Exosomes And Related MiRNAs In The "bystander Effect" Of Ultraviolet Radiation

Background and ObjectiveUltraviolet（UV）is a natural radiation source which is widespread.The researchers defined it as a class of non-ionizing radiation with a wavelength between 180 and400 nm.We can divide UV into UVC（200～280 nm）,UVB（280～320 nm）and UVA（320～400 nm）according to its wavelength.Researches find that UV can cause varying degrees of photo aging when irradiating cells.In addition to direct damage to irradiated cells,it can also produce indirect damage to adjacent cells,which is called bystander effects.Bystander effects（BE）is an interesting phenomenon referring to similar responses which is displayed by cells adjacent to irradiated cells or even cultured with the supernatant of irradiated cells.Those non-irradiated cells having similar responses induced by signals from irradiated cells are called bystander cells.Recent researches indicated that this phenomenon is possibly caused by micro RNAs（mi RNAs）.Mi RNAs can play essential roles in biological processes as it can be wrapped and exported into the extracellular environment in this process.Mi RNAs can be packed into extracellular vesicle（EVs）which can function as cell-to-cell transmitters and convey their contents in bystander effects and generally it regulates the biological features of recipient cells negatively.In this study,we chose two specific mi RNAs,mi R-4655-3p and mi R-769-5p screened by our research group,using mi RNA mimics for the functional verification test.In addition,we investigate the role of exosome-mediated mi RNA in bystander effect involved in acute photo damage.MethodsWe set up a UV-induced bystander effect model of human skin fibroblasts in vitro.Differentially expressed mi RNAs in irradiated cells,bystander cells and culture medium were identified by using microarray and quantitative real-time polymerase chain reaction（q RT-PCR）.Exosomes were extracted and transferred to induce bystander effects.In this study,we verified the mediating regulation of exosomes by extracting exosomes from supernatant of directly irradiated cells and transferring to non-irradiated cells.In addition,mi R-4655-3p and mi R-769-5p were overexpressed using mi R-4655-3p and mi R-769-5p mimics.Research methods included CCK-8,ROS kit,annexin V/PI method,q RT-PCR and Western blotting analysis.Results1.Exosomes induced bystander effects in ultraviolet irradiationIn acute ultraviolet irradiation,transferring exosomes from supernatant in dicrectly irradiated cells could induce photo damage showed by several biological indicators.CCK-8 showed a decrease on cell proliferation in bystander cells,which was significantly different from non-irradiated cells（P<0.05）.Results of ROS showed an increase on uorescence intensity in bystander cells,which was significantly different from non-irradiated cells（P<0.05）.Results of Annexin V-FITC/PI showed an increase on cell apoptosis in bystander cells,which was significantly different from non-irradiated cells（P<0.05）.2.Up-regulation of mi R-4655-3p and mi R-769-5p by mi RNA mimics in non-irradiated cells can induce photo damageUp-regulation of mi R-4655-3p and mi R-769-5p by mi RNA mimics in non-irradiated cells could induce photo damage showed by several biological indicators.Results of CCK-8 showed a decrease on cell proliferation in bystander cells,which was significantly different from non-irradiated cells（P<0.05）.Results of ROS showed an increase on uorescence intensity in bystander cells,which was significantly different from non-irradiated cells（P<0.05）.Results of Annexin V-FITC/PI showed an increase on cell apoptosis in bystander cells,which was significantly different from non-irradiated cells（P<0.05）.ConclusionTaken together,our research revealed that bystander effects could be induced by transferring exosomes.Morever,we demonstrated that mi R-4655-3p and mi R-769-5p were involved in bystander effects.