Investigation of the effects of B16F10 derived exosomes in induction of immunosuppressive phenotype in the hematopoietic stem cells

Objective: This study aimed to elucidate the effects of melanoma-derived exosomes on modulating the differentiation of hematopoietic stem cells (HSCs) towards immunosuppressive myeloid-derived suppressor cells (MDSCs). Materials and Methods: Exosomes were isolated via ultracentrifugation from conditioned media of the B16F10 murine melanoma cell line after adaptation to exosome-free culture conditions. HSCs were extracted from the bone marrow of adult C57BL/6 mice through density gradient separation and MACS column isolation of CD133+ and CD34+ populations. HSCs were cultured with or without B16F10 exosomes for 24 hours. Flow cytometry analyzed the expression of canonical MDSC surface markers CD11b, Ly6G, and Ly6C. Levels of the immunosuppressive cytokines interleukin-10 (IL-10) and tumor necrosis factor beta (TGF-β) in HSC culture supernatants were quantified by ELISA. Results: Compared to untreated controls, HSCs treated with B16F10 exosomes displayed significantly increased percentages of CD11b+Ly6G+ granulocytic MDSCs and CD11b+Ly6C+ monocytic MDSCs, with a notable predominance of the Ly6G+ granulocytic subtype. Additionally, exosome-treated HSCs secreted markedly higher levels of the cytokines IL-10 and TGF-β, which are involved in MDSC-mediated immunosuppression. Conclusions: Our findings demonstrate that melanoma-derived exosomes can orchestrate the differentiation of HSCs into MDSCs with an immunosuppressive phenotype, as evidenced by the upregulation of MDSC surface markers and secreted cytokines. This supports a role for tumor-derived exosomes in driving the systemic expansion and accumulation of immunosuppressive MDSCs through the reprogramming of HSC fate. Elucidating the exosome contents and HSC signaling pathways involved could reveal therapeutic strategies to block this pathway and enhance anti-tumor immunity

Direct and indirect effects of exposure to 900 MHz GSM radiofrequency electromagnetic fields on CHO cell line: Evidence of bystander effect by non-ionizing radiation

© 2019 Introduction: The rapid rise in global concerns about the adverse health effects of exposure to radiofrequency radiation (RFR)generated by common devices such as mobile phones has prompted scientists to further investigate the biological effects of these environmental exposures. Non-targeted effects (NTEs)are responses which do not need a direct exposure to be expressed and are particularly significant at low energy radiations. Although NTEs of ionizing radiation are well documented, there are scarcely any studies on non-targeted responses such as bystander effect (BE)after exposure to non-ionizing radiation. The main goal of this research is to study possible RFR-induced BE. Material and methods: Chinese hamster ovary cells were exposed to 900 MHz GSM RFR at an average specific absorption rate (SAR)of 2 W/kg for 4, 12 and 24 hours (h). To generate a uniformly distributed electromagnetic field and avoid extraneous RF exposures a cavity was desined and used. Cell membrane permeability, cell redox activity, metabolic and mitotic cell death and DNA damages were analyzed. Then the most effective exposure durations and statistically significant altered parameters were chosen to assess the induction of BE through medium transfer procedure. Furthermore, intra and extra cellular reactive oxygen species (ROS)levels were measured to assess the molecular mechanism of BE induced by non-ionizing radiation. Results: No statistically significant alteration was found in cell membrane permeability, cell redox activity, metabolic cell activity and micronuclei (MN)frequency in the cells directly exposed to RFR for 4, 12, or 24 h. However, RFR exposure for 24 h caused a statistically significant decrease in clonogenic ability as well as a statistically significant increase in olive moment in both directly exposed and bystander cells which received media from RFR-exposed cells (conditioned culture medium; CCM). Exposure to RFR also statistically significant elevated both intra and extra cellular levels of ROS. Conclusion: Our observation clearly indicated the induction of BE in cells treated with CCM. To our knowledge, this is the first report that a non-ionizing radiation (900 MHz GSM RFR)can induce bystander effect. As reported for ionizing radiation, our results proposed that ROS can be a potential molecule in indirect effect of RFR. On the other hand, we found the importance of ROS in direct effect of RFR but in different ways

Enterolactone: A novel radiosensitizer for human breast cancer cell lines through impaired DNA repair and increased apoptosis

Introduction Radiotherapy is a potent treatment against breast cancer, which is the most commonly diagnosed cancer among women. However, the emergence of radioresistance due to increased DNA repair leads to radiotherapeutic failure. Applying polyphenols combined with radiation is a more promising method leading to better survival. Enterolactone, a phytoestrogenic polyphenol, has been reported to inhibit an important radioresistance signaling pathway, therefore we conjectured that enterolactone could enhance radiosensitivity in breast cancer. To assess this hypothesis, radiation response of enterolactone treated MDA-MB-231 and T47D cell lines and corresponding cellular mechanisms were investigated. Methods Cytotoxicity of enterolactone was measured via MTT assay. Cells were treated with enterolactone before X-irradiation, and clonogenic assay was used to evaluate radiosensitivity. Cell cycle distribution and apoptosis were measured by flow cytometric analysis. In addition, DNA damages and corresponding repair, chromosomal damages, and aberrations were assessed by comet, micronucleus, and cytogenetic assays, respectively. Results Enterolactone decreased the viability of cells in a concentration- and time dependent manner. Enterolactone significantly enhanced radiosensitivity of cells by abrogating G2/M arrest, impairing DNA repair, and increasing radiation-induced apoptosis. Furthermore, increased chromosomal damages and aberrations were detected in cells treated with enterolactone combined with X-rays than X-ray alone. These effects were more prominent in T47D than MDA-MB-231 cells. Discussion To our knowledge, this is the first report that enterolactone is a novel radiosensitizer for breast cancer irrespective of estrogen receptor status. Authors propose enterolactone as a candidate for combined therapy to decrease the radiation dose delivered to patients and subsequent side effects. © 2016 Elsevier Inc