Lymphocyte Surface Membrane Research Articles

Binding of purified lectins to guinea pig lymphocytes: Studies of the number, binding constant, and distribution of Lens culinaris lectin A and Agaricus bisporus lectin molecules on lymphocyte surfaces

The surface membrane of guinea pig lymphocytes contains discrete receptors for several different homogeneous lectins. Two very similar lectins from the lentil, LcL-A and LcL-B, bind to the same receptor. Three other lectins, AbL, WGA, and Con A, do not bind detectably to the LcL receptor. AbL binds to another discrete receptor to which none of the other lectin molecules bind. The LcL and AbL receptors are contained on different units, each of which is capable of independent movement on the cell surface. The normal distribution of LcL molecules on the surface is different than the distribution of AbL molecules. However, when lymphocytes are simultaneously incubated with LcL and AbL, the distribution of bound LcL molecules on the cell surface parallels the normal distribution of AbL molecules, which indicates that the movement of AbL molecules and AbL receptors on the surface can strongly influence the movement of LcL and LcL receptors. When lymphocytes are held in close contact by LcL or AbL, the lectin molecules and receptors appear to concentrate at the area of contact between two cells. A model is presented which suggests that the migration of lectins and lectin receptors can occur by a multicellular process.

Cell surface galactosyltransferase and lectin agglutination of thymus and spleen lymphocytes.

Lectin agglutination and cell surface galactosyltransferase (EC 2.4.1.67; 1-O-alpha-D-galactosyl-myo-inositol:raffinose galactosyltransferase) enzyme activity have been studied with thymus and spleen lymphocytes of neonatal rats. Thymus lymphocytes were more agglutinable by concanavalin A than by wheat germ agglutinin, whereas spleen lymphocytes were more agglutinable by wheat germ agglutinin than by concanavalin A. Thymus lymphocytes, but not spleen lymphocytes, of neonatal rats could be blast transformed by concanavalin A. Cell surface galactosyltransferase activity was present on both types of lymphocytes, but was greatly increased on thymus cells after blast transformation. The differences in lectin agglutination suggest a difference in the surface membranes of thymus and spleen lymphocytes. The increase in cell surface galactosyltransferase activity with blast transformation of thymus lymphocytes may be related to the exteriorization of the Golgi apparatus into the plasma membrane.

Difference in the cellular cholesterol to phospholipid ratio in normal lymphocytes and lymphocytic leukaemic cells.

FLUIDITY of the fatty acyl chains of phospholipids in cell membranes is required for essential cellular activities, and the degree of fluidity can be influenced by cholesterol. Changes in the ratio of cholesterol to phospholipids can, therefore, affect the internal viscosity and molecular motion of lipids within membranes and this may function as a regulator in cell behaviour1–4. Our studies of the surface membrane of normal lymphocytes and lymphocytic leukaemic cells have shown that the surface membrane of the leukaemic cells has a lower rotational mobility of concanavalin A binding sites5,6, and a higher fluidity of the lipid layer6,7. We have now found that these differences in fluidity are associated with a difference in the cholesterol to phospholipid ratio in these cells.

Surface immunoglobulin on rabbit lymphoid cells: I. Ultrastructural distribution and endocytosis of b4 allotypic determinants on peripheral blood lymphocytes

Immunoglobulin (Ig) b4 allotypic determinants are detected on the surface membrane of rabbit peripheral blood lymphocytes by an indirect immunoferritin labeling technique. Cells coated with antiallotype antibodies are labelled with soluble complexes of ferritin and rabbit antiferritin of a given allotype. At 0 °C a patchy distribution of labeled surface immunoglobulin is visualized on 80% of the lymphocytes examined. Warming of the cells for 1–5 min at 37 °C causes rapid endocytosis of surface label in a perinuclear fashion. Cap formation is not observed. Cross-linking of immunoferritin labelled surface determinants with sheep anti-rabbit Ig (SARG) inhibits endocytosis and promotes aggregation of small surface patches. Indirect evidence suggests that sloughing and/or stripping of labelled surface Ig can occur after this aggregation. These surface changes may be the first step in the induction of lymphocyte activation.

Comparison of Normal and CLL Lymphocyte Surface Ig Determinants Using Peroxidase-labeled Antibodies. I. Detection and Quantitation of Light Chain Determinants

Abstract Chronic lymphocytic leukemia (CLL) and normal human blood lymphocytes were investigated with regard to their membrane-associated light (L) chains. Peroxidase-labeled anti-kappa or anti-lambda antibodies were used to visualize by light microscopy the cells bearing L chain determinants. In addition, the activity of the enzyme coupled to the antibody was measured spectrophotometricly in order to determine the number of antigenic determinants on each positive cell. The results demonstrated that 16%-25% of lymphocytes from six normal control subjects were positive for kappa chains, and 4%-10% were positive for lambda chains. The average number of antigenic sites on the surface of each positive cell was calculated to be 90,000. Lymphocytes of each of the 14 CLL patients studied carried either kappa or lambda antigenic determinants on their surface, but not both; 50%-70% of the lymphocytes were stained. The average number of antigenic sites per positive CLL was calculated to be 9000. These results confirm previously reported studies indicating that CLL is a monoclonal proliferative disease. In addition, the quantitative results demonstrate that the CLL lymphocyte surface membrane bears, on the average, only 10% of the L chain determinants present on the normal lymphocyte.

The lymphocyte uropod: a specialized surface site for immunologic recognition.

Functional specialization of the cell surface of motile protozoa such as the Paramecium and amoeba are generally recognized (Ambrose and Forrester, 1968; Wolpert and Gin-gell, 1968). Less well appreciated is the existence of differentiated surface membranes on mammalian leukocytes. In this laboratory we have been interested in a modification of the lymphocyte surface called the uropod, an area consisting of microvillus projections of cell membrane adjacent to the golgi-associated cell pole. The cytoplasm contained in the uropod is rich in microtubules, mitochondria, endoplasmic reticulum, and numerous endocytic vesicles, and is quite distinct from the pseudopod region which contains few such subcellular organelles. Moreover, the uropod is not engaged in cell motility as is the pseudopod. The lymphocyte surface membrane functions in immunologic recognition, cell cooperation, and cell-mediated cytotoxicity in an as yet undefined manner. Circumstantial evidence suggests that the uropod may represent the site of such interactions.

Antibody-mediated internalization of B lymphocyte surface membrane immunoglobulin

The ultrastructural correlates of antibody-mediated internalization of membrane immunoglobulin determinants has been studied in the guinea pig using a horseradish peroxidase conjugate of rabbit anti-guinea pig immunoglobulin. We have observed that polar flow of ligand-induced micro-aggregated membrane immunoglobulin proceeds by local deformation of the plasma membrane into open-ended endocytic vesicles which in turn accumulate at the Golgi-associated pole prior to completion of endocytosis. ‘Capping’ does not appear to result from simple diffusion of the aggregates within the plane of the membrane. In addition, the question of whether anti-immunoglobulin can induce the formation of uropods on B lymphocytes in the guinea pig was examined. Formation of uropod-like structures on B lymphocytes appears to be factitious and induced by accumulation at the Golgi-associated cell pole of non-internalizable aggregates of membrane immunoglobulin linked to cell debris. These observations are interpreted as supporting the concept that spontaneous uropod formation by lymphocytes in the absence of anti-immunoglobulin reagents is a characteristic of antigen-reactive thymus-derived lymphocytes in the guinea pig.

Immunoglobulin nature of PPD receptors on human T lymphocytes.

READILY demonstrable immunoglobulin (Ig) determinants on mouse and human lymphocyte surface membrane suggests that they are probably bone-marrow-derived (B) cells1–4. There is much controversy about the presence of Ig on the cell surface and no conclusive evidence has been obtained either by direct methods of detecting Ig determinants5–10 or by inhibition of lymphocyte function by anti-Ig sera11–17. We studied an indisputable result of cell activity, human tuberculin hypersensitivity and measured the effects of antisera directed against various human Ig fragments on macrophage migration inhibition18 by human lymphocytes in the presence of purified protein derivative (PPD), as described previously19. This is an acceptable in vitro correlate of cell-mediated immunity in man20–22.

Rotational diffusion of lectins bound to the surface membrane of normal lymphocytes

Rotational diffusion of lectins bound to the surface membrane of normal lymphocytes

Difference in the mobility of lectin sites on the surfact membrane of normal lymphocytes and malignant lymphoma cells.

AbstractBinding sites for Concanavalin A (Con A) can move on the surface membrane. This induction of movement resulted in the formation of caps of Con A membrane‐site complexes in normal lymph‐node or spleen lymphocytes, but not in a line of malignant lymphoma cells. This indicates a difference in the mobility of Con A sites on the surface membrane of normal lymphocytes and lymphoma cells. Binding sites for the lectin from wheat germ (WGA) did not form caps either on normal lymphocytes or on lymphoma cells. Normal lymphocytes with caps induced by Con A still showed binding of WGA to the rest of the membrane. The mobility of Con A sites was, therefore, independent of WGA sites. The difference in the mobility of binding sites for Con A on the surface membrane of normal lymphocytes and lymphoma cells suggests that changes in the fluid state of the carbohydrate‐containing structures on the surface where the Con A sites are located, are associated with malignant cell transformation.

IgD on the surface of peripheral blood lymphocytes of the human newborn.

Immunoglobulin determinants have been demonstrated on the surface membrane of lymphocytes in man and other species1. Immunoglobulin on individual human lymphocytes is restricted to a single class, subclass and allotype2. Here we report the occurrence of IgD on a large percentage of the lymphocytes of human umbilical cord blood.

Cytotoxic and stimulatory effects of antisera to membrane associated antigens of lymphocytes.

Antisera against subcellular rat lymphocyte fractions, an unfractionated lymphocyte homogenate, a purified microsomal fraction and a soluble cytoplasmic fraction were produced in rabbits. The highly cytotoxic antisera against the microsomal fraction exhibited only a low stimulatory activity. The antisera against the cytoplasmic fraction were less cytotoxic, but stimulated the DNA-synthesis in rat lymphocyte cultures very effectively. Antisera against the ULH showed high cytotoxic titers and strong stimulatory properties. The cytotoxic activity could be removed by absorption with the ULH and the microsomal fraction; the stimulatory activity was reduced only by absorption with the ULH. Absorption with the cytoplasmic fraction was ineffective. In immunofluorescence studies and with the immunoferritin technique it could be demonstrated that the microsomal as well as the soluble cytoplasmic antigen is a constituent of the lymphocyte surface membrane which is distributed in patches on the membrane in different patterns. The immunofluorescence studies indicate that thymus cells lack some antigenic determinants which are present on membranes of lymph node and spleen cells.

Location of the antigen, antibody and antigen-antibody complexes in delayed-type hypersensitivity skin reactions to horseradish peroxidase.

The sites of intradermally injected horseradish peroxidase (HRP), of its antibody and of HRP-anti-HRP antibody complexes were investigated in the dermis of rabbits showing strong skin reactions of delayed hypersensitivity against HRP. Comparison was made with the location of intradermally injected HRP and its antibody in the dermis of immunized rabbits showing slight degrees of hypersensitivity of a nondelayed type and with the location of intradermally injected HRP in the dermis of nonimmunized animals. In all skin reactions of delayed hypersensitivity, a considerable proportion of the injected antigen was associated with collagen fibers, the reaction product appearing as "dots," "rodlets" and "strands" in linear array on individual collagen fibers in the interior of the bundles or as strands coating the outermost surface of the bundles. The reaction for the antibody was positive at the same sites. It was suggested that receptors for antigen-antibody complexes may be associated with collagen fibers. Free granular deposits containing antigen-antibody complexes seemed to originate from the collagen-associated granules after their enlargement and release. The extracellular granules often adhered to the antibody-positive surface membranes of lymphocytes. Other granular deposits (antigen-antibody complexes) seemed to be phagocytosed by monocytes (macrophages) and fibroblasts. In areas of tissue injury in the upper dermis, many mononuclear cells showed the reaction for the antibody on their cell surface. Aggregations of such cells were often located close to disintegrating bundles of collagen fibers. It was suggested that the encounter of the antigen and the antibody (antigen-antibody complexes) on the surfaces of cells and of collagen may trigger the release of cytotoxic factors. A few lymphocytes in the dermis of hypersensitive rabbits contained the specific antibody on their cell surface before skin lesions were elicited.

Surface membrane of lymphocytes. 1. Confirmation of antigenicity of lymphocytes.

Injection of serum or immunoglobulin, obtained after immunization of lymphocytes or thymocytes, caused lymphopenia either in normal rats or in RES‐blocked rats. Control rabbit serum, immunized with rat liver, caused leucocytosis instead of lymphopenia. The dose of administration was related to the degree of leucopenia.The presence of lymphocytic specific antigen determinants was confirmed with Ouchterlony agar plate method as well as other various heterologous antigens. Antilymphocytic antibody disappeared after absorption with lymphoid cells.Ferritin‐conjugated‐ALG were adsorbed along the plasma membrane of lymphocyte and did not infiltrate into the cytoplasm.

Interaction of phytohemagglutinin with plasma membranes of pig lymphocytes and thymus cells

Phytohemagglutinin (PHA) stimulated pig small lymphocytes and thymus cells to transform and synthesise DNA. The lymphoagglutinating and lymphocyte-stimulating activities of PHA were adsorbed by whole lymphocytes and by the purified plasma membranes of lymphocytes and thymus cells. The lymphocyte plasma membrane was 22 times as effective as whole cells (on a dry wt basis) in removing the lymphocyte-stimulating activity. The capacities of the thymus plasma membrane to adsorb the PHA activities were similar to those of the lymphocyte plasma membrane. It was concluded that thymus cells are stimulated to transform by PHA, and that the surface membranes of lymphocytes and thymus cells possess a similar number of receptors for PHA. Sucrose density gradient centrifugation of lymphocyte plasma membrane dissolved in 2% sodium deoxycholate gave a fraction which inhibited the lymphoagglutinating and lymphocyte-stimulating activities of PHA; the sedimentation coefficient of the inhibitory material was between those of immunoglobulin G and ovalbumin. The results suggest that the initial event in the transformation of lymphocytes by PHA is the selective binding of the PHA mitogen by a receptor on the cell surface.

Fine Structural Localization of Adenosine Triphosphatase in Human Platelets and Other Blood Cells

Abstract Adenosine triphosphatase activity has been localized in human blood cells by combined histochemical and electron microscopic technics. Deposits of lead phosphate, indicative of enzymic activity, were observed on the surface membranes of erythrocytes, lymphocytes and platelets. Intracellular ATPase activity was found in lymphocyte mitochondria and nucleoli. Blood platelets contained organelles in their hyaloplasm with dense accumulations of the enzyme reaction product. The distribution of lead phosphate in platelet organelles suggested that they were mitochondria, but some platelet granules may also contain ATPase activity.