Impact on U251 glioma cell radiosensitivity and CFL1 level via inhibiting cell motility regulator ROCKII of RhoA-RockII-CFL1 pathway.

Abstract

e14630 Background: In response to ionizing radiation, cofilin1 (CFL1) is essential for actin dynamics and tumor metastasis. In our previous study, CFL1 was revealed up-regulated in radioresistant astrocytomas of surgical samples and positively correlated with glioma cell radioresistance. Methods: The present study aims to clarify the potential mechanism of CFL1-mediated radioresistance and to find new molecular targets to improve the radiosensitivity of U251 glioma cell. Radioresistant U251 (RR-U251) was established by serially irradiation with 5Gy until 60 Gy of irradiation was accumulated on human U251 glioma cell. Y-27632, PAK1-shRNA and PAK1-shRNA were respectively utilized to inhibit CFL1 upstream regulators in normal U251 and RR-U251 cells. CFL1 expression level and the alteration of radiosensitivity were evaluated; the cell radiosensitivity was assessed through cell vitality, invasion and migration experiments after irradiation. Results: 20 ¦Ìmol/L Y-27632 was screened and administrated to inhibit ROCKII in glioma cells followed by 5Gy radiation treatment. The CFL1 level was reduced in both normal U251 and RR-U251 cells. Meanwhile, cell viability, migration and invasion abilities significantly decreased in RR-U251 cells. After being transfected with PAK1-shRNA, cell radiosensitivity were significantly improved in both normal U251 and RR-U251 cells; while CFL1 level showed no significant difference after PAK1 silence . After inhibiting MRCK¦Á expression, cell viability, migration and invasion capabilities significantly reduced in both normal and radioresistant cells; while CFL1 levels showed no significant decrease. Conclusions: The results of the present study indicate that inhibiting ROCKII of RhoA-RockII-CFL1 pathway could improve radiosensitivity of radioresistant glioma cells and simultaneously down-regulate CFL1 expression; while the inhibiton of PAK1 or MRCK¦Á ameliorated the radiosensitivity with no significant regulation effect on CFL1. In conclusion, the present study emphasize the involvement of ROCKII in CFL1-mediated radioresistant phenotype and provide potential biomarkers for improvement of further clinical radiotherapy.