| ObjectiveTo investigate the effect of 280nm Light Emitting Diode-Ultraviolet（LED-UV）on the formation of reactive oxygen species and the change of mitochondrial membrane potential on human leukemia HL-60 cells,and research the relation between HL-60 cell apoptosis and mitochondrial membrane potential and generation of reactive oxygen species.MethodsLogarithmic growth of HL-60 cells as the research object,280nm LED-UV ultraviolet light as a light source,they were irradiated with different doses（J/m2）.The experiment was divided into two groups,control group and experimental group.Group A（0j/m2）was the control group,the experimental group divided into B-E subgroup（8J/m2,15J/m2,30J/m2,60J/m2）.After irradiation,all cells were cultured in IMDM culture containing 10%fetal bovine serum and incubated for 2 hours at 37℃,5%CO2 and saturated humidity condition.JC-1 staining and flow cytometry were used to detect the changes of mitochondrial membrane potential,flow cytometry and Annexin V-FITC/PI double staining to detect apoptosis,DCFH-DA（2,7dichiorofluorescein diaceate）and fluorescence microscopy to detect the change of reactive oxygen.Results1.HL-60 cell growth state:the control group cells were round and bright,regular shape and arranged closely.When the 280nm LED-UV was 8J/m2,the cells began to appear shrinkage,with the dose increasing,brightness reduction,cell shape became irregular.There were a lot of debris,especially in the group D and group E.2.The apoptosis rate of HL-60 cells:group A was（3.01+0.72）%,group B-E were（7.45+0.31）%、（11.86+0.56）%、（21.29+0.25）%、（7.80+0.14）%.Among five groups had significant difference（F=3314.97,P<0.01）.The group D apoptosis rate was the highest,while the group E was significantly lower than that in group D（P<0.01）.3.The changes of mitochondrial membrane potential of HL-60 cells:group A was126.22+6.81,group B-E were 97.86+2.25、82.65+4.22、72.35+3.12、80.28+3.52,The changes of mitochondrial membrane potential among five groups had significant difference（F = 623.09,P<0.01）.But there were no singnificant difference between group C and group E（P>0.05）.The mitochondrial membrane potential of group D was the lowest,while group E was significantly higher than that in group D（P<0.01）.4.The changes of HL-60 cells reactive oxygen species:group A was（1.21±0.11）%,group B-E were（8.05±0.81）%、（18.56±0.31）%、（29.80±0.15）%、（17±0.22）%,Among five groups had significant difference（F=12502.94,P<0.01）.The group D ROS generation rate was the highest,while the group E was significantly lower than that in group D（P<0.01）.5.The relationship between HL-60 apoptosis and mitochondrial membrane potential:there is a highly negative correlation between cell apoptosis rate and mitochondrial membrane potential,r=-0.82,P<0.01.6.The relationship between HL-60 apoptosis and reactive oxygen species:there is a highly positive correlation between cell apoptosis rate and reactive oxygen species,r=0.94,P<0.01.Conclusions1.280nm LED-UV can induce apoptosis in HL-60 cells,in the range of 8-30J/m2the apoptosis rate was in a dose-dependent manner.The apoptosis rate was the highest when the dose was 30J/m2,while lower in dose of 60J/m2.2.280nm LED-UV can make the mitochondrial membrane potential of HL-60cells decreased,in the range of 0-30J/m2,they were in a dose-dependent manner.At the dose of 30J/m2,the mitochondrial membrane potential was the lowest,while significantly increased in 60J/m2.3.280nm LED-UV can make the HL-60 cell reactive oxygen species increased.In the range of 0-30J/m2,it is in a dose dependent manner.At the dose of 30J/m2,the reactive oxygen species generated the highest,while significantly increased in 60J/m2.4.There is a highly negative correlation between cell apoptosis rate and mitochondrial membrane potential,the higher of the mitochondrial membrane potential,the lower of the rate of cell apoptosis.5.There is a highly positive correlation between cell apoptosis rate and reactive oxygen species,the higher of the reactive oxygen species,the higher of the rate of cell apoptosis. |
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