Light-emitting diode photomodulation of uterine adenocarcinoma cells inhibited angiogenesis capacity via the regulation of exosome biogenesis.

This study was conducted to investigate the inhibitory effects of light-emitting diodes (LEDs) on exosome biogenesis and angiogenesis capacity in Ishikawa endometrial cancer cells. To this end, cells were exposed to different energy densities (fluences) of 4, 8, 16, 32, and 64 J/cm2 for 5 days (once every 24 h), and cell viability was determined using an MTT assay. Based on data from the MTT panel, cells were exposed to 4 and 16 J/cm2 for subsequent analyses. Exosome biogenesis was also monitored via monitoring the expression of CD63, ALIX, and Rab27a and b. The size and morphology of exosomes in the supernatant were measured using scanning electron microscopy (SEM), and dynamic light scattering (DLS). Using Transwell insert, the migration capacity of these cells was studied. The angiogenic effects of irradiated Ishikawa cell secretome at different fluences were monitored on human endothelial cells using in vitro tubulogenesis. Results indicated LED can reduce the viability of Ishikawa cells in a dose-dependent manner. According to our data, 4 and 64 J/cm2 groups exhibited minimum and maximum cytotoxic effects compared to the control cells. Data revealed a close proportional relationship between the power of laser and exosome average size compared to the non-treated control cells (p < 0.05). Real-time PCR analysis showed the suppression of Rab27b and up-regulation of Rab27a in irradiated cells exposed to 4 and 16 J/cm2 (p < 0.05). These effects were evident in the 16 J/cm2 group. Likewise, LED can inhibit the migration of Ishikawa cells in a dose-dependent manner (p < 0.05). Tubulogenesis activity of endothelial cells was suppressed after incubation with the secretome of irradiated Ishikawa cells (p < 0.05). These data showed tumoricidal properties of LED irradiation on human adenocarcinoma Ishikawa cells via the inhibition of exosome biogenesis and suppression of angiogenesis capacity.

Expression of recombinant G-CSF receptor domains and their inhibitory role on G-CSF function.

BACKGROUND AND PURPOSE: Granulocyte colony-stimulating factor (G-CSF) is routinely used in combination with chemotherapy to battle neutropenia. However, studies suggest that this chemokine may increase the risk of metastasis and malignancy in many cancers. To counteract the adverse effects of G-CSF in cancer, antibodies have been used to block its action. However, antibodies are large and complex molecules which makes their production expensive. Thus in this study, we aim to construct different structure variants of the G-CSF receptor containing different domains and select the best variant that prevents the adverse actions of this chemokine. These novel structures are smaller than antibodies and easier to produce. EXPERIMENTAL APPROACH: Different domains of the G-CSF receptor were designed and cloned into the pET28a expression vector. These recombinant receptor subunits were then expressed in Escherichia coli and purified using standard affinity chromatography techniques. Interaction of recombinant receptor subunits with G-CSF was assessed using enzyme-linked immunosorbent assay and NFS60 cells. FINDINGS / RESULTS: Two recombinant receptor subunits containing D1 + D2 + D3 domains and D2 domain showed the strongest inhibitory activity to G-CSF. CONCLUSION AND IMPLICATIONS: These novel recombinant receptor variants could be candidates for further studies in the development of novel therapeutics.