Objective: In this study, bystander effects in MCF-7 breast cancer cells and hFOB 1.19 normal osteoblast cells irradiated with gamma emitting HDR Brachytherapy Ir-192 source were investigated.
Material and Methods: In this in-vitro study, bystander effect stimulation was conducted using medium transfer technique of irradiated cells to the non-irradiated bystander cells. Cell viability, reactive oxygen species (ROS) generation and colony forming assay was employed to evaluate the effect.
Results: Results indicate that the exposure to the medium irradiated MCF-7 induced significant bystander killing and decreased the survival fraction of bystander MCF-7 and hFOB from 1.19 to 81.70 % and 65.44 %, respectively. A significant decrease in survival fraction was observed for hFOB 1.19 bystander cells (p < 0.05). We found that the rate of hFOB 1.19 cell growth significantly decreases to 85.5% when added with media from irradiated cells. The ROS levels of bystander cells for both cell lines were observed to have an increase even after 4 h of treatment. Our results suggest the presence of bystander effects in unirradiated cells exposed to the irradiated medium.
Conclusion: These data provide evidence that irradiated MCF-7 breast cancer cells can induce bystander death in unirradiated MCF-7 and hFOB 1.19 bystander cells. Increase in cell death could also be mediated by the ROS generation during the irradiation with HDR brachytherapy.
MATERIALS AND METHODS: Cytotoxicity was measured by the MTT assay and further confirmed via apoptosis, ROS, cell cycle, DNA fragmentation and cytokine assays.
RESULTS: ITHB4 demonstrated a lower IC50 compared to zerumbone in inhibiting the proliferation of MCF-7 cells. ITHB4 showed no toxicity against normal breast and human immune cells. Apoptosis assay revealed that ITHB4, at a concentration equal to the IC50, induces apoptosis of MCF-7 cells and cell cycle arrest at the sub-G1 and G2/M phases. ITHB4 triggered accumulation of intracellular ROS and nuclear DNA fragmentation. Secretion of pro-inflammatory cytokines induced inflammation and potentially immunogenic cell death.
CONCLUSION: ITHB4 has almost similar chemotherapeutic properties as zerumbone in inhibiting MCF-7 growth, and hence provide the basis for further experiments in animal models.
Methods: Chemical profiling of P. blanda was carried out using gas chromatography mass spectrometry (GCMS) followed by isolation of bioactive compounds by column chromatography. DPPH• and FRAP assays were used to evaluate antioxidant activity and the MTT assay was performed to estimate the cytotoxicity activity against three cancer cell lines, namely MCF-7, HL-60 and WEHI-3, and three normal cell lines, MCF10A, WRL-68 and HDFa.
Results: X-ray crystallographic data for peperomin A is reported for the first time here and N,N'-diphenethyloxamide was isolated for the first time from Peperomia blanda. Methanol and dichloromethane extracts showed high radical scavenging activity with an IC50 of 36.81 ± 0.09 µg/mL, followed by the dichloromethane extract at 61.78 ± 0.02 µg/mL, whereas the weak ferric reducing activity of P. blanda extracts ranging from 162.2 ± 0.80 to 381.5 ± 1.31 µg/mL were recorded. In addition, petroleum ether crude extract exhibited the highest cytotoxic activity against all the tested cancer cell lines with IC50 values of 9.54 ± 0.30, 4.30 ± 0.90 and 5.39 ± 0.34 µg/mL, respectively. Peperomin A and the isolated mixture of phytosterol (stigmasterol and β-sitosterol) exhibited cytotoxic activity against MCF-7 and WE-HI cell lines with an IC50 of (5.58 ± 0.47, 4.62 ± 0.03 µg/mL) and (8.94 ± 0.05, 9.84 ± 0.61 µg/mL), respectively, compared to a standard drug, taxol, that has IC50 values of 3.56 ± 0.34 and 1.90 ± 0.9 µg/mL, respectively.
Conclusion: The activities of P. blanda extracts and isolated compounds recorded in this study underlines the potential that makes this plant a valuable source for further study on anticancer and antioxidant activities.