Fig. 4

a, Apoptotic potential of SNME on NCI/ADR-RES cells. Cells were treated with 7.8125, 15.625, 31.25, 62.5, 125 and 250 μg/ml of methanolic extract for 6, 12 and 24 h, stained with ethidium bromide-acridine orange. Control cells were maintained in the vehicle for the indicated time periods. Results are represented as mean ± SEM of three assays. Representative microscopic image showing ethidium bromide – acridine orange stained apoptotic cells after treatment with 125 μg/ml SNME on NCI/ADR-RES cells. b, Nuclear condensation studies of SNME on NCI/ADR-RES cells. Cells were treated with 7.8125, 15.625, 31.25, 62.5, 125 and 250 μg/ml of methanolic extract for 6, 12 and 24 h, stained with Hoescht. The number of bright blue condensed spots indicating apoptotic cells with condensed chromatin increases with Solanum nigrum treatment in a time and concentration dependent manner. Control cells were maintained in the vehicle for the indicated time periods. Results are represented as mean ± SEM of three assays. Representative microscopic image showing nuclear condensation (→) in Hoescht stained apoptotic cells after treatment with 125 μg/ml SNME on NCI/ADR-RES cells. c, SNME induces apoptosis in NCI/ADR-RES cell. FITC-conjugated annexin binding to phosphatidyl serine, exposed to the outer leaflet, is measured by FACS analysis in NCI/ADR-RES cells after treatment with methanolic extract of S. nigrum 125 μg/ml for 12 h and 24 h. Apoptotic population (Q4) FACS histogram plotted against PI vs. FITC staining with a notable increase in the number of apoptotic cells after treatment for 24 h. Staurosporine (1 μg/ml) treated cells were used as positive control. d, DNA fragmentation of NCI/ADR-RES cells exposed to methanolic extract of unripe Solanum nigrum. Cells were incubated with 125 μg/ml at 0, 6, 12, 24 and 48 h. DNA ladders reflecting the presence of DNA fragments were viewed on ethidium –bromide stained 2% agarose gel