Encystation of Entamoeba invadens IP‐1 Is Induced by Lowering the Osmotic Pressure and Depletion of Nutrients from the Medium¹

Abstract

Trophozoites of Entamoeba invadens IP-1 can be induced to encyst in simple solutions composed of semipermeable constituents (buffer, salts, or sugars) provided that their osmotic pressure is in the range of 60-160 mosmol/kg. Optimal yield of mature cysts was obtained when the osmotic pressure of the medium was 110 mosmol/kg. Encystation could be obtained in the absence of serum although higher yields were obtained in its presence. No difference in the yield of mature cysts was found when either dialyzed or full serum was used. High yields of encystation were obtained (greater than 70%) in the presence of 5% serum in solutions of NaCl, KCl, or MgSO4, suggesting that the mechanism of encystation is not induced via sodium or potassium channels. Cysts were obtained in the presence of 72 mM glucose, indicating that depletion of a carbon source is not the only requirement for encystation. A rapid change in the density of the Entamoeba cells was observed upon transfer of trophozoites (density 1.061-1.073 g/ml) from growth medium to the low osmotic pressure encystation solutions. Within the first 2 min their density decreased (to 1.050 g/ml), but it soon increased, reaching within 30 min a density higher than 1.120 g/ml. As the encystation process continued to completion, the density of the cells gradually decreased, the mature cysts reaching a density of 1.049-1.061 g/ml.

Current Therapeutics, Their Problems, and Sulfur-Containing-Amino-Acid Metabolism as a Novel Target against Infections by “Amitochondriate” Protozoan Parasites

“…Although signals leading to encystation and excystation of E. histolytica are still poorly understood, osmolality changes, nutrient depletion (278), and adherence via galactose-binding lectin (79) are likely involved in encystation. In vitro encystation of the related reptilian species Entamoeba invadens was established by manipulating osmotic and nutrient conditions of axenic cultures (20,278,317). However, encystation has not been achieved with an axenic E. histolytica line.…”

Proteasome‐dependent cyst formation and stage‐specific ubiquitin mRNA accumulation in Entamoeba invadens

“…This second differentiation process involves organelle rearrangement [1], novel gene expression and protein synthesis [2,3], the formation of an external chitinaceous cell wall [4], and ultimately results in the creation of the infectious form of the parasite [5]. Entamoeba invadens, a parasite of reptiles, will undergo this process in vitro in response to carbon source deprivation [6] or osmotic shock [7], and is used as a model of the human parasite, E. histolytica, which does not efficiently encyst under similar axenic culture conditions.The bulk of the turnover of intracellular proteins in eukaryotic cells is carried out by two self-contained proteolytic systems, the lysosomes and proteasomes. Most proteins to be degraded by the proteasome/ubiquitin system are covalently conjugated to ubiquitin and then unfolded and threaded into the proteasomes where they are broken down by the three types of proteolytic activities expressed by the b-subunits of the multisubunit 20S proteasome complex [8,9].…”

“…This second differentiation process involves organelle rearrangement [1], novel gene expression and protein synthesis [2,3], the formation of an external chitinaceous cell wall [4], and ultimately results in the creation of the infectious form of the parasite [5]. Entamoeba invadens, a parasite of reptiles, will undergo this process in vitro in response to carbon source deprivation [6] or osmotic shock [7], and is used as a model of the human parasite, E. histolytica, which does not efficiently encyst under similar axenic culture conditions.…”

Untitled

“…By applying the same methodology as well as other factors, such as the presence of metal ions in the serum 20 , osmotic shock 21,22 and fasting glucose 23 , the presence of mucin glycoprotein acts as a ligand for the lectin on the membrane and results in the aggregation of Entamoeba trophozoites and short-chain fatty acids 24 .…”