Effects of leptospiral lipopolysaccharide on rabbit platelets.

Leptospiral lipopolysaccharides (LPSs) extracted from Leptospira interrogans serovar copenhageni virulent strain Shibaura, serovar canicola virulent strain Moulton, and serovar hebdomadis strain Hebdomadis, were tested for their ability to induce platelet aggregation and/or lysis in rabbit platelet-rich plasma (PRP). All showed positive reactions with a release of adenosine triphosphate (ATP) and serotonin. The values, however, were different from each other. The ability of leptospiral LPS extracted from serovar copenhageni virulent strain Shibaura (I-LPS) to induce platelet aggregation was the highest of all. After treatment of I-LPS, the platelets developed a ruffled surface with appearance of pseudopodia as observed by transmission electron microscopy (TEM) and scanning electron microscopy (SEM). I-LPS also showed cytotoxicity for the platelets. Degenerative or lytic changes were recognized in 44.5% of the platelets which were observed 60 min after I-LPS treatment.

Lack of effectiveness of ofloxacin against experimental syphilis in rabbits.

Ofloxacin, a new pyridone-carboxylic acid derivative, was evaluated in experimental syphilis in rabbits in comparison with penicillin G. Experimental syphilis was established by intradermal injection of Treponema pallidum subsp. pallidum Nichols. Ten days after infection, the dermal lesions were characterized by syphilitic papula accompanied with central necrosis. These animals were subsequently treated either with ofloxacin twice a day at an oral dose of 10 mg/kg or with penicillin G once a day at an intramuscular dose of 10,000 U/kg for 21 consecutive days. In penicillin G-treated animals, the dermal lesions became smaller as early as day 3 of treatment and almost disappeared during the therapy. In marked contrast to remarkable efficacy of penicillin G was further development of the lesions in ofloxacin-treated animals, showing no difference in pathological manifestations as compared to untreated animals. The results of nontreponemal serologic test correlated well with the response of animals to treatment.

Chemotherapeutic efficacy of ofloxacin on renal and subcutaneous infection models with Staphylococcus aureus in mice.

Ofloxacin, a new pyridone-carboxylic acid derivative, was evaluated in descending nephritis and subcutaneous abscess models with Staphylococcus aureus in mice in comparison with norfloxacin. Descending nephritis was produced by intravenous injection of S. aureus 39 (MIC 0.78 microgram/ml for ofloxacin and 3.13 micrograms/ml for norfloxacin). Subcutaneous abscess was established by subcutaneous injection of soft agar containing S. aureus 56230 (MIC 0.39 microgram/ml for ofloxacin and 1.56 micrograms/ml for norfloxacin). Three days after infection, the lesions of both models were characterized by purulent inflammation accompanied with massive infiltration of neutrophils and bacterial multiplication. The animals were treated twice a day orally with each compound for 4 consecutive days, and subjected to bacteriological examination 18 h later. In the renal model, the 50% effective doses calculated on the basis of clearance of bacteria from kidneys were 38.4 mg/kg for ofloxacin and greater than 100 mg/kg for norfloxacin. In the subcutaneous model, the 50% effective doses based upon 90% reduction of viable bacteria as compared with untreated controls were 25.2 mg/kg for ofloxacin and greater than 100 mg/kg for norfloxacin. The excellent efficacies of ofloxacin in both infection models are attributed to its high oral absorbability and tissue distribution.

Preparation of monoclonal antibody to hepatocellular membranes and its application to induction of liver cell membrane damage.

Monoclonal antibody (MoAb) to rat liver plasma membranes was prepared by hybridization of mouse immune lymphocytes with mouse myeloma cells, and was identified by the immunodiffusion method in a fraction of IgM secreted from the hybridoma thus obtained. In indirect immunofluorescence tests, specific fluorescence was detected only on the surface of rat hepatocytes, but neither on the cells from other organs of the rat nor on the hepatocytes of other species of animals, suggesting that the antibody may be organ- and species-specific. When the primary culture rat hepatocytes, labelled with isotopic chromium (51Cr), were treated with the MoAb together with complement, a specific release of 51Cr from the cells was found shortly after treatment, accompanied with bubbling of the cell membranes, and a significant release of 51Cr was observed at an MoAb concentration of 15 micrograms/ml or more. Without complement, or with inactivated complement, these reactions were not observed. These facts suggest strongly that the cell surface of the hepatocytes was damaged by the MoAb in the presence of complement.

Action of malotilate on reduced serum cholesterol level in rats with carbon tetrachloride-induced liver damage.

The mode of action of malotilate in normalizing serum cholesterol in hypocholesterolemic rats with fatty liver was examined by determination of biosynthesis, catabolism and excretion of cholesterol. Fatty liver was produced by subcutaneous injection of CCl4 at the dose of 1 ml/kg into male rats (SLC-SD) twice a week for 3 weeks. Daily administration of malotilate (100 mg/kg) in rats with hypocholesterolemia resulted in a rapid normalization of lowered serum cholesterol. Such a recovery of cholesterol level in serum coincided in time with normalization of the decreased cholesterol level of each lipoprotein fraction, VLDL-triglycerides secretion and the decreased apolipoprotein A1 value. Histopathological improvement in liver was also confirmed by a decrease in the size of fat droplets stored within the hepatocytes. The malotilate treatment gave a tendency to facilitate hepatic cholesterol synthesis in rats with fatty liver. Malotilate at a concentration of 0.5-2 micrograms/ml also stimulated cholesterol biosynthesis in cultured normal hepatocytes. The drug had the action to accelerate the catabolic excretion of 3H-labeled cholesterol into feces. These results suggest that the mode of action by which serum cholesterol is normalized in rats with fatty liver is probably due to a stimulative effect of malotilate on hepatic cholesterol synthesis and cholesterol secretion from the liver.

Stimulation by staphylococcal enterotoxin A of nonspecific resistance of mice to microbial infection.

The effects of staphylococcal enterotoxins A, B, and C2 (SEA, SEB, and SEC2) on the resistance of mice to microbial infections were studied. SEA stimulated the resistance strongly, whereas SEB and SEC2 had no such effect. Treatment with SEA increased the number of peripheral blood polymorphonuclear leukocytes significantly within 4 h, and these polymorphonuclear leukocytes exhibited a higher chemiluminescence response than those of the controls. Furthermore, a significant increase in spleen weight was also observed in mice treated with SEA, and histologically that increase was characterized by a proliferation of lymphoblast-like cells which were stained with antibody to mouse Thy-1 but not with antibody to mouse immunoglobulin G by indirect immunofluorescence. As expected from the above findings, the treatment of nude mice (nu/nu) with SEA failed to protect them against Escherichia coli infections, whereas treatment of heterozygous (nu/+) controls afforded such protection. This was in part supported by the fact that the chemiluminescence response of peripheral blood polymorphonuclear leukocytes was increased significantly by treatment with SEA in nu/+mice but not in nu/nu mice.

Nonbacteremic pseudomonas pneumonia in immunosuppressed guinea pigs.

An experimental model of nonbacteremic pneumonia with a virulent strain of Pseudomonas aeruginosa was successfully established in guinea pigs immuno-suppressed with cortisone acetate although the organisms were eliminated rapidly from the lungs without cortisone treatment. Using a pocket nebulizer, almost all the animals which received 10(6) organisms/g-lung developed bronchopneumonia without any septic findings as long as 10 days after challenge. The lesions produced in such animals were characterized by dissemination of multiple purulogranulomatous changes. In the early stage of infection, infiltration of polymorphonuclear cells (PMNs) in the bronchiolar and alveolar spaces was diffuse, later showing multifocal accumulation with the formation of central spherical grains enclosing bacterial colonies. In the later stage, granulation tissue consisting of large mononuclear cells, fibroblasts and collagen fibers developed around the PMN accumulation. The animals which received 10(7) organisms/g-lung, on the other hand, developed severe pulmonary hemorrhages and necrosis followed by septic death.

Acute demyelination in mice inoculated intraspinally with mouse hepatitis virus, JHM strain.

After intracerebral or intralumbar inoculation of mouse hepatitis virus, JHM strain, into weanling mice, viral growth and lesions in the central nervous system were comparatively studied for 5 days postinoculation. In the intralumbar group the virus titer of the spinal cord exceeded that of the brain until 24 hr postinoculation, and declined later. In the intracerebral group the brain virus titer was always higher, and the cord titer attained the same level as that of intralumbar group at 72 hr. Demyelinating lesions in the spinal cord appeared 48 hr after intralumbar inoculation, that is, 36 hr earlier than after intracerebral inoculation. Virus-specific fluorescence was detected within glia cells in the cord white matter 48 hr intralumbar inoculation. Electron microscopy revealed a number of virions within oligodendroglia cells, suggesting that the acute phase demyelination might be due to viral growth within this type of cells.

Giant cell formation in the brain of suckling mice infected with mouse hepatitis virus, JHM strain.

After intranasal inoculation with a neurotropic mouse hepatitis virus, JHM strain, suckling mice less than 2 weeks of age died in 4 or 5 days postinoculation, while those 3 weeks or more of age survived. In the brain lesions of mice 8 days or less of age, there appeared two different types of multinucleated giant cells. One of them which was irregular in shape with many virus particles, appeared mostly in the subependymal or perivascular areas after 72 h postinoculation and it seemed to have resulted from a fusion of two or more infected macrophages. The other type of giant cells which was oval in shape with a small number of virus particles, appeared within degenerative lesions at 96 h postinoculation. It consisted of two morphologically different parts and seemed to be a complex of infected neurons and reacting macrophages. In the cerebral lesions of mice 2 to 4 weeks of age, only the former type of giant cells appeared.

Vertical transmission of mouse hepatitis virus infection in mice.

Transplacental infection with mouse hepatitis virus, JHM strain was studied by intravenous inoculation of pregnant dams. Inoculation on day 9 or 12 of gestation brought about the death of more than 50% of the fetuses at 4 days postinfection while inoculation on day 6 or 15 of gestation effected the death of 12% of fetuses or neonates. Inoculation of day 12 of gestation resulted in markedly higher virus titers. At 72 h postinfection in the placentas, fetal membranes and fetuses than in the maternal livers and blood. Virus-specific antigen and virus particles were noted in the placentas, visceral yolk sac and fetal livers by immunofluorescence and electron microscopy. Histopathology revealed degenerative and necrotic changes in these tissues and in the fetal bone marrow.