NF-kappaB mediates the survival of human bronchial epithelial cells exposed to cigarette smoke extract.
BACKGROUND
We have previously reported that low concentrations of cigarette smoke extract induce DNA damage without leading to apoptosis or necrosis in human bronchial epithelial cells (HBECs), and that IL-6/STAT3 signaling contributes to the cell survival. Since NF-kappaB is also involved in regulating apoptosis and cell survival, the current study was designed to investigate the role of NF-kappaB in mediating cell survival in response to cigarette smoke exposure in HBECs.


METHODS
Both the pharmacologic inhibitor of NF-kappaB, curcumin, and RNA interference targeting p65 were used to block NF-kappaB signaling in HBECs. Apoptosis and cell survival were then assessed by various methods including COMET assay, LIVE/DEAD Cytotoxicity/Viability assay and colony formation assay.


RESULTS
Cigarette smoke extract (CSE) caused DNA damage and cell cycle arrest in S phase without leading to apoptosis in HBECs as evidenced by TUNEL assay, COMET assay and DNA content assay. CSE stimulated NF-kappaB -DNA binding activity and up-regulated Bcl-XL protein in HBECs. Inhibition of NF-kappaB by the pharmacologic inhibitor curcumin (20 microM) or suppression of p65 by siRNA resulted in a significant increase in cell death in response to cigarette smoke exposure. Furthermore, cells lacking p65 were incapable of forming cellular colonies when these cells were exposed to CSE, while they behaved normally in the regular culture medium.


CONCLUSIONS
The current study demonstrates that CSE activates NF-kappaB and up-regulates Bcl-XL through NF-kB activation in HBECs, and that CSE induces cell death in cells lacking p65. These results suggest that activation of NF-kappaB regulates cell survival following DNA damage by cigarette smoke in human bronchial epithelial cells.
Liu
Xiangde
X
Pulmonary, Critical Care and Sleep Medicine, Department of Internal Medicine, University of Nebraska Medical Center, Omaha, Nebraska, USA. srennard@unmc.edu.
Togo
Shinsaku
S
Al-Mugotir
Mona
M
Kim
Huijung
H
Fang
Qiuhong
Q
Kobayashi
Tetsu
T
Wang
Xingqi
X
Mao
Lijun
L
Bitterman
Peter
P
Rennard
Stephen
S
D001980:Bronchi / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D016328:NF-kappa B / Q000378:metabolism*; D010936:Plant Extracts / Q000494:pharmacology*; D012906:Smoke / Q000032:analysis*; D013638:Tars; D014026:Nicotiana / Q000737:chemistry*; D014028:Tobacco Smoke Pollution / Q000032:analysis*
D016328:NF-kappa B; D010936:Plant Extracts; D012906:Smoke; D013638:Tars; D014028:Tobacco Smoke Pollution; C024746:tobacco tar