The toxicological properties of Scilla nervosa (Burch.) Jessop (Hyacinthaceae) in cultured HepG2 liver cells.

Abstract

Background and Aims of Study. Scilla nervosa is a member of the Hyacinthaceae plant family that has been naturalised in the grasslands of Southern Africa. The bulbs are traditionally used to treat a variety of ailments. For example, the Zulu people use aqueous decoctions of the bulbs as analgesics in the treatment of rheumatic fever, crushed bulbs are used by the Sotho people as laxatives and the Tswana people use cooked bulbs to treat infertility in women as well as cold aqueous extracts to treat infections. It was recently demonstrated in our laboratory that extracts prepared from the bulbs possess potent anti-inflammatory properties, and this may therefore provide a rationale for the traditional use of the plant as an analgesic. Several studies have demonstrated that the bulbs contain homoisoflavanones and stilbenoids that could be responsible for their therapeutic effects. Although the plant has diverse medicinal applications, and despite it being recognised as a poisonous species particularly in livestock, little is known about its toxicity in human liver cells. The objectives of this study were therefore to investigate the potential toxicity of the bulbs on the liver, a major detoxifying organ. A human liver cell line was treated with an aqueous extract of the bulbs to investigate (1) cell viability, (2) potential mechanisms of cytotoxicity, (3) DNA integrity, and (4) changes in the cytochrome P450 enzyme activity. Materials and Methods. This study was conducted on the cultured HepG2 human hepatocellular carcinoma cell line, a model system to investigate the cytotoxicity of xenobiotics. The viability of cultured HepG2 liver cells in the presence of varying concentrations of an aqueous extract of the bulbs was determined after 24 hours treatment, and the concentration that reduced viability to 50% (IC50) was derived. Potential mechanisms of cytotoxicity at the IC50 were investigated. These included changes in metabolic activity (intracellular adenosine triphosphate (ATP) quantification), apoptosis induction (phosphatidylserine (PS) externalisation, caspase-8 and -9 induction and changes in mitochondrial membrane potential), and oxidative damage via free radical formation (lipid peroxidation). Genotoxicity was investigated by determining changes in DNA integrity (DNA fragmentation). The ability of the extract to stimulate or {¹ Du Toit, K., Kweyama, A., Bodenstein, J. 2011. Anti-inflammatory and antimicrobial profiles of Scilla nervosa (Burch.) Jessop (Hyacinthaceae). South African Journal of Science, 107:96-100.} inhibit enzymes commonly involved with drug metabolism was investigated by determining cytochrome P450 3A4 (CYP3A4) activity. Results Cell-viability decreased in a concentration-dependent manner and the IC50 was determined as 0.03 mg/ml. Treating the cells at the IC50 resulted in (1) a 1.2-fold increase in intracellular ATP levels, (2) no significant change in PS externalisation, (3) a 1.3-fold increase in caspase-8 activity, (4) a 1.1-fold decrease in caspase-9 activity, (5) no significant change in mitochondrial membrane potential, (6) a 1.9-fold increase in lipid peroxidation, (7) evidence for genotoxicity as demonstrated by DNA fragmentation, and (8) no evidence of changes in CYP3A4 activity. Conclusion. Results suggest that HepG2 liver cells are sensitive to an aqueous extract of the bulbs of S. nervosa. The extract has the potential to (1) induce apoptosis, (2) increase oxidative stress and (3) cause genotoxicity in vitro. peroxidation, genotoxicity