Genes Nutr (2010) 5:331–341 Fig. 1 a A log plot of concentrations of the T. indica fruit pulp extracts (lg/ml) versus percentage (%) of HepG2 cells viability. From the log10 plot, the IC50 was found to be 4.57 mg/ml. b Examination under light microscope of morphological changes in HepG2 cells incubated for 48 h in the presence of 300 lg/ml and 4.57 mg/ml of the T. indica fruit pulp extracts In addition, a set of genes that are involved in the cell cycles CCNG1, CCNG2, CCNA2, CCNB1, CCNB2 and CDC2 were also signiﬁcantly down-regulated. Gene ontology: biological process Gene ontology analysis of the differentially expressed genes in HepG2 cells in response to treatment with the methanol extracts of the T. indica fruit pulp is shown in 335 Figs. 4 and 5. In Fig. 4, genes (and subsequently their gene products) were categorized into their participations in general metabolisms. It was found that 60 genes that are related to reactive oxygen species were up-regulated, whereas 13 were down-regulated. In addition, 95 genes that are involved in lipid metabolism were down-regulated, and another 62 were up-regulated. Figure 5 shows the number of signiﬁcantly regulated genes that are categorized according to their roles in major biological processes (a), molecular functions (b) and major cellular components (c). The green bars represent the up-regulated genes, and the purple bars depict the down-regulated genes. Validation of the microarray data Semi-quantitative RT–PCR analysis was performed on selected signiﬁcantly regulated genes as follows: PDE3A, GSTA1 and MTTP (Fig. 6). The sizes of the PCR products that corresponded to the selected genes were 180, 230 and 180 bps, respectively. The PCR product for the house- keeping gene, GADPH, which is 457 bp, is shown by the arrow. The relative expression of the PDE3A, GSTA and MTTP genes to that of the housekeeping gene were -0.69-, 1.401- and -0.439-fold, respectively. The expression of the selected genes PDE3A, GSTA1, CYP1A1 and MTTP as well as the interferon-induced protein (IFIT1) relative to that of GADPH was also quantitated using real-time RT–PCR. As shown in Fig. 7, the pattern of expression of the genes analysed using microarray technology was similar to that of the real-time RT–PCR. The MTTP gene was down-regulated by -1.25- fold in real-time RT–PCR compared with -1.96-fold in microarray. PDE3A gene was down-regulated by about -5.06-fold in RT–PCR when compared with -5.10 in microarray analysis. CYP1A1 gene was also shown to be down-regulated when analysed using both RT–PCR and microarray methods by -2.73- and -3.61-fold, respec- tively. GSTA1 was up-regulated by 1.54-fold in RT–PCR analysis compared with 2.8-fold in microarray. In addition, IFIT1 was up-regulated by fourfold in RT–PCR compared with 6.41-fold in microarray. All data were normalized to the reference gene, GAPDH. The expression patterns obtained with semi-quantitative RT–PCR and real-time RT–PCR were consistent with the microarray results. Discussion The MTT assay is a rapid colorimetric assay developed by Mosman [28] and had since been widely used to measure cell viability/proliferation in living cells [2, 5, 15]. In this study, the methanol extracts of the T. indica fruit pulp were shown to have an IC50 of 4.57 mg/ml. For the microarray 123