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Biomedical subjects

E Orozco

Publications and source records attributed to E Orozco.

At least 109 records · Page 6Linked to original sources
PubMed1987

Entamoeba histolytica: cytopathogenicity and lectin activity of avirulent mutants.

Three clones of Entamoeba histolytica (L-6, C93, C919) were isolated by mutagenesis with ethyl methanesulfonate from the axenic strain HM1:IMSS and were studied for adherence, cytolytic, and soluble galactose inhibitable lectin activity. Avirulent clones adhered to and killed fewer Chinese hamster ovary cells than HM1:IMSS (P less than 0.01). However, only C919 was deficient in adherence to red blood cells. Galactose (1.0 g) completely inhibited adherence of all the mutants to Chinese hamster ovary cells; however, adherence to erythrocytes was only partially inhibitable by galactose. Avirulent mutants were more susceptible to being killed by human neutrophils in vitro (P less than 0.01 compared to HM1:IMSS). Soluble protein preparations from all the avirulent mutants were markedly less mitogenic for human lymphocytes and had lower lectin activity for Chinese hamster ovary cells compared to the HM1:IMSS wild type (P less than 0.01 for each activity with each mutant). Indirect immunofluorescence with a monoclonal antibody (F-14) that recognizes the Gal/GalNAc lectin was positive for L-6 and C919. These findings utilizing avirulent mutants of E. histolytica further support a role for the amebic galactose inhibitable lectin in the in vivo pathogenesis of amebiasis.

Animals↗
PubMed1987

Localization and identification of an Entamoeba histolytica adhesin.

The adherence of Entamoeba histolytica trophozoites to target cells was studied by using monoclonal antibodies (MAbs) and adhesion-deficient mutants of the parasite. MAbs Adh-1 and Adh-2 reacted with a surface protein of approximately 112 kDa of the total proteins of trophozoites from the wild type strain, clone A, strain HM1:IMSS. Both MAbs reacted weakly with the adhesion-deficient mutant clones, C-98, C-919 and C-923, all derived from HM1:IMSS. MAbs Adh-1 and Adh-2 incubated with trophozoites from clone A inhibited adherence to red blood cells, erythrophagocytosis and cytopathic effect on cell culture monolayers. Antibodies against a approximately 112 kDa polypeptide were found in the sera from patients with hepatic abscess. These results demonstrate that the adherence of trophozoites to target cells is a necessary event in order for cytopathogenicity to occur.

Adhesiveness↗
PubMed1986

Isolation and characterization of phagocytosis- and virulence-deficient mutants of Entamoeba histolytica.

We have used chemical mutagenesis to isolate 10 independent phagocytosis-deficient mutants of Entamoeba histolytica from a highly virulent clone (strain HM1:IMSS). Phagocytosis-deficient mutants were characterized by several biologic properties related to amoebic pathogenic mechanisms: virulence, adhesion to red blood cells, cytopathic effect of live trophozoites on cell culture monolayers, ability of amoebic extracts to destroy cell culture monolayers, and growth in semisolid agar. All of these mutants were also deficient in in vivo virulence and in vitro cytopathic effect. Three were also deficient in adhesion, and the extracts of three others were unable to destroy cell culture monolayers. These findings suggest a strong correlation between phagocytosis and in vivo and in vitro cytopathogenicity. Both adherence of trophozoites to target cells and toxin activity of crude extracts of amoebae seem to be necessary factors for the expression of the virulence of this parasite. Traits responsible for growth in semisolid agar do not seem to be involved in virulence.

Cell Adhesion↗
PubMed1985

Isolation and characterization of Entamoeba histolytica mutants resistant to emetine.

Isolation of emetine-resistant mutants of Entamoeba histolytica is described. Spontaneous rate of mutation, obtained from a Luria-Delbrück fluctuation test was 2.5 X 10(-7). The mutagenesis frequency obtained from the number of colonies grown in the presence of emetine, divided by the number of viable trophozoites inoculated in semisolid agar was less than or equal to 10(-7) and it was increased from 10 to 100 fold with ethyl methanesulfonate. Two types of clones were isolated: one was cross-resistant to colchicine and the other was not, indicating that mutation took place at different loci. Protein synthesis in the presence of emetine remained unaltered for colchicine-sensitive mutants, suggesting that the molecular lesion lay in the protein synthesis machinery. Colchicine-resistant mutants showed a lower level of [35S]methionine incorporation, probably due to alterations in the molecular transport of some substances across the membrane. This mutant clone was also deficient in red blood cell adhesion, suggesting membrane alterations.

Animals↗
PubMed1985

Structural bases of the cytolytic mechanisms of Entamoeba histolytica.

The cellular bases of the powerful cytolytic activity of the human protozoan parasite Entamoeba histolytica were explored by studying the effect of the virulent strain HM1:IMSS on epithelial monolayers of MDCK cells using a combination of time-lapse microcinematography and transmission and scanning electron microscopy. Early alterations of the epithelial cell membranes were detected by measuring changes in the transepithelial electrical resistance of MDCK monolayers mounted in Ussing chambers. The aggressive mechanism of E. histolytica trophozoites was found to be a complex, multifactorial phenomenon that included hit-and-run damage to the plasma membrane of effector cells mediated through contact, phagocytosis of lysed or apparently intact, but detached, MDCK cells, and intracellular degradation of ingested cells. Following contact with amebas, the epithelial monolayers showed a pronounced lowering of transepithelial resistance, opening of tight junctions, distortion of microvilli, surface blebbing, and the presence of minute focal discontinuities in the plasma membrane. There was no evidence of amebic exocytosis, membrane fusion, or junction formation between the parasite and host plasma membranes. Although modifications in the epithelial cell membranes usually preceded lysis, the cytolytic activity of the parasite did not exclusively involve damage to the plasma membrane of the cultured host cells but also was mediated by avid phagocytosis, the displacement and separation of neighboring cells by means of pseudopodial activity, and the "pinching-off" of the peripheral cytoplasm of epithelial cells.

Animals↗
PubMed1983

Entamoeba histolytica. Phagocytosis as a virulence factor.

In this paper, we attempted to define the role of phagocytosis in the virulence of Entamoeba histolytica. We have isolated, from a highly phagocytic and virulent strain, a clone deficient in phagocytosis. Trophozoites of wild-type strain HM1:IMSS were fed with Escherichia coli strain CR34-Thy- grown on 5-bromo,2'-deoxyuridine. The trophozoites that had incorporated the base analog through phagocytosis of the bacteria were killed by irradiation with 310 nm light. The survivors, presumably trophozoites defective in phagocytosis, were grown until log phase and submitted two more times to the selection procedure. Clone L-6, isolated from a subpopulation resulting from this selection procedure, showed 75-85% less erythrophagocytic activity than the wild-type strain. The virulence of clone L-6 and strain HM1:IMSS was measured. The inoculum required to induce liver abscesses in 50% of the newborn hamsters inoculated (AD50) of HM1:IMSS was 1.5 X 10(4) trophozoites. Clone L-6 trophozoites failed to induce liver abscesses in newborn hamsters even with inocula of 5 X 10(5) trophozoites. Virulence revertants were obtained by successive passage of L-6 trophozoites through the liver of young hamsters. The trophozoites that recovered the ability to produce liver abscesses simultaneously recuperate high erythrophagocytic rates. These results show that phagocytosis is involved in the aggressive mechanisms of E. histolytica.

Animals↗
PubMed1978

[An in vitro model for the quantitative study of E. histolytica virulence].

An experimental model for the quantitative study of the cytopathic effect of E. histolytica is described. Confluent cultures of the MDCK cell line, which form a monolayer with morphological and electrophysiological features of an epithelium were used. Trophozoites of the strains HM1:IMSS, HM2:IMSS and HM3:IMSS, axenic produced variable degrees of cytopathic effects of MDCK cultures each one characteristic for a given strain. The degree of damage induced by the different strains of amebas was measured quantitatively. The results demonstrate that the degree of the cytopathic effect produced in vitro by the various strains tested correlates with the virulence shown in vivo by the same strains, judged as the size of the inoculum required to induce liver cytopathic effect in newborn hamsters.

Animals↗