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Enhancement of hemin-induced membrane damage by artemisinin
1Department of Pathology, New England Deaconess Hospital, Boston, MA 02215.
Abstract:
Artemisinin is an effective antimalarial agent, and its action on the malarial parasite is suggested to be mediated by oxidative processes. Since malarial parasites contain a high concentration of hemin, and hemin may induce the formation of reactive oxygen species, we investigated the interaction of artemisinin, iron and hemin. We used erythrocyte membrane-bound Ca2+ pump ATPase (basal) and calmodulin (CaM)-activated Ca2+ pump ATPase as our model. Membranes were incubated with artemisinin in the presence or absence of iron-ascorbate or hemin at 37 degrees for 1 hr. Following incubation, ATPase activity was measured. Our results showed that artemisinin (500 microM) had no effect on ATPase activities. However, artemisinin enhanced the inhibitory effect of iron (50 microM)-ascorbate (500 microM) on ATPase activity (46.3 +/- 3.9 vs 63 +/- 2.1% for basal; 57.2 +/- 2.5 vs 74.8 +/- 2.1% for CaM-activated). Desferrioxamine (DFO, 200 microM) blocked significantly the effect of iron-ascorbate-artemisinin on ATPases (P < 0.01). Hemin inhibited ATPase activity in a concentration-dependent fashion. Artemisinin enhanced hemin (10 microM)-induced inhibition of basal (36.0 +/- 6.0 vs 73.7 +/- 3.0%) and CaM-activated Ca2+ pump ATPase (31.6 +/- 2.8 vs 70.0 +/- 1.5%). Iron chelators (DFO, ferene, 8-hydroxyquinoline, 1,10-phenanthroline, and 1,2-dimethyl-3-hydroxypyrid-4-one) had no effect on artemisinin plus hemin-induced enzyme inhibition. Catalase (2000 U/mL) had a minor effect on the artemisinin-hemin or hemin-mediated effect. Thiourea (1 mM) had no effect. However, superoxide dismutase (500 U/mL) and dithiothreitol blocked artemisinin-hemin or hemin-mediated ATPase inhibition significantly (P < 0.001). In conclusion, these results suggest that, in our model, artemisinin enhances the damage of hemin-induced ATPases via oxidation of thiol groups on the enzymes. Free iron or hydroxyl radical does not seem to be involved. This interaction between artemisinin and hemin may contribute to the antimalarial action of artemisinin against malarial parasites.
Insights
Artemisinin enhances hemin-induced damage to erythrocyte ATPases, suggesting a mechanism for its antimalarial action. This interaction involves oxidation of enzyme thiol groups, not free iron or hydroxyl radicals.
Area of Science:
- Biochemistry
- Pharmacology
- Malariology
Background:
- Artemisinin is a key antimalarial drug.
- Its mechanism involves oxidative processes.
- Malarial parasites have high hemin concentrations, which can generate reactive oxygen species.
Purpose of the Study:
- Investigate the interaction between artemisinin, iron, and hemin.
- Determine the effect on erythrocyte membrane-bound Ca2+ pump ATPase (basal and calmodulin-activated).
Main Methods:
- Incubation of erythrocyte membranes with artemisinin, iron-ascorbate, or hemin.
- Measurement of ATPase activity.
- Use of iron chelators, catalase, superoxide dismutase, and dithiothreitol to probe mechanisms.
Main Results:
- Artemisinin alone did not affect ATPase activity.
- Artemisinin enhanced the inhibitory effects of iron-ascorbate and hemin on ATPase activity.
- Superoxide dismutase and dithiothreitol blocked the artemisinin-hemin-mediated inhibition.
- Iron chelators and catalase had no significant effect.
Conclusions:
- Artemisinin enhances hemin-induced damage to ATPases via oxidation of enzyme thiol groups.
- Free iron or hydroxyl radicals are unlikely to be involved.
- This artemisinin-hemin interaction may contribute to artemisinin's antimalarial efficacy.