Roles of phosphatidylinositol 3 kinase in silica-induced DNA double strand breaks damage repair in human embryo lung fibroblasts

Abstract

To study the role of Phosphatidylinositol 3 kinase (PI3K) in silica-induced DNA double strand break repair in human embryo lung fibroblasts (HELF). 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. 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%. 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.