Background: Breast cancer is the major cause of death from cancer among women around the world. Given the drug resistance in the treatment of this disease, it is very important to identify new therapies and anticancer drugs. Many studies demonstrated that hypericin could induce apoptosis in different cancer cell lines; however, the underlying mechanism is not well understood yet. Therefore, this study aimed to evaluate the anticancer effect of hypericin in two breast cancer cell lines, one with wild type P53 and the other with mutant P53. Methods: In this study, the MDA-MB-231 and MDA-MB-175-VII cell lines were treated with different concentrations of hypericin for 24 and 48 hours. The measurement of cell death was performed by MTT assay. The cell apoptosis rate was measured using annexin V/propidium iodide assay through flow cytometry. The level of expression in P21 and P53 genes was evaluated by real time PCR. Immunocytochemistry (ICC) analysis was performed for P21 (direct target for P53 protein) to confirm the results. Results: The results showed that hypericin could have dose-dependent cytotoxic effects on the MDA-MB-231 and MDA-MB-175-VII cell lines, and its cytotoxicity is much higher in the latter cells. According to flow cytometry results, 86% of MDA-MB-175-VII cells underwent apoptosis with IC50 dose of hypericin for MDA-MB-231 cells after 24 hours. Moreover, after 24 hours of exposure to hypericin with MDA-MB-231 IC50 concentration, the expression of P53 and P21 genes upregulated in MDA-MB-175-VII much more than MDA-MB-231 when both cell lines were treated with 24 hours IC50 dose of MDA-MB-231. The ICC analysis on P21 confirmed that by treating both cell lines with MDA-MB-231 IC50 dose of hypericin for 24 hours, this protein is overexpressed much more in MDA-MB-175-VII cells. Conclusion: The results of this study demonstrated that hypericin’ s apoptotic and cytotoxic effects on cancer cells may be mediated via P53 overexpression, cell cycle arrest and the subsequent apoptosis. Therefore, it is of great importance to consider that hypericin would have better impact on cells or tumors with wild type P53.

Background: Although several roles of 27-hydroxycholesterol (27-HC), the most abundant oxysterol in blood circulation, in cancers have been elucidated, its impact on breast cancer proliferation and its pathway remain unknown. Materials and Methods: The effect of 27-HC on breast cancer cell proliferation and its pathway was evaluated using Michigan Cancer Foundation-7 (MCF-7) and M. D. Anderson-Metastatic Breast 231 (MDA-MB-231) cell lines. The MTT assay was applied after 24-and 48-hour incubation to distinguish cell proliferation. To determine the cause of different viability results from the MTT assay, the Annexin-FITC/PI test was used at concentrations of 0. 1, 1, and 10 μM after 24-and 48-hour incubation. Results: 27-HC in concentrations of 5, 10, and 20 μM induced cell cytotoxicity compared with control. Also, the annexin V conjugated with fluorescein isothiocyanate/propidium iodide (Annexin-FITC/PI) test revealed an increase in total apoptotic cells treated with 0. 1, 1, and 10 μM of 27-HC after 48 hours (P value < 0. 05). Besides, the cytotoxic effect of 27-HC was observed at 10 μM concentration in both cell lines, MCF-7 and MDA-MB-231 (P value < 0. 05). Conclusion: The identification of 27-HC’s cytotoxic effects on both estrogen receptor (ER)-negative and ER-positive breast cancer cell lines is a novel discovery that may be linked to LXRβ.

Background: Exosomes are membrane nanovesicles, 30 to 100 nm in diameter, secreted by most cell types. Besides playing many biological roles, especially in cell-cell communication, scientific proof indicated that the pathology of so many human cancers is closely related tomanybiologic elements in exosomes. Theymayserve as useful biomarkers for treatment, prognosis, anddetection. Cancer cells produce more exosomes, inducing changes in target cells (near or distant from the tumor), such as metastasis and chemotherapy resistance. Therefore, isolation, identification, and analysis of these microvesicles seem essential. Objectives: The current study aimed at collecting and purifying microvesicles secreted from breast cancer cells and confirming the identity of the obtained exosomes using methods assessing size and morphology. Methods: In recent research, the MDA-MB-231 cell line was grown under standard conditions. Released exosomes were collected and ultra-centrifuged. Scanning (SEM) and transmission (TEM) electron microscopes, atomic force microscopy (AFM), and dynamic light scattering (DLS) were used to assess exosome size. Results: The obtained data revealed that MDA-MB-231 cells produced exosomes. The nanovesicles were isolated from the culture medium of MDA-MB-231 cells by applying different strategies, including differential centrifugation, filtration, and ultracentrifugation. The exosomes were characterized; they had a size of 30-100 nm and spherical shape. Conclusions: Intercellular communication can be mediated through direct cell-cell contact or transfer of secreted molecules. In the last two decades, a third mechanism for intercellular communication has emerged that involves intercellular transfer of extracellular vesicles (exosomes). Due to their many functions in the body, it is of great importance to purely isolate and recognize exosomes to understand their modes of action as the first step in the advancement of researches. However, more research is required to obtain cost-effective and efficient methods. It was found that MDA-MB-231 cells release exosomes. They are spherical and 30-100nmin diameter. The use of a combination strategy for the first time was useful in isolating exosomes derived from MDA-MB-231 cells without disturbing their structure. Further studies are required to compile a uniform protocol for exosome isolation in medical research.