Calcofluor white alters the assembly of chitin fibrils in Saccharomyces cerevisiae and Candida albicans cells

In the presence of calcofluor white, budding scars and dividing cross-walls of Saccharomyces cerevisiae exhibited fluorescence, indicating that the brightener was a specific marker of fungal chitin. In addition, incubation of cells in the presence of the brightener did not stop protein and wall-polymer formation, but abnormal deposition of chitin occurred. Chitin synthesis was normal in regenerating protoplasts of Candida albicans in the presence of calcofluor, but formation of the crystalline lattice was blocked. These results suggest that crystallization of nascent subunits may occur by a self-assembly mechanism that was blocked by the stain.     

Corpus allatum activity during overlapping cycles of oöcyte growth in adult female Melanoplus sanguinipes

Cycles of oögenesis in Melanoplus sanguinipes overlap to the extent that there are always 2 and occasionally 3 sets of vitellogenic oöcytes in the ovarioles at any one time. Three phases of vitellogenic oöcyte development can be distinguished: (1) An initial 24-hour phase of slow development (1.0–1.2 mm, 0.05–0.10 mm³). (2) A phase of rapid oöcyte growth (1.2–3.5 mm, 0.1–1.3 mm³). The duration of this phase is 2 days in the first cycle and 3 days in subsequent cycles. (3) A final phase of rapid oöcyte growth and maturation (3.5–4.5 mm, 1.3–2.8 mm³). Including the time taken for oviposition the duration of this latter phase is 3 days. Phases 1, 2 and 3 of cycles n + 2, n + 1 and n, respectively, overlap entirely. Activity of the corpora allata was measured using a radio-biosynthetic technique. A period of increased corpus allatum activity coincides with the initial part of phase 2 in each cycle. Each set of oöcytes is, thus, subject to 2 and occasionally 3 peaks of corpus allatum activity during development. Using these data a model of the control of oöcyte development has been devised     

Composition, secretion, and fate of the glands in the miracidium and sporocyst of Fasciola hepatica L

The anterior end of the miracidium of Fasciola hepatica contains a large flask-shaped apical gland and four unicellular lateral glands, all of which have ducts which pass to the tip of the apical papilla. These glands appear to be involved in penetration of the larva into the snail host. The apical gland secretes as the miracidium proves the epidermis of the host before attachment. It seems likely that its secretion is a chemical which lyses the epidermal cells. The lateral glands are PAS-positive and may contain a neutral mucopolysaccharide. They also secrete as the miracidium probes the snail and a layer of PAS-positive material may be seen at the leading edge of the apical papilla as the larva penetrates into the host. Both the apical gland and the lateral glands may be visible in the sporocyst for several days after penetration.     

Crystals of the human heavy chain disease protein Riv and human Fc fragment are isomorphous: further evidence for conformational flexibility in the hinge region of immunoglobulins

Protein Riv is a human gamma 1 heavy chain disease immunoglobulin variant with a deletion of the entire VH and CH1 domains and consisting of most of the hinge region plus the CH2 and CH3 domains. Crystals of this protein are orthorhombic, belonging to the space group P2(1)2(1)2(1), with a = 80.1 A, b = 145.5 A, c = 50.1 A. These crystals are shown to be isomorphous with crystals of a human Fc fragment, indicating that the hinge region and the initial part of the CH2 domain of protein Riv do not assume a unique conformation in the crystalline state.     

Blocking of human T lymphocyte functions by anti-Leu-2 and anti-Leu-3 antibodies: differential inhibition of proliferation and suppression

We have shown previously that monoclonal antibodies to the Leu-2 and Leu-3 T cell antigens block the response of their respective subsets in allogeneic MLR. The present study was an effort to explore the mechanism of inhibition and to determine if anti-Leu-2 and anti-Leu-3 antibodies affect the responses to stimuli in addition to alloantigens. Our results indicate that antibodies to Leu-2 and Leu-3 have profound inhibitory effects on proliferation by their respective T cell subsets responding to a variety of stimuli, including specific soluble antigens and alloantigen. This effect was characterized by the following features: a) For optimal inhibition of proliferation, antibody must be present at the onset of antigenic stimulation. b) Inhibition is augmented by increasing the concentration of antibody or decreasing the concentration of antigen. c) Fab fragments of both anti-Leu-2a and anti-Leu-3a antibodies also block proliferation. In addition to their effects on T cell proliferation, anti-Leu-3 antibody blocked T cell-dependent lg synthesis induced in MLR, and anti-Leu-2 antibody prevented the induction, in vitro, of Leu-2+3- suppressor cells of lg synthesis. Taken together, these results suggest that antibodies to antigenic determinants on the Leu-2 and Leu-3 molecules competitively block segments of these structures that bind to alloantigen or nominal antigen. On the other hand, anti-Leu-2a antibody failed to block suppression of the MLR by in vivo activated, antigen-specific Leu-2+3- suppressor cells, which suggests that the Leu-2a epitope does not transmit antigen-specific signals from these differentiated suppressor T cells.     

The "lazy" NK cells of Chediak-Higashi syndrome

Natural killer (NK) function, measured in a short-term (4-hr) 51Cr-release assay, is profoundly depressed in circulating PBL of donors with Chediak-Higashi syndrome (CHS). In this study, we demonstrate that CHS NK cells can express relatively normal lytic function after prolonged exposure in vitro to high levels of activating as well as cytotoxic stimuli. After activation with the human cloned interferon (B1) for 24 hr, CHS NK cells have lytic activity comparable to unactivated normals in a 4-hr 51Cr-release assay. In addition, after 5 days of activation with mitomycin C-treated B cell lines, CHS NK cells have levels of activity similar to those of activated normals but are defective in generating cytotoxic cells capable of lysing the stimulator B cell. Even though CHS NK cells are defective in a 4-hr 51Cr-release assay, after 16 hr they enhance their killing capability 200 to 400-fold. In fact, after 16 hr of interaction with K562 target cells, CHS NK cells are capable of releasing NK soluble cytotoxic factors. These results are consistent with the hypothesis that CHS NK cells have all the necessary cellular structures and molecules required for them to function as lytic effector cells, but their lack of cytotoxic function is due to a relative refractoriness in initiating the post-binding lytic mechanism.     

Further characterization of cooperative interactions of monoclonal antibodies

We reported previously that mixtures of some monoclonal antibodies directed against the glycoprotein hormone human chorionic gonadotropin (hCG) had a higher affinity for the antigen than either monoclonal antibody separately. The synergistic interaction could no longer be detected when one of the antibodies was replaced with its F(ab) fragment. This cooperative interaction has now been further characterized. One-half of 10 possible pairs prepared from five IgG1 monoclonal antibodies against hCG result in a synergistic interaction. The addition of an IgG2b monoclonal antibody to one of the IgG1 monoclonal antibodies also induces a cooperative interaction, which shows that the effect is not subclass restricted. Cooperative interactions between antibodies are also not restricted to solution conditions; adsorption of one antibody to a solid support appears to increase the cooperative effect. Indeed, one pair of antibodies that failed to bind hCG synergistically in solution did so when one antibody was bound to a solid surface. The liquid phase antibody also has an effect on the specificity of the solid phase antibody. The sensitivity of the solid phase assay system has enabled us to develop a rapid method of determining if two monoclonal antibodies can bind to an antigen simultaneously. A quantitative theoretical model has been devised that successfully predicts the cooperative behavior observed between antibodies and should be useful in devising conditions that result in sensitive solid phase radioimmunoassays.     

Abnormal binding of soluble IgG immune complexes to hepatic nonparenchymal cells of autoimmune mice

Because the liver is the major organ responsible for removal of soluble immune complexes (IC), the surface binding characteristics of preformed model IC to unstimulated mouse liver nonparenchymal cells (NPC) in suspension were studied. NPC of non-autoimmune C3H/FeJ, C3H/HeJ, A/J, DBA/2 and the autoimmune NZB/W F1 and MRL/lpr female mice of various ages were isolated by perfusion of the portal vein with collagenase followed by separation of NPC from hepatocytes with a metrizamide gradient. Thirty-five percent of NPC of all mouse strains were nonspecific esterase-positive and phagocytosed latex beads. Radiolabeled mouse IgG anti-DNP covalently cross-linked stable IC were separated by gel filtration and bound to NPC under various conditions. Marked differences were noted in maximal number of IC bound per cell between the autoimmune and non-autoimmune mouse strains: 3.3 to 4.0 X 10(5) in the non-autoimmune strains vs 0.3 to 1.4 X 10(5) molecules of IC bound per cell in the autoimmune strains at 1 to 6 mo. Insignificant differences were noted in Ka by Scatchard plot analysis (3.5 to 5.0 X 10(8) M-1) and rate of reversibility of binding as determined by dissociation of surface-bound IC with an excess of heat-aggregated gamma-globulin (T 1/2:1.5 to 2 min). These data demonstrate a decreased number of available binding sites for IC in unstimulated NPC from NZB/W F1 and MRL/lpr female mice throughout their life spans. Although the findings are consistent with saturation of binding sites of the NPC with native IC, the abnormality found in the 1-mo-old autoimmune mice (who do not have detectable autoantibodies) suggests a primary defect in FC receptor expression or an altered state of activation of NPC that may contribute to the disease process.     

Autoimmune effector cells. IV. Induction of experimental allergic encephalomyelitis in Lewis rats without adjuvant

Experimental autoimmune encephalomyelitis (EAE) was induced in Lewis rats without the aid of adjuvant. Radioresistant cells were activated from spleens of nonimmune donor rats by in vitro culture with myelin basic protein (BP). After adoptive transfer, these cells recruited EAE effector cells in normal syngeneic recipients, as demonstrated after transfer to secondary hosts. The induction of EAE without adjuvant may lead to a better understanding of the mechanisms of autoimmune disease induction.