[Roles of phosphatidylinositol 3 kinase in silica-induced DNA double strand breaks damage repair in human embryo lung fibroblasts].
OBJECTIVE
To study the role of Phosphatidylinositol 3 kinase (PI3K) in silica-induced DNA double strand break repair in human embryo lung fibroblasts (HELF).


METHODS
Control HELF cells and DN-Deltap85 (HELF transfected with Dominant negative mutant of PI3K) were treated with 200 microg/ml silica for different times. The expression levels of phosphor-H2AX (H2AX), Ku70, Ku80 and DNA-PKcs were determined by Western blot. Furthermore, DNA double strand breaks were measured by neutral comet assay after cells were treated with 200 microg/ml silica for 0, 12 and 24 h.


RESULTS
After treatment with 200 microg/ml silica for different times, the levels of H2AX were increased in a time-dependent manner and the expression levels of H2AX were obviously suppressed in DN-Deltap85 compared with control cells. The levels of Ku70 and Ku80 were also significantly suppressed in DN-Deltap85 (0.37 +/- 0.14, 0.55 +/- 0.17) compared with control cells (0.58 +/- 0.09, 0.95 +/- 0.21) after treatment with 200 microg/ml silica for 12 h (P < 0.05). Both the percentage of tail DNA in HELF and DN-Deltap85 increased significantly at 12 h (9.78 +/- 1.15, 11.79 +/- 4.90) compared with groups without treatment with silica (2.40 +/- 0.69, 3.31 +/- 1.35) and then decreased at 24 h (4.19 +/- 0.47, 7.58 +/- 4.32), but only the decrease of HELF at 24 h was significant compared with HELF at 12 h (P < 0.05). DNA repair competence of HELF was 75.74% and that of DN-Deltap85 declined to 49.64%.


CONCLUSIONS
Silica dust can induce DNA double strand breaks in human embryo lung fibroblasts. PI3K might play a role in silica-induced DNA double strand break repair by regulating the expression levels of Ku70 and Ku80.
Liu
Hai-Feng
HF
National Institute of Occupation Health and Poison Control, Chinese Center for Disease Control and Prevention, Beijing 100050, China.
Zhang
Feng-Mei
FM
Liu
Bing-Ci
BC
Jia
Xiao-Wei
XW
Ye
Meng
M
D034961:Antigens, Nuclear / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000378:metabolism; D002478:Cells, Cultured; D020552:Comet Assay; D053903:DNA Breaks, Double-Stranded; D004249:DNA Damage; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000378:metabolism; D005347:Fibroblasts / Q000201:enzymology*; D006657:Histones / Q000378:metabolism; D006801:Humans; D000072200:Ku Autoantigen; D008168:Lung / Q000166:cytology; D058539:Phosphatidylinositol 3-Kinase / Q000378:metabolism*; D012822:Silicon Dioxide / Q000633:toxicity*
D034961:Antigens, Nuclear; C109198:CIB1 protein, human; D002135:Calcium-Binding Proteins; D004268:DNA-Binding Proteins; C468783:H2AX protein, human; D006657:Histones; D012822:Silicon Dioxide; D058539:Phosphatidylinositol 3-Kinase; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen