Evaluation of the antioxidant activity and antiproliferative effect of the jaboticaba (Myrciaria cauliflora) seed extracts in oral carcinoma cells

Abstract

It is becoming increasingly evident that certain phytochemicals possess cancer chemopreventive properties. In this study, the antiproliferative activity of extracts from different parts of the jaboticaba (Myrciaria cauliflora) plant was evaluated for its effect on human oral carcinoma cell lines. The cytotoxicities of various plant extract concentrations were examined and the 50% maximal inhibitory concentration (IC50) was determined. Water extracts of jaboticaba seeds showed concentration-dependent antiproliferative effects. Annexin V/propidium iodide positivity with active caspase-3 induction indicated that the treated cells underwent apoptosis. Several important regulatory proteins (Bcl-2, Bcl-xL, Bid, and survivin) involved in apoptosis were also evaluated. The antioxidant activity of jaboticaba was investigated using 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2'-azinobis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) assays, and the drug concentration eliciting 50% maximum stimulation (SC50) was determined. The present findings suggest that water extracts of jaboticaba seeds exhibit an antiproliferative effect against oral cancer cells by inducing apoptosis through downregulating survivin expression and thereby activating caspase-mediated Bid cleavage.

Figure 1

Antioxidative activity of different parts of jaboticaba.Using DPPH and ABTS assays, the water extract of jaboticaba seeds was found to have the best scavenging activity compared to other portions of the plant. Trolox served as a positive control.

Figure 2

Water extract of jaboticaba seeds possessed the most potent cytotoxic activity. Cell survival rate of HSC-3 cells after 24 h treatment with various extracts showed that the water extract of jaboticaba seeds had the most significant cytotoxic activity compared to extracts from other portions of the plant. The IC₅₀ of water extract of jaboticaba seeds in HSC-3 cells was approximately 15 μg/mL. Each point represents the mean and SD of six determinations.

Figure 3

Apoptosis induction by jaboticaba seed water extract treatment of HSC-3 cells. (a) HSC-3 cells treated with jaboticaba seed water extract for 24 h at desired concentrations were stained with Annexin V-FITC and propidium iodide (PI). The Annexin V-FITC signal is shown on the x-axis and PI signal is shown on the y-axis. The apoptotic populations of cells were significantly increased in treatment groups in a dose-dependent manner. (b) Flow cytometric analysis was performed after a 48 h treatment to determine caspase-3 activity, which was found to be activated in a dose-dependent manner.

Figure 4

Effects of jaboticaba seed water extract on apoptosis regulatory proteins in HSC-3 cells. HSC-3 cells were treated with 5, 10, 25, 50, and 100 μg/ml of jaboticaba seed extract or ddH₂O as a control for 48 h, and then total proteins were isolated. Equal amounts of cell lysates were analyzed for PARP, survivin, Bid, Bcl-xL, and Bcl-2 expression by Western blotting with corresponding antibodies. β-actin served as the loading control. The fold change was calculated as the ratio of the target proteins in the presence of indicated concentration of Jaboticaba seed extract after normalization of the target proteins to β-actin in each lane.