The microaerophilic flagellate Giardia intestinalis: oxygen and its reaction products collapse membrane potential and cause cytotoxicity

摘要

Trophozoites of the microaerophilic flagellate parasitic protozoon Giardia intestinalis have only a limited capacity to detoxify O2. Thus, when exposed to controlled concentrations of dissolved O2 8?μM, they gradually lose their ability to scavenge O2. In a washed cell suspension stirred under 10% air in N2 (equivalent to 25?μM O2), inactivation of the O2-consuming system was complete after 3·5 h; during this period accumulation of H2O2 (3?μmol per 106 organisms) and oxidation of cellular thiols to 16% of their initial level occurred. Under 20% air (50?μM O2), respiratory inactivation was complete after 1·5?h, and under air (258?μM O2), after 50?min. Loss of O2-consuming capacity was accompanied by loss of motility. Use of the fluorogen 2,7-dichlorodihydrofluorescein acetate indicated that intracellular H2O2 is produced at extranuclear sites. Flow cytometric estimation of the plasma membrane electrochemical potentials using bis(1,3-dibutylbarbituric acid) trimethine oxonol, DiBAC4(3), showed that values declined from ?134?mV to ?20?mV after 4·5?h aeration. Incubation of organisms with 60?μM H2O2 for 10?min gave partial collapse of plasma membrane potential and complete loss of O2 uptake capacity; motility and viability as assessed by DiBAC4(3) exclusion were completely lost after 1?h. Inactivation of the O2-consuming system and loss of viability were also observed on exposure to singlet oxygen photochemically generated from rose bengal or toluidine blue.