Endogenous fibronectin of blood polymorphonuclear leukocytes: immunochemical characterization and subcellular localization.

Fibronectin, a large dimeric glycoprotein synthesized and secreted by several cell types, mediates cell adherence to surfaces. In infections and inflammatory responses, blood polymorphonuclear leukocytes (PMNs) adhere to cells and matrix proteins during extravasation and accumulation at inflammatory sites. The presence of fibronectin in blood PMNs has been poorly studied, and the characteristics and subcellular localization of this endogenous adhesive molecule are practically unknown. By immunofluorescence flow cytometry, purified rabbit antibodies and a monoclonal antibody to plasma fibronectin reacted with isolated blood PMNs, only after permeabilization of the cells. By Western blot analysis, the antibodies recognized, under reducing conditions, a protein with an apparent molecular mass of 230 kDa in the cell lysate. Eleven monoclonal antibodies to common frame fibronectin epitopes, including the RGD-containing cell-binding domain, also reacted with PMN fibronectin by Western blotting. In contrast, two antibodies to ED-A, the alternatively spliced region characteristic of "cellular" fibronectin, were unreactive, but recognized platelet fibronectin. On average, 1 million PMNs contained 6.8 ng +/- 1.4 (SD) of fibronectin, as measured by sandwich ELISA. Immunogold labeling and electron microscopy studies indicated localization of most fibronectin in PMN granules. Moreover, double-immunofluorescence and digital image analysis demonstrated colocalization of fibronectin with lactoferrin, a marker of specific (secondary) granules. The results indicate that blood PMNs contain approximately 8000 molecules per cell of intact ED-A-negative fibronectin localized mainly in their specific granules.

Intracellular co-localization of Trypanosoma cruzi and inducible nitric oxide synthase (iNOS): evidence for dual pathway of iNOS induction.

Evidence is presented from studies in vitro and in vivo for a dual pathway of inducible nitric oxide synthase (iNOS) induction during Trypanosoma cruzi infection, one of which is interferon (IFN)-gamma dependent and the other not. In vitro, the IFN-gamma-dependent iNOS induction decreases parasite multiplication, and is in vivo associated with protection. iNOS induced by this pathway mediated a high NO output and showed a diffuse, cytoplasmic immunostaining in IFN-gamma-activated macrophages in vitro as well as in cell infiltrates or infected tissues. Surprisingly, in such tissues, iNOS co-localized with parasite nests, and by immunoelectromicroscopy, iNOS was demonstrated on the parasite surface. iNOS co-localization with parasites was also seen in tissues from T. cruzi-infected IFN-gamma receptor (R) knockout mice suggesting an IFN-gamma-independent pathway of induction. However, no cytoplasmic iNOS was seen in inflammatory infiltrates of these tissues. IFN-gammaR(-/-) mice displayed a dramatically enhanced susceptibility to infection with T. cruzi, diminished accumulation of iNOS mRNA in skeletal muscle and spleen cells, and reduced release of NO and peroxynitrite. Expression of iNOS around intracellular parasites was also observed after infection of peritoneal macrophages or L-929 fibroblasts in vitro in the absence of other exogenous stimuli. A time-dependent NO release and enhanced accumulation of iNOS mRNA also was observed in infected peritoneal cells and fibroblasts. Cultured T. cruzi amastigotes, trypomastigotes, and epimastigotes were not labeled by the anti-iNOS antibodies and contained no iNOS mRNA, indicating that the iNOS detected actually originated from the mammalian cell. A pathogenic effect of low NO levels is suggested by the arresting effect of NOS inhibitors and the enhancing consequences of low concentrations of NO donors on intracellular parasite multiplication.

[Evaluation of the incidence and toxicity of samples of Bacillus cereus in various classes of foods sold and consumed in the State of Rio de Janeiro]. / Avaliacão da incidência e da toxicidade de amostras de Bacillus cereus em diferentes classes de alimentos comercializados e consumidos no Estado do Rio de Janeiro.

One hundred and fourteen strains of Bacillus cereus were isolated during presumptive plate-counts from 18 groups of industrialized, non-industrialized, crude or cooked food, belonging to 10 separate classes. Specific presumptive counts ranged from 10(2) to 6 X 10(3)/g or ml. Among these isolates, 13 strains were derived from 3 outbreaks of food poisoning (involving a minimum of 57 people), as determined by the assayed bacteriological quality of the ingested foods. As an adopted procedure to correlate toxicity and ability to promote illness in man, culture fluids of all strains were assayed to determine their ability to increase vascular permeability (APC) to cause necrosis in rabbits skin and to kill albino mice. APC was positive in 86.85% of the 114 strains, death of albino mice occurred in 65.79% and a combination of APC and death was observed in 59.65%. APC plus necrosis, or only necrosis, occurred with 34.21% of the culture fluids. Death, APC and death with or without necrosis, were demonstrated in the strains implicated with illness. This confirmed the known individuality of action exhibited by certain B. cereus food-borne toxigenic factors. The low presumptive counts of this bacterium in the order of 10(2)-10(3)/g or ml found in food, implicated or not with illness, suggests that the recommended number of B. cereus per g or ml of food sample should be reevaluated in our country. Furthermore, a wider range of food should be brought under bacteriological sanitary control for this species.

Local effects of silica on tumor growth inhibition. A histological study.

The subcutaneous implant of a fibrosarcoma covered with silica, in rats, resulted in a temporary retardation of tumor growth. The histological study demonstrated that the silica provoked, within and at the periphery of the tumor, inflammatory foci which altered the composition of the fundamental substance and which promoted the accumulation of principally macrophages in the interior as well as on the surface of the tumoral mass. Besides the macrophages of the host, a number of tumor cells do also phagocyte silica crystals, leading to destruction of all those cells. The authors are of the opinion that the local modifications provoked by silica, could have interfered in the adaptation of the tumor with its nutrient stroma. Of these modifications, the stroma sclerosis seems to be the most effective in the inhibition of tumor growth. Some correlations were suggested between the here described results and those obtained by other workers who have used other inflammatory agents of different nature.

Growth inhibition of sarcoma 180 induced by silica and talc.

In applying amorphous silica and talc in the site of solid Sarcoma 180 implants, the authors found a suppression and delay of tumor growth which was proveked by peritumoral foreign-body granulomas. Correlating these findings which those experiments which resulted in granulomas of hypersensitivity type, it may be suggested that any kind of peritumoral chronic inflammation enables an anti-tumor effect.