RESUMO
Entamoeba histolytica is a parasite which presents capacity to degrade tissues and therefore has a pathogenic behavior. As this behavior is not shown by all strains, there have been several studies investigating molecular basis of the cytotoxicity process. Using the suppression subtractive hybridization (SSH) technique, differential gene expressions of two E. histolytica strains, one virulent (EGG) and one nonvirulent (452), have been analyzed with the purpose of isolating genes which may be involved with amoebic virulence. Nine cDNA fragments presenting high homology with E. histolytica previously sequenced genes were subtracted. Of these, four genes were confirmed by RT-PCR. Two coding for hypothetical proteins, one for a cysteine-rich protein, expressed only in the virulent strain, EGG and another one, coding for grainin 2 protein, exclusive from 452 strain. This study provided new insight into the proteins differences in the virulent and nonvirulent E. histolytica strains. We believe that further studies with these proteins may prove association of them with tissue damage, providing new perceptions to improve treatment or diagnosis of the invasive disease.
Assuntos
Entamoeba histolytica/genética , Entamoeba histolytica/patogenicidade , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Técnicas de Hibridização Subtrativa/métodos , Animais , Células CHO , Cricetinae , Cricetulus , Efeito Citopatogênico Viral/genética , Eletroforese em Gel de Ágar , Eletroforese em Gel de Poliacrilamida , Humanos , Fígado/parasitologia , Masculino , Trofozoítos/fisiologia , Virulência/genéticaRESUMO
Entamoeba histolytica is a protozoan parasite that presents a risk to the health of millions of people worldwide. Due to the existence of different clinical forms caused by the parasite and also different virulence levels presented by one strain, one would expect differences in the profile of gene transcripts between virulent and nonvirulent cultures. In this study we used the differential display to select gene segments related to invasiveness of amoeba. One Brazilian strain of E. histolytica in two conditions, able or not to cause lesions in experimental animals, was used. RNA from this strain, was used to study the differential expression of genes. 29 specific gene fragments differentially expressed in the virulent strain were selected. By real-time PCR, six of these genes had confirmed their differential expression in the virulent culture. These genes may have important roles in triggering invasive amoebiasis and may be related to adaptation of trophozoites to difficulties encountered during colonization of the intestinal epithelium and liver tissue. Future studies with these genes may elucidate its actual role in tissue invasion by E. histolytica generating new pathways for diagnosis and treatment of amoebiasis.
Assuntos
Entamoeba histolytica/metabolismo , Entamebíase/metabolismo , Regulação da Expressão Gênica , RNA de Protozoário/biossíntese , Animais , Entamoeba histolytica/genética , Entamoeba histolytica/patogenicidade , Entamebíase/genética , Entamebíase/terapia , Humanos , Camundongos , RNA de Protozoário/genética , RatosRESUMO
Cysteine proteinase (CP) activity and CP5 mRNA levels were analyzed in eleven samples of Entamoeba histolytica isolated from patients presenting different clinical profiles. The virulence degree of the isolates, determined in hamster liver, correlated well with the clinical form of the patient and culture conditions. CP5 mRNA levels were also determined in sample freshly picked up directly from liver amoebic abscess. Differences were not observed in the levels of CP5 mRNA and CP specific activity among the cultured samples. However, different levels of CP5 mRNA were observed in trophozoite freshly isolated from hepatic amoebic lesions. These results reinforce the importance of CP5 for the virulence of amoebae and the need for studies with the parasite present in lesions to validate mechanisms involved in pathogenesis of amoebiasis.
