Localization of Entamoeba histolytica amebopore in amebic liver abscesses in hamsters.

Amebopore was purified from axenically grown trophozoites of the Entamoeba histolytica strain HM1:IMSS. The purification procedure involved Mono Q anion-exchange chromatography and electroelution. Sequence analysis of the final product revealed that amebopore A was completely pure. Polyclonal antibodies against the purified amebopore were obtained from rabbits, and Western blot studies demonstrated their specificity. Sections of experimental, acute (1, 2, 3, and 4 days), amebic liver abscesses produced in hamsters were stained with the anti-amebopore antibody; in all the analyzed stages, amebopore appeared as a constitutively expressed cytoplasmic molecule in trophozoites. No extracellular or hepatocyte-membrane amebopore was found. This study is the first to trace amebopore in an in vivo model of amebic liver abscesses.

Inflammation, complement, ischemia and amoebic survival in acute experimental amoebic liver abscesses in hamsters.

We have examined the role of inflammatory cells, ischemia and serum complement on the development of acute experimental amoebic liver abscess in hamsters (AEALAH). In hamsters made leukopenic by whole body radiation (800 rad) and daily intraperitoneal glycogen injections, the absence of inflammatory cells and liver tissue damage surrounding the parasites resulted in their rapid (24 h) disappearance from the liver, which showed no lesions. Focal liver ischemia, always present in control AEALAH with inflammation and tissue destruction, was reproduced in radiated hamsters by injection of amoebae mixed with Superdex microspheres, but again in the absence of inflammation, amoebae caused no liver damage and disappeared in 24 h. In hamsters made hypocomplementemic by injection of purified cobra venom factor (CVF), amoebae caused AEALA indistinguishable from controls, but in leukopenic + hypocomplementemic hamsters, amoebae were unable to produce lesions and disappeared from the liver in 48 h. We conclude that inflammation and tissue damage are required for the survival of amoebae in AEALAH and for the progression of the experimental disease.

Entamoeba histolytica: localization of a 30-kDa cysteine proteinase using a monoclonal antibody.

We produced a monoclonal antibody against a major cysteine proteinase of 30kDa from trophozoites of Entamoeba histolytica strain HM1:IMSS. The specificity of the monoclonal antibody was confirmed by specific inhibition of azocasein digestion and by electrophoretic analysis, in the presence of sodium dodecyl sulfate or on a substrate gel, of the antigen precipitated by the antibody. Immunofluorescent staining of trophozoites with the monoclonal antibody revealed heterogeneity in the intensity of whole cell fluorescence and subcellular localization of the stain. The latter was also observed in trophozoites, which were stained by conventional immunohistochemical methods, from experimental liver abscesses in hamsters. Ultrastructural analysis showed antigen distributed mainly in clear amorphous zones in the cytoplasm, which were not limited by a visible membrane. Proteinases are translocated from these compartments to phagocytic vacuoles after trophozoites ingest erythrocytes, suggesting that these regions might be a lysosomal equivalent of this primitive eukaryotic cell.

[Role of leukocytes and amebic proteases in experimental testicular necrosis produced in the rat by Entamoeba histolytica]. / Papel de los leucocitos y de las proteinasas amibianas en la necrosis testicular experimental producida en la rata por Entamoeba histolytica.

We examined the participation of polymorphonuclear leucocytes and amebic proteinases upon tissue damage by means of an experimental model of acute amebic lesions developed in rat's testicle. In leukopenic rats (less than 1,000 leucocytes/ml) intratesticular injection of axenic E. histolytica's trophozoites (HM-1) produced lesions undistinguishable from the normal controls. On the other hand, inhibition of 80% (average) of the proteinase activity by means of previous incubation of the trophozoites with human a2M gave way to minimal inflammatory lesions almost undistinguishable from the controls which were injected with PBS-A. Our data suggest that in this experimental model of acute amebiasis polymorphonuclear leukocytes do not participate in the tissue damage and that amebic proteinases are responsible for Entamoeba histolytica's virulence.

[The mechanism of natural resistance in the rat to Entamoeba histolytica]. / Estudios sobre el mecanismo de la resistencia natural de la rata a Entamoeba histolytica.

The data presented in this paper may be summarized as follows: 1) intraportal injection of a virulent strain (HM-1) of Entamoeba histolytica in Wistar rats, of both sexes, gives way to non-progressive microscopic changes, characterized by a rapid leukocytic reaction surrounding the amebas, disappearance of the parasites within 5 hours, and total lack of hepatic damage; 2) leucopenia only modifies the previous description by the fact that there are no leucocytes around the amebas, although these disappear in the same time, showing, at this moment, an early and prominent vacuolar degeneration; 3) hypocomplementaemia shows the same results as leucopenia; 4) fragments and extracts from various tissues from the rat and hamster show variable degrees of interference with the viability of axenic amebas of Entamoeba histolytica conserved in culture; only a minimal part of the interference to be due to the activity of the complement which is present in the tissue extracts.

Increase of biosynthesis and degradation of collagen in normal lungs induced by soluble factors obtained from experimental pulmonary silicosis.

We have studied the effects of soluble factors obtained from rat lungs with experimentally-induced pulmonary fibrosis of 2 months duration on the in vitro rates of biosynthesis and degradation of collagen in normal rat lung preparations. Factors soluble in phosphate-buffered saline were prepared from the minced lungs of normal controls and of silicotic animals. The in vitro rate of collagen biosynthesis of normal rat lung explants was measured as the rate of incorporation of radioactive proline into total and collagenous protein. The in vitro rate of collagen degradation in normal rat lung homogenates was measured as the rate of release of hydroxyproline-containing materials of less than 100,000 daltons to the supernatant. Our results suggest that in this experimental model of pulmonary fibrosis there are soluble factors that stimulate both collagen biosynthesis and collagen degradation in in vitro preparations of normal rat lung.