The effect of selected medicinal plants on rotenone-induced toxicity in SH-SY5Y neuroblastoma cells

Abstract

Parkinson's disease (PD) is the second most common chronic neurodegenerative disease characterized by dopamine decrease in the substantia nigra. Currently, there is no promising cure for PD and this has resulted in extensive research into alternative medicines. The aim of this study was to investigate the effect of methanol and ethyl acetate extracts of Lannea schweinfurthii (Engl. Engl) (Anacardiaceae), Zanthoxylum capense (Thunb. Harv) (Rutaceae), Scadoxus puniceus ((L.) Friis & Nordal) (Amaryllidaceae) and Crinum bulbispermum (Burm. f.) Milne-Redh. & Schweick) (Amaryllidaceae) on rotenone-induced toxicity in SH-SY5Y neuroblastoma cells. The latter which mimics PD symptoms in vitro. Cytotoxicity of the plant extracts was assessed using sulforhodamine B (SRB) assay. Intracellular reactive oxygen species (ROS) were measured fluorometrically with the use of the fluorescent dye 2‟,7‟-dichlorodihydrofluorescein diacetate (H2DCF-DA). Intracellular glutathione content was measured fluorometrically after staining with monochlorobimane (MCB). Fluorescent dye 5,5‟ ,6,6‟ -tetrachloro-1,1‟ ,3,3‟ -tetraethylbenzimidazolcarbocyanine iodide (JC-1) was used to assess the mitochondrial membrane potential (MMP) status of cells. Apoptosis was assessed by determining caspase-3 activity through detection of 7-amino-4-methylcoumarin (AMC) which is a product of caspace-3 substrate, acetyl-Asp-Glu-Val-Asp 7-amino-4-methylcoumarin (Ac-DEVD-AMC), cleaved by the caspase-3 enzyme. Rotenone was used as an in vitro model to induce PD-like symptoms. Cytotoxicity studies for methanol extract of Zanthoxylum capense revealed the highest IC50 value of 121.3 μg/mL, indicating low toxicity. The ethyl acetate extract of Crinum bulbispermum was observed to have no effect on the normal proliferation of the SH-SY5Y cells and produced an IC50 value >100 μg/mL. The calculated IC50 value obtained from rotenone cytotoxicity studies was 112 iv nM. Zanthoxylum capense and Scadoxus puniceus plant extracts were observed to be neuroprotective against rotenone-induced toxicity. A decrease in intracellular glutathione content as well as MMP was also observed in cells exposed to rotenone alone (50 nM). There was no intracellular ROS generation observed in cells exposed to rotenone alone (50 nM) after 24 h and 72 h. However, apoptotic cell death was observed in cells treated with rotenone (50 nM). Intracellular ROS production was observed to be elevated by methanol and ethyl acetate extracts of C. bulbispermum. Methanol extracts of Z. capense was observed to increase intracellular glutathione content. MMP was increased effectively following treatment with ethyl acetate extract of C. bulbispermum. Moreover, both methanol and ethyl acetate plant extracts were found to decrease caspase-3 activity significantly (p<0.05), in cells exposed to 50 nM rotenone. Z. capense methanol extract reduced caspase-3 activity the most effectively. Treatment with plant extracts was protective and decreased cell death. Furthermore, L. schweinfurthii, Z. capense, S. puniceus and C. bulbispermum, demonstrated strong antioxidant and anti-apoptotic effects against rotenone-toxicity, making them potential agents in developing therapies for treating PD.