[Antisense ERCC1 RNA decreases the repair capability of damaged DNA in lung cancer cells induced by benzo[a]pyrene].
OBJECTIVE
To investigate the effect of ERCC1 gene on the repair capability of damaged DNA in lung cancer A549 cells induced by benzo[a]pyrene.


METHODS
Recombinant plasmid expressing ERCC1 antisense RNA was constructed and transfected into A549 cells by Lipofectin reagent. The stable-transfected cell colonies were selected by hygromycin. Cell viability was determined by the MTT assay. The level of ERCC1 mRNA was measured by Northern Blot analysis. Single cell gel electrophoresis assay was applied to determine the cellular DNA damage and fifty cells for each group were counted.


RESULTS
Seven positive colonies expressing ERCC1 antisense RNA were screened. There was no growth rate difference between the antisense-transfected cells and the parental cells. The endogenous mRNA level in transfected colonies decreased in varied degrees, i.e. 12% approximately 86% of that of the parental cells in Northern Blot assay. After 24 h treatment of 10 micro mol/l benzo[a]pyrene, the repair capability for DNA damage in transfected colonies was reduced to 29% approximately 71% of that of the parental cells. Also, a statistically significant correlation was observed between expression of ERCC1 mRNA and repair capability (r = 0.84).


CONCLUSIONS
Antisense ERCC1 RNA decreased the repair capability for damaged DNA in lung cancer cells induced by benzo[a]pyrene.
Wu
Xiao-ming
XM
Institute of Environmental Medicine, Tongji Medical College of Huazhong University of Science and Technology, Wuhan 430030, China.
Zhou
Yi-kai
YK
Xu
Shun-qing
SQ
Hao
Qiao-ling
QL
Ren
Shu
S
D001564:Benzo(a)pyrene / Q000633:toxicity*; D045744:Cell Line, Tumor; D004249:DNA Damage / Q000187:drug effects; D004260:DNA Repair / Q000187:drug effects*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D004720:Endonucleases / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D006801:Humans; D008175:Lung Neoplasms / Q000473:pathology*; D010957:Plasmids; D016372:RNA, Antisense / Q000494:pharmacology*; D012333:RNA, Messenger / Q000378:metabolism; D012097:Repressor Proteins; D014162:Transfection
D004268:DNA-Binding Proteins; D016372:RNA, Antisense; D012333:RNA, Messenger; D012097:Repressor Proteins; D001564:Benzo(a)pyrene; C071447:ERCC1 protein, human; D004720:Endonucleases