The cytotoxic effects of Scilla nervosa (Burch.) Jessop (Hyacinthaceae) aqueous extract on cultured HepG2 cells Prishania Pillay a , Alisa Phulukdaree b , Anil A. Chuturgoon b , Karen Du Toit a , Johannes Bodenstein a,n a Discipline of Pharmaceutical Sciences, University of KwaZulu-Natal, Private Bag X54001, Durban 4000, South Africa b Discipline of Medical Biochemistry, University of KwaZulu-Natal, Private Bag X54001, Durban 4000, South Africa article info Article history: Received 22 May 2012 Received in revised form 25 October 2012 Accepted 27 October 2012 Available online 2 November 2012 Keywords: Scilla nervosa (Hyacinthaceae) HepG2 liver cells Caspase Apoptosis Genotoxicity CYP3A4 abstract Ethnopharmacological relevance: Bulbs of Scilla nervosa, a medicinal plant indigenous to Southern Africa, are traditionally used in aqueous decoctions to treat a diverse range of illnesses. The bulbs contain homoisoflavanones and stilbenoids. Little information is known about the plant’s toxicity on the liver, a major detoxifying organ. This study investigated the effects of an aqueous extract of the bulbs in cultured HepG2 liver cells, a model system for investigating the toxicity of xenobiotics. Materials and methods: The concentration that reduced cell viability to 50% (IC 50 ) after 24 h treatment was derived. Potential mechanisms of toxicity using the IC 50 were investigated as changes in metabolic activity, apoptosis, oxidative damage and DNA fragmentation. In addition, cytochrome P450 3A4 (CYP3A4) activity, which is implicated in drug metabolism and interactions, was also assayed. Results: Cell viability decreased in a concentration-dependent manner and the IC 50 was determined as 0.03 mg/mL. Treating the cells at the IC 50 for 24 h resulted in increased intracellular ATP levels, no significant change in phosphatidylserine externalisation, increased caspase-8 activity, decreased caspase-9 activity, no significant change in mitochondrial membrane potential, increased lipid peroxidation, evidence for genotoxicity as demonstrated by DNA fragmentation, and slightly induced CYP3A4 activity. Conclusion: Results suggest that liver cells are sensitive to an aqueous extract of the bulbs and there is an increased potential to induce apoptosis, oxidative stress and genotoxicity in vitro. & 2012 Elsevier Ireland Ltd. All rights reserved. 1. Introduction Scilla nervosa (Burch.) Jessop (Hyacinthaceae)[ ¼ Schizocarphus nervosus (Burch.) Van der Merwe] is a monocotyledonous per- ennial plant originally endemic to Botswana and has naturalised in the grasslands of the eastern parts of Southern Africa (Hutchings et al., 1996). The bulbs of the plant are considered to be a valuable medicinal species and have been used by traditional healers of different cultures to treat infertility in women, constipation, dysentery, nervous conditions, rheumatic fever and pain (Watt and Breyer-Brandwijk, 1962). In livestock it is used to treat gall sickness (Watt and Breyer-Brandwijk, 1962). It has been recently demonstrated that extracts prepared from the bulbs possess potent anti-inflammatory properties and these findings may therefore rationalise the traditional use of the plant as an analgesic for rheumatic fever (Du Toit et al., 2011). Previous studies have revealed that the bulbs contain homoisoflavanones and stilbenoids (Bangani et al., 1999; Silayo et al., 1999). The chemical structures, plant origins, ethnobotany and biological activities of homoisoflavanones have been reviewed by Du Toit et al. (2010). Studies by Bangani et al. (1999) demonstrated 5 homoisoflavanones and 2 stilbenoids, while Silayo et al. (1999) demonstrated 13 homoisoflavanones and 3 stilbenoids. These could individually or in combination be responsible for the observed therapeutic effects. However, little information is known about the plant’s toxicity. Only one previous report suggested that 0.5–1 kg of the fresh plant in the flowering stages was toxic to sheep (Van der Walt and Steyn, 1946). Therefore, a current investigation of the potential toxicity of the bulbs on the liver, a major detoxifying organ, is required. The aim of this study was to investigate the toxic effects of an aqueous extract prepared from the bulbs in HepG2 liver cells, a model system to investigate the toxicity of xenobiotics in vitro. Specifically, the viability of HepG2 cells in the presence of varying concentrations of the extract for 24 h was investigated to determine the IC 50 -value. Contents lists available at SciVerse ScienceDirect journal homepage: www.elsevier.com/locate/jep Journal of Ethnopharmacology 0378-8741/$ - see front matter & 2012 Elsevier Ireland Ltd. All rights reserved. http://dx.doi.org/10.1016/j.jep.2012.10.053 Abbreviations: DCm, mitochondrial membrane potential; CCM, complete culture medium; CYP3A4, cytochrome P450 3A4; IC 50 , concentration of extract that reduced cell viability to 50%; JC-1, 5,5 0 ,6,6 0 -tetrachloro-1,1 0 ,3,3 0 -tetraethylbenzimi- dazolcarbocyanine; MDA, malondialdehyde; MTT, 3-(4,5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2H-tetrazolium bromide; PBS, phosphate buffered saline; RLU, relative light units n Corresponding author. Tel.: þ27 31 260 7500; fax: þ27 31 260 7907. E-mail address: bodensteinj@ukzn.ac.za (J. Bodenstein). Journal of Ethnopharmacology 145 (2013) 200–204