Hexane fraction of Globimetula braunii induces mitochondria-mediated apoptosis in Plasmodium berghei-infected mice

Abstract

BackgroundApoptosis is a common pathology in malaria and most antimalarial drugs induce apoptosis during chemotherapy. Globimetula braunii is an African mistletoe used for the treatment of malaria but its effect on mitochondria-mediated apoptosis is not known. MethodsMalarial infection was induced by the intraperitoneal injection of NK 65 strain Plasmodium berghei-infected erythrocytes into mice which were treated with graded doses (100–400 mg/kg) of methanol extract (ME), and fractions of n-hexane, dichloromethane, ethylacetate and methanol (HF, DF, EF and MF) for 9 days after the confirmation of parasitemia. Artequine (10 mg/kg) was used as control drug. The fraction with the highest antiplasmodial activity was used (same dose) to treat mice infected with chloroquine-resistant (ANKA) strain for 5 consecutive days after the confirmation of parasitemia. P-alaxin (10 mg/kg) was used as control drug. On the last day of the treatment, liver mitochondria were isolated and mitochondrial Permeability Transition (mPT) pore opening, mitochondrial F0F1 ATPase (mATPase) activity, lipid peroxidation (mLPO) and liver deoxyribonucleic acid (DNA) fragmentation were assessed spectrophotometrically. Caspases 3 and 9 were determined by Enzyme-Linked Immunosorbent Assay (ELISA) technique. Cytochrome c, P53, Bcl-2-associated X protein (Bax), and B-cell lymphoma-2 (Bcl2) were determined via immunohistochemistry. Phytochemical constituents of the crude methanol extract of Globimetula braunii were determined via the Gas Chromatography-Mass Spectrometry (GC-MS) analysis ResultsThere was large amplitude mPT induction by malaria parasites, extract and fractions of Globimetula braunii. At 400 mg/kg, HF significantly (p < 0.01) downregulated mATPase activity, and mLPO in both (susceptible and resistant) models, caused DNA fragmentation (P < 0.0001), induced caspases activation, P53, bax and cytochrome c release but downregulated Bcl2 in both models. The GC-MS analysis of methanol extract of Globimetula braunii showed that α-amyrin is the most abundant phytochemical. ConclusionThe n-hexane fraction of Globimetula braunii induced mitochondrial-mediated apoptosis through the opening of the mitochondrial pore, fragmentation of genomic DNA, increase in the levels of P53, bax, caspase 3 and 9 activation and cytochrome c release with concomitant decrease in the level of Bcl2. α-Amyrin is a triterpene with apoptotic effects.