In vivo exocytosis of lysosomes by the endothelium of the venous sinuses of bone marrow and liver: visualization at normal and low body temperature

We have visualized the exocytosis of lysosomes into the peripheral circulation by the phagocytic endothelia of the venous sinuses of liver and bone marrow of rats. Perfusion fixation at normal body temperature produced images of the earliest stages of lysosomal exocytosis. After fixation at low body temperatures (7-12 degrees C), advanced stages of this process became evident, showing extrusion of lysosomes and their contents into the circulation. It is postulated that this form of exocytosis has escaped structural detection because of its rapidity and relative infrequency as compared to merocrine secretory exocytosis, and that fixation at low body temperatures arrests or slows down these exocytic events in sufficient measure for ultrastructural visualization. The possibility that this lysosomal exocytosis contributes to the presence of lysosomal enzymes detected in the peripheral blood should be considered. In addition, it is likely that lysosomal degradation products may be discharged by exocytosis into the circulation.     

Mycological survey of Korean cereals and production of mycotoxins by Fusarium isolates.

The fungal species isolated from Korean cereals (barley, polished barley, wheat, rye, and malt) were Alternaria spp., Aspergillus spp., Chaetomium spp., Drechslera spp., Epicoccum sp., Fusarium spp., and Penicillium spp., etc. The number of Fusarium strains isolated was 36, and their ability to produce Fusarium mycotoxins on rice was tested. Nivalenol (NIV) was produced by Fusarium graminearum (7 of 9 isolates), Fusarium oxysporum (3 of 10 isolates), and Fusarium spp. (7 of 15 isolates). Of 15 isolates of Fusarium spp., 6 formed deoxynivalenol (DON). Fusarenon-X and 3-acetyl-DON were produced by most NIV- and DON-forming isolates, respectively. Zearalenone was produced by 3 isolates of F. graminearum, 1 isolate of Fusarium equiseti, and 11 isolates of Fusarium spp. T-2 toxin was not produced by any Fusarium isolates. The highest concentrations of mycotoxins produced by Fusarium isolates were 77.4 (NIV), 5.3 (DON), 138.3 (fusarenon-X), 40.6 (3-acetyl-DON), and 23.2 (zearalenone) micrograms/g.     

Measurement and prediction of flow through a replica segment of a mildly atherosclerotic coronary artery of man

Pressure distributions were measured along a hollow vascular axisymmetric replica of a segment of the left circumflex coronary artery of man with mildly atherosclerotic diffuse disease. A large range of physiological Reynolds numbers from about 60 to 500, including hyperemic response, was spanned in the flow investigation using a fluid simulating blood kinematic viscosity. Predicted pressure distributions from the numerical solution of the Navier-Stokes equations were similar in trend and magnitude to the measurements. Large variations in the predicted velocity profiles occurred along the lumen. The influence of the smaller scale multiple flow obstacles along the wall (lesion variations) led to sharp spikes in the predicted wall shear stresses. Reynolds number similarity was discussed, and estimates of what time averaged in vivo pressure drop and shear stress might be were given for a vessel segment.     

Multifaceted freezing injury in human polymorphonuclear cells at high subfreezing temperatures

Extracellular freezing injury at high subzero temperatures in human polymorphonuclear cells (PMNs) was studied with a cryomicroscope, electron microscope, and functional assays (phagocytosis, microbicidal activity, and chemotaxis). There are at least four major factors in freezing injury: osmotic stress, chilling, cold shock, and dilution shock. Extracellularly frozen PMNs lose functions when cooled to -2 degrees C without a cryoprotectant. Cells lose volume on freezing to the same degree as in hypertonic exposure. PMNs have a minimum volume to which they can shrink without injury. Greater dehydration produces irreversible injury to cellular functions, and cells eventually collapse under high osmotic stress. Chilling sensitivity is seen in slowly chilled, supercooled PMNs below -5 degrees C; at -7 degrees C, functions are lost in 1 h. This injury can be prevented by the addition of Me2SO but not glycerol. Me2SO does not, however, prevent cold shock (injury due to rapid cooling), which is seen during cooling at 10 degrees C/min to -14 degrees C, but not during slow cooling at 0.5 degrees C/min. One of the problems of using glycerol as a cryoprotectant stems from the high sensitivity of PMNs to dilution shock during the dilution or removal of glycerol.     

The amino acid transport systems of the autotrophically grown green alga Chlorella

The kinetic analysis of l-amino acid uptake by the green alga Chlorella revealed at least seven different uptake systems to be present in cells grown autotrophically with nitrate as nitrogen source. There is a ‘general system’ which transports most neutral and acidic amino acids, a system for short-chain neutral amino acids including proline, a system for basic amino acids including histidine, a special system for acidic amino acids, and specific systems for methionine, glutamine and threonine. The ‘general system’ is possibly the same as that which can be stimulated by incubation of cells in glucose plus ammonium (Sauer, N. (1984) Planta 161, 425–431). The incubation of Chlorella in glucose induces the increased synthesis of six amino acid uptake systems, namely the above-mentioned system for short-chain neutral amino acids, a threonine system, a methionine system, and a glutamine system. These results indicate that the uptake of l-amino acids by the green alga Chlorella is as complex as in other free-living organisms such as bacteria or yeast. The small number of amino acid uptake systems found in cells of higher plants, i.e. two or three, seems therefore to be a consequence of integration of the cells in a tissue supplying a relatively constant environment, and not a consequence of autotrophic growth on mineral carbon and mineral nitrogen.     

Inhibition of virus multiplication by immunoactive peptides

In the present study we show that peritoneal macrophages obtained from the mice treated with the immunoactive peptides inhibit the multiplication of Herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2), but not that of vesicular stomatitis virus (VSV), and that the intraperitoneal administration of the peptides suppresses the infection with HSV-1 in mice.     

The fermentation process for l-phenylalanine production using an auxotrophic regulatory mutant of Escherichia coli

Summary A process for l-phenylalanine production was studied using a tyrosine auxotrophic regulatory mutant of Escherichia coli, resistant to both -2-thienyl-dl-alanine and p-fluoro-dl-phenylalanine. Fermentations were carried out in a 30-1 fermentor with intermittent feeding of glucose plus phosphate. The mutant accumulated l-phenylalanine in the fermentation broth up to 15 g/l at pH 7.0 and 33°C. Column chromatography on a strong cation exchanger was employed as the most effective step in the purification of l-phenyl-alanine from the broth. This step brought about 4-fold concentration of the product with 96% recovery.     

Serological survey of T-lymphocyte differentiation antigens in wild mice

Allelic distributions of Thy-1, Ly-l, and Ly-2 antigens in wild mice are characteristic of each Mus musculus subspecies. Eastern mice (M.m.molossinus, M.mmusculus, M.m.castaneus, M.m.bactrianus) express the Thy-1.1 antigen, whereas Western mice (M.m. domesticus, M.m.brevirostris) express the Thy-1.2. All mice from wild populations examined in this survey express the Ly-1.2. The Ly-2.1 is distributed in Eastern mice and some Western mice, and the Ly-2.2 is found in the remaining Western mice. Allelic distributions of these antigens were also examined in two other species, Mus spretus and Mus spicilegus. Allelic constitutions of Thy-1 and Ly-1 in these species are similar to those of Eastern mice. Some M.spicilegus, however, express the Ly-1.1 antigen. This antigenic type is not found in M.musculus. Some Eastern mice related to M.m.castaneus react weakly to Ly-1.2-specific and Ly-2.1-specific monoclonal antibodies in both the complement-mediated cytotoxicity test and the absorption test. These results suggest that M.m.castaneus has unique alleles in the Ly-1 and Ly-2 loci.     

Production of human c-myc protein in insect cells infected with a baculovirus expression vector.

A cDNA fragment coding for human c-myc was inserted into the genome of the baculovirus Autographa californica nuclear polyhedrosis virus adjacent to the strong polyhedrin promoter. Insect cells infected with the recombinant virus produced significant amounts of c-myc protein, which constituted the major phosphoprotein component in these cells. By immunoprecipitation and immunoblot analysis, two proteins of 61 and 64 kilodaltons were detected with c-myc-specific antisera. The insect-derived proteins were compared with recombinant human c-myc-encoded proteins synthesized in Escherichia coli and Saccharomyces cerevisiae cells. The c-myc gene product was found predominantly in the nucleus by subcellular fractionation of infected insect cells.     

Immobilization of enzymes in porous supports: Effects of support-enzyme solution contacting

Proteins have been immobilized in porous support particles held in a fixed-bed reactor through which protein solution is continuously circulated. Changing the recirculation flow rate alters the observed immobilization kinetics and the maximum enzyme loading which can be achieved for glucose oxidase and glucoamylase on carbodiimide-treated activated carbon and for glucoamylase immobilized on CNBr-Sepharose 4B. Direct microscopic examination of FITC-labelled protein in sectioned Sepharose particles and indirect activity-loading studies with activated carbon-enzyme conjugates all indicate that immobilized enzyme is increasingly localized near the outer surface of the support particles at larger recirculation flow rates. Restricted diffusion of enzymes may be implicated in this phenomenon. These contacting effects may be significant considerations in the scaleup of processes for protein impregnation in porous supports, since apparent activity and stability of the final preparation depend on internal protein distribution.