Growth inhibition of Entamoeba histolytica by rat colon components.

Susceptibility to invasive amebiasis has been suggested to be due to intrinsic amebic factors and/or to such host factors as intestinal microflora, mucus and colonic redox potential. We investigated the effect of rat colon components on the growth of axenically cultured E. histolytica trophozoites. Extracts of rat colon tissue produced a 57% amebic growth inhibition. The main growth inhibitory components were precipitated by 65% ammonium sulfate and were heat-sensitive. These components were partially separated by ultrafiltration and gel filtration chromatography. Thus, we found colonic components (Mr 50-100 kDa) that produced strong growth inhibition (75%). These results suggest that rat colonic products may play an active role in resistance to amebic infection.

Purification and partial characterization of an hemolytic activity from Entamoeba histolytica.

It is generally accepted that hydrolytic and cytolytic amebic components are involved in the pathogenic mechanisms of E. histolytica. We have now identified a lytic activity in two membrane proteins of 23.5 kDa and 25 kDa, which are able to lyse rat erythrocytes. The activity was purified from total homogenates of the virulent strains HM1:IMSS and HM38:IMSS, and the erythrocyte lysis was directly related to protein concentration. The hemolytic activity was heat-sensitive and resistant to reduction by 2-mercaptoethanol. Total amino acid analysis of pure proteins showed a high hydrophobic amino acid content: 36% for 23.5 kDa and 50% for 25 kDa. This hemolytic activity could be related, along with other amebic factors, to tissue damage.

Variation of proteins and proteinases in Entamoeba histolytica lysates containing a protease inhibitor.

Sodium dodecyl sulfate (SDS)-lysates of E. histolytica trophozoites were analyzed by electrophoresis in simple and gelatin-containing ("substrate") SDS-polyacrylamide gels. In simple gels, boiled lysates with para hydroxymercuribenzoate (pHMB) had a complex pattern of apparently undegraded proteins; boiled lysates without pHMB showed a major 30 kDa and four minor (43, 46, 63 and 117 kDa) proteins, whereas unheated lysates displayed only the 117 kDa protein. Using substrate gels no gelatinases were detected in heated lysates; unheated lysates without pHMB showed a major 30 kDa and three minor (33, 46 and 68 kDa) gelatinases, whereas those with pHMB presented a major 56 kDa and two minor (70 and 105 kDa) gelatinases. Three caseinase peaks were separated by Sephadex G-75 chromatography from unheated lysates: peak I contained 46, 56 and 117 kDa pHMB-sensitive gelatinases and peaks II and III contained smaller pHMB-resistant caseinases. We conclude that proteins remaining in lysates after SDS-induced proteolysis appear to be mainly proteases relatively resistant to self-digestion whose type and amount changes with the conditions of lysis and the presence of inhibitors; this is exemplified by the finding of the major gelatinase of lysates with pHMB being larger (56 kDa) than in lysates lacking the inhibitor (30 kDa).