Direct membrane effects of morphine and endorphins on Amoeba proteus.

Morphine, leu-enkephalinamide, met-enkephalin, alpha-neoendorphin and its Arg8 1-8 fragment increase contractile vacuole output in the freshwater Amoeba proteus at 18 microM. Significant effects of leu-enkephalin and naloxone are obtained at 180 microM. All compounds have reached their maximal activity at 720 microM. Alpha-neoendorphin and leu-enkephalin are inactive in the presence of isotonic, non-penetration sucrose, hence these compounds increase plasma membrane permeability to water. Results from molecular modeling show a clear correlation of activity with amphiphilicity, charge distribution and general flexibility of molecules. We conclude that, like previously-studied vasopressin analogues and non-hormonal amphiphilic peptides, active opioids embed themselves into the Amoeba plasma membrane, disrupting the lipid bilayer and increasing its permeability. In our Amoeba system, naloxone, a general morphine-like inhibitor, blocks active opioids as well as a vasopressin analogue. Naloxone, being less active than other tested amphiphiles, acts as a membrane stabilizer, protecting the lipid bilayer against the disruption action of more active compounds.     

Immunohistochemical characterization of delta crystallin-containing retina/optic nerve "boundary" cells in the chick embryo

The term "transdifferentiation" has been used to describe the apparent phenotypic conversion of chick embryo neural retina Müller glial cells into lens-like cells in vitro. This phenotypic conversion is characterized by expression of such lens-specific proteins as delta crystallin and has been viewed as an example of cells transforming from the phenotype of a given tissue to that of another. We have identified a population of neuroglia-like cells in the embryonic chick retina which express high levels of delta crystallin as a function of normal development. The position and morphology of these cells is quite distinctive in that they form a loose meshwork which defines the boundary between the neural retina and the optic nerve head. These "boundary" cells are detectable as early as Day 5 of development through hatching. However, the meshwork structure formed by the cells is only readily observed between Days 8 and 9 of development. Double-immunolabeling procedures comparing delta crystallin staining to that of glial and neuronal markers suggest that these cells are a form of retinal Müller glial cell. The results show that under appropriate microenvironmental conditions, expression of delta crystallin falls into the normal repertoire of retinoblast cells. The results also demonstrate the presence of a cellular boundary defining the junction between the neural retina and the optic nerve, tissues that are ontogenetically and structurally continuous but functionally distinct.     

Selective solubilization of xylan in pulp using a purified xylanase fromTrichoderma harzianum

Summary The specificity of action of a cellulase-free xylanase preparation on pulp fibers was revealed by the composition of the solubilized products after enzyme treatment. The neutral carbohydrates released by the treatment of two hardwood kraft pulps were composed exclusively of xylooligomers. A similar treatment of Solka Floc showed no detrimental effect on the degree of polymerization of the cellulose fibers, as determined by size exclusion chromatographic analysis.     

Treatment of canine aspiration pneumonitis: fluid volume reduction vs. fluid volume expansion

The aspiration of gastric acid causes pulmonary edema and hypoxemia. One approach to the management of this syndrome is to raise cardiac output (Qt) and O2 delivery (QO2) to ensure tissue oxygenation (VO2) at the risk of increasing the edema. Another approach reduces the edema by reducing pulmonary microvascular pressure (Pmv) at the risk of reducing QO2 and VO2. We compared these approaches in 24 anesthetized, ventilated dogs with pulmonary wedge pressure (Ppw), a clinical approximation of Pmv, of 12.5 mmHg. Before and again 1 h after endobronchial instillation of 0.1 N HCl, we measured Qt, QO2, VO2, venous admixture, and in vivo extravascular lung liquid. The dogs were then randomly divided into four equal groups: 1) 12.5 mmHg Ppw, high Qt; 2) 7.5 mmHg Ppw, intermediate Qt; 3) 4.5 mmHg Ppw, low Qt; and 4) 4.5 mmHg Ppw plus dopamine, intermediate Qt. Measured values were followed for 4 more h, after which the lungs were excised to compare wet weight-to-body weight ratios (W/B). When plasmapheresis reduced Ppw at 1 h, edema did not increase further and W/B of groups 2 (21 +/- 3), 3 (18 +/- 3), and 4 (22 +/- 3) were significantly less than in group 1 (27 +/- 3) (P less than 0.001). Although Qt decreased with Ppw, increased hematocrit and reduced venous admixture maintained QO2 in group 2 but not in group 3. In group 4 an intermediate Qt maintained QO2 even at 4.5 mmHg Ppw but edema increased to the group 2 level presumably because Pmv rose with Qt on dopamine. VO2 remained constant over time in each group. These data demonstrate that canine HCl-induced pulmonary edema, measured in vivo or gravimetrically, is very sensitive to reductions in Pmv. Moreover, the lowest Pmv (and QO2) was well tolerated because an O2 supply dependency of VO2 was not observed.     

Antibody response of BALB/c mice to dextran B1355S: alterations in the expression of an idiotype associated with the depletion of idiotype-binding cells

In this study, we established that BALB/c mice recognize and respond to the idiotype (M104E IdI) of a major dextran-specific clonotype within the BALB/c mouse repertoire. This idiotype recognition is established by demonstrating the presence of idiotype-binding cells and by the production of antibodies specific for the private M104E idiotype. To determine whether or not the idiotype-recognizing cells play a regulatory role during an immune response to dextran, the idiotype-binding cells were selectively removed either by panning or by radiation-induced killing. Two significant effects are observed when the depleted spleen cells are immunized with dextran. First, there is a substantial increase in the proportion of anti-dextran antibodies that are M104E IdI+. The second effect of the idiotype-specific cell depletion is the production of significant amounts of M104E IdI+ immunoglobulin molecules which do not bind dextran. The depletion experiments produced no alteration in the concentration of anti-dextran antibodies found in the serum or in the number of dextran-specific PFC in the spleen. The data indicate that idiotype-reactive cells can play a role in regulating the level of individual clonotype expression (i.e., the M104E clonotype), but that an alternative mechanism must exist for regulating the absolute amount of anti-dextran antibody produced after immunization.     

Failure of PFC inhibition assays to distinguish idiotypically between clonotypes that are readily distinguishable by RIA analysis

Two different hemolytic plaque assay protocols that are commonly used to quantitate idiotype-positive antibody-secreting cells have been compared to a standard radioimmunoassay (RIA) to test for their ability to discriminate between related, but idiotypically distinct, clonotypes. The idiotype proband used in this analysis is the individual specific idiotype associated with the dextran-binding myeloma protein M104E (M104E IdI). Antibodies specific for this private idiotype (anti-M104E IdI) were purified by a combination of adsorption and affinity chromatography of the immunoglobulin fraction isolated from the sera of rabbits repeatedly immunized with M104E. The same affinity-purified anti-M104E IdI antibodies were used in the hemolytic plaque assays and in the RIA. In one of the plaque assays, the putative idiotype-positive antibody-forming cells were scored by lysis of target erythrocytes to which the anti-idiotype had been covalently coupled. In the other plaque assay, the idiotype-positive cells were determined indirectly by anti-idiotype inhibition of PFC produced on dextran-coupled target erythrocytes. The fidelity of these two assays to quantitate the M104E private idiotype expression in individual BALB/c mice after a single immunization with dextran B-1355S was determined by comparing the plaque assay data to the data generated by a double-antibody, post-precipitation RIA of either the antibodies in the serum or of monoclonal antibodies produced by hybridomas. Our data indicate that both plaque assay protocols reflect an overestimate of the actual expression of M104E private idiotype. By using a library of dextran-specific hybridomas (that have been characterized in an RIA with respect to their M104E IdI cross-reactivity), we have shown that the PFC overestimate of the M104E expression observed in dextran-immune mice is due to the inability of both plaque assay protocols to distinguish between dextran-specific clonotypes that express idiotypes cross-reactive with, but not identical to, the 104E IdI. We conclude that the plaque assay should be used only in conjunction with an RIA to estimate the idiotype expression. This is especially true in situations where closely related cross-reactive idiotype families exist.     

Mechanism of adhesion of Alysiella bovis to glass surfaces

Alysiella bovis adheres to surfaces by means of short, ruthenium red-staining, rod-like fimbriae. The fimbriae remain associated with the cell envelope of A. bovis, even when sonicated or exposed sequentially to toluene, Triton X-100, lysozyme, ribonuclease, and deoxyribonuclease. Adhesion of outer membrane-derived cell wall ghosts of A. bovis to glass was inhibited by IO4-, sodium dodecyl sulfate, urea, pronase, and trypsin. Protease treatment digested the fimbriae from the distal end, and exposure to sodium dodecyl sulfate depolymerized the fimbriae. Exposure of ghosts to 1% sodium dodecyl sulfate preferentially solubilized a 16,500-dalton protein which was subsequently purified by gel filtration and demonstrated to be a glycoprotein (ca. 17% carbohydrate). Antibodies raised against the 16,500-dalton glycoprotein agglutinated whole cells and inhibited adhesion of ghosts to glass.