3T3 cultured mouse fibroblasts appear to be different, as shown by a shadow-cast replica technique using concanavalin A and a hemocyanin marker, or as shown previously on isolated membranes with concanavalin A coupled to ferritin. However, chemical fixation of cells before labeling with concanavalin A and hemocyanin, or labeling exclusively at 40, allows one to distinguish between inherent concanavalin A-binding-site topography and potential rearrangement of sites induced by the action of the multivalent concanavalin A molecule itself. The inherent distribution of binding sites on 3T3, protease-treated 3T3, and transformed cells is actually the same on all cells, i.e., dispersed and random. Treatment of unfixed transformed or protease-treated 3T3 cells, but not normal 3T3 cells, with concanavalin A and hemocyanin at 370 (or at 40 with subsequent warming to 37°), however, results in clustering of binding sites, presumably due to crosslinking of neighboring lectin-binding sites by the quadrivalent concanavalin A. Thus, the underlying difference between concanavalin A-binding sites on normal as compared with transformed or protease-treated normal cells lies not in the inherent topography of binding sites, but rather in the susceptibility of the sites to aggregation by concanavalin A. The latter may reflect an increased mobility of lectin-binding sites on transformed or protease-treated cells.

The effect of phagocytosis on lectin binding to plasma membranes of polymorphonuclear leukocytes was examined. The specific activities of binding sites of concanavalin A and Ricinus communis agglutinin (defined as M&g of lectin bound per 100 Mg of membrane protein) were measured on isolated membranes; they decreased in parallel with phagocytosis. Our data suggest that this removal occurs by concentration of binding sites into internalized membrane. Colchicine and vinblastine, which did not inhibit phagocytosis, prevented the selective removal of lectin-binding sites from the surface. It was also shown that at 370 lectins induce their own internalization. This property was used to define operationally three classes of lectin receptors, one of which is most extensively removed from plasma membrane during phagocytosis. Based on other morphological studies in which it is shown that before phagocytosis the surface distribution of concanavalin-binding sites is random, it is inferred that phagocytosis alters surface topography by inducing the selective movement of binding sites into membrane undergoing internalization and that colchicine-sensitive proteins are essential for this imposed topographical reorganization.According to the fluid mosaic model of cell surfaces, proteins are free to diffuse in the membrane of cells and thus should assume a random or homogeneous distribution (1). Yet, several studies have indicated that certain components (2) and functions (3) dependent on proteins are disposed over the surface nonrandomly. The resolution of this paradox is suggested by some morphological studies of the distribution of concanavalin A (Con A)-binding sites (CABS) on transformed cells (4, 5); the inherent distribution of CABS is random but CABS may be induced to cluster (i.e., assume a nonrandom distribution) by addition of exogenous Con A. We have also presented evidence that colchicine-sensitive proteins may modify the movement of CABS (6, 7) and membrane-transport proteins (8).During phagocytosis, particles are enveloped by cytoplasmic membrane which is then withdrawn into membranebound intracellular vesicles. Suitable particles can stimulate the internalization of a large portion of the membrane (3). Thus, phagocytosis causes the removal of an operationally defined region of the cell surface, permitting examination of the properties and composition of the residual surface mem-

Data in the literature suggest that circulating levels of lipoprotein(a) [Lp(a)] and insulinlike growth factor I (IGF-I) respond similarly to therapy with growth hormone, estrogen, or tamoxifen. To more clearly document these relations, we designed a randomized, double-blind, placebo-controlled study of the effects of tamoxifen and continuous estrogen on circulating levels of Lp(a), IGF-I, and IGF binding protein 3 (IGFBP-3) in healthy postmenopausal women. Both estrogen and tamoxifen decreased serum levels of IGF-I to 30% below baseline during the 3 months of treatment, while IGFBP-3 levels were unchanged. Plasma Lp{a) levels decreased to 24% below baseline after 1 month of treatment with either estrogen or tamoxifen (P<.05 for estrogen only); after 3 months Lp(a) decreased to C irculating concentration of lipoprotein (a) [Lp(a)] has been positively correlated with the incidence of coronary artery disease, 1 restenosis of coronary angioplasty, 2 vein graft stenosis after coronary artery bypass surgery, 3 and stroke/ The structure of Lp(a) consists of a low-density lipoprotein (LDL)-like particle, with a novel protein [apolipoprotein(a), or apo(a)] that is disulfide-linked to the apo B-100 moiety of the lipoprotein. The structure of the apo(a) gene is strikingly homologous to that of plasminogen, although there is a deletion of kringles 1 to 3 and a large number of tandem repeats of the kringle 4-like domain.5 However, unlike plasmin, the derivative of plasminogen, Lp(a) does not appear to demonstrate proteolytic activity that could influence fibrinolysis. 6Details of the metabolic regulation of plasma Lp(a) level are only partially understood. A major part of the variability in circulating Lp(a) level is determined by the size of the apo(a) isoform, with serum concentration being inversely correlated with apo(a) molecular weight. 7In general, plasma levels seem to be determined by the rate of secretion and not by the catabolic rate.

The distribution of surface-bound concanavalin A on the membranes of 3T3, and simian virus 40-transformed 3T3 cultured mouse fibroblasts was examined using a shadow-cast replica technique with a hemocyanin marker. When cells were prefixed in paraformaldehyde, the binding site distribution was always random on both cell types. On the other hand, labeling of transformed cells with concanavalin A (Con A) and hemocyanin at 37°C resulted in the organization of Con A binding sites (CABS) into clusters (primary organization) which were not present on the pseudopodia and other peripheral areas of the membrane (secondary organization). Treatment of transformed cells with colchicine, cytochalasin B, or 2-deoxyglucose did not alter the inherent random distribution of binding sites as determined by fixation before labeling. However, these drugs produced marked changes in the secondary (but not the primary) organization of CABS on transformed cells labeled at 37°C. Colchicine treatment resulted in the formation of a caplike aggregation of binding site clusters near the center of the cell, whereas cytochalasin B and 2-deoxyglucose led to the formation of patches of CABS over the entire membrane, eliminating the inward displacement of patches observed on untreated cells. The distribution of bound Con A on normal cells (3T3) at 37°C was always random, in both control and drug-treated preparations. Pretreatment of cells with Con A enhanced the effect of colchicine on cell morphology, but inhibited the morphological effects of cytochalasin B. The mechanisms that determine receptor movement and disposition are discussed.