Genotoxicity and reduced heat shock protein 70 in human airway smooth muscle cells exposed to cigarette smoke extract

Abstract

Cigarette smoke is associated with the development of several diseases, such as chronic obstructive pulmonary disease (COPD). The purpose of this study was to investigate genotoxicity and heat shock protein 70 (Hsp70) in human airway smooth muscle cells (HASMCs) exposed to cigarette smoke extract (CSE). HASMCs was exposed to CSE with different doses for 24 h. The level of 8-hydroxydeoxyguanosine (8-OHdG) was determined by using HPLC-ECD, the DNA damage was analyzed by using comet assay, and apoptosis was examined by using Annexin-FITC/PI staining. The production of Hsp70 after CSE stimulation was tested. Results indicated that CSE significantly increased the level of 8-OHdG, DNA damage and cell apoptosis, and reduced the production of Hsp70. In particular, levels of Hsp70 were inversely correlated with 8-OHdG, DNA damage and cell apoptosis. It was concluded that cigarette smoke induced genotoxicity and decreased the production of cell protective protein Hsp70, which may contribute to the development of some airway diseases.

Keywords

airway smooth muscle / cigarette smoke extract / 8-hydroxydeoxyguanosine / DNA damage / heat shock protein 70

References

[1] RepineJE, BastA, LankhorstI. Oxidative stress in chronic obstructive pulmonary disease. Oxidative Stress Study Group. Am J Respir Crit Care Med, 1997, 156(2Pt1): 341-357

[2] LofrothG. Environmental tobacco smoke: overview of chemical composition and genotoxic components. Mutat Res, 1989, 222(2): 73-80

[3] PryorWA. Biological effects of cigarette smoke, wood smoke, and the smoke from plastics: the use of electron spin resonance. Free Radic Biol Med, 1992, 13(6): 659-676

[4] AgbajeIM, McVicarCM, SchockBC, et al.. Increased concentrations of the oxidative DNA adduct 7,8-dihydro-8-oxo-2-deoxyguanosine in the germ-line of men with type 1 diabetes. Reprod Biomed Online, 2008, 16(3): 401-409

[5] ChungKF, AdcockIM. Multifaceted mechanisms in COPD: inflammation, immunity, and tissue repair and destruction. Eur Respir J, 2008, 31(6): 1334-1356

[6] VayssierM, BanzetN, FrancoisD, et al.. Tobacco smoke induces both apoptosis and necrosis in mammalian cells: differential effects of HSP70. Am J Physiol, 1998, 275(4Pt1): L771-779

[7] HightowerLE. Heat shock, stress proteins, chaperones, and proteotoxicity. Cell, 1991, 66(2): 191-197

[8] WuT, ChenS, XiaoC, et al.. Presence of antibody against the inducible Hsp71 in patients with acute heat-induced illness. Cell Stress Chaperones, 2001, 6(2): 113-120

[9] SchroderHC, HassaneinHM, LauenrothS, et al.. Induction of DNA strand breaks and expression of HSP70 and GRP78 homolog by cadmium in the marine sponge Suberites domuncula. Arch Environ Contam Toxicol, 1999, 36(1): 47-55

[10] ZhangK, ShanL, RahmanMS, et al.. Constitutive and inducible thymic stromal lymphopoietin expression in human airway smooth muscle cells: role in chronic obstructive pulmonary disease. Am J Physiol Lung Cell Mol Physiol, 2007, 293(2): L375-382

[11] TwortC, Van BreemenC. Human airway smooth muscle in culture. Tissue Cell, 1988, 20(3): 339-344

[12] LeeSD, LeeDS, ChunYG, et al.. Cigarette smoke extract induces endothelin-1 via protein kinase C in pulmonary artery endothelial cells. Am J Physiol Lung Cell Mol Physiol, 2001, 281(2): L403-411

[13] HelbockHJ, BeckmanKB, ShigenagaMK, et al.. DNA oxidation matters: the HPLC-electrochemical detection assay of 8-oxo-deoxyguanosine and 8-oxo-guanine. Proc Natl Acad Sci USA, 1998, 95(1): 288-293

[14] PougetJP, DoukiT, RichardMJ, et al.. DNA damage induced in cells by gamma and UVA radiation as measured by HPLC/GC-MS and HPLC-EC and Comet assay. Chem Res Toxicol, 2000, 13(7): 541-549

[15] KindzelskiiAL, PettyHR. Ultrasensitive detection of hydrogen peroxide-mediated DNA damage after alkaline single cell gel electrophoresis using occultation microscopy and TUNEL labeling. Mutat Res, 1999, 426(1): 11-22

[16] HodgeGL, FlowerR, HanP. Optimal storage conditions for preserving granulocyte viability as monitored by Annexin V binding in whole blood. J Immunol Methods, 1999, 225(1-2): 27-38

[17] WuT, YuanY, WuY, et al.. Presence of antibodies to heat stress proteins in workers exposed to benzene and in patients with benzene poisoning. Cell Stress Chaperones, 1998, 3(3): 161-167

[18] CarnevaliS, PetruzzelliS, LongoniB, et al.. Cigarette smoke extract induces oxidative stress and apoptosis in human lung fibroblasts. Am J Physiol Lung Cell Mol Physiol, 2003, 284(6): L955-963

[19] LiuX, ConnerH, KobayashiT, et al.. Cigarette smoke extract induces DNA damage but not apoptosis in human bronchial epithelial cells. Am J Respir Cell Mol Biol, 2005, 33(2): 121-129

[20] ShenbergerJS, DixonPS. Oxygen induces S-phase growth arrest and increases p53 and p21(WAF1/CIP1) expression in human bronchial smooth-muscle cells. Am J Respir Cell Mol Biol, 1999, 21(3): 395-402

[21] PytelD, WysockiT, MajsterekI. Comparative study of DNA damage, cell cycle and apoptosis in human K562 and CCRF-CEM leukemia cells: Role of BCR/ABL in therapeutic resistance. Comp Biochem Physiol C Toxicol Pharmacol, 2006, 144(1): 85-92

[22] CheethamME, AndertonBH, JacksonAP. Inhibition of hsc70-catalysed clathrin uncoating by HSJ1 proteins. Biochem J, 1996, 319Pt1: 103-108

[23] HartlFU. Molecular chaperones in cellular protein folding. Nature, 1996, 381(6583): 571-579

[24] Vayssier-TaussatM, CamilliT, AronY, et al.. Effects of tobacco smoke and benzo[a]pyrene on human endothelial cell and monocyte stress responses. Am J Physiol Heart Circ Physiol, 2001, 280(3): H1293-1300

[25] GaoYJ, XiaoCF, ChenS, et al.. In vitro study on role of Hsp70 expression in DNA damage of human embryonic lung cells exposed to Benzo[a]pyrene. Biomed Environ Sci, 2004, 17(2): 144-152

[26] ChenYC, Lin-ShiauSY. Involvement of heat-shock protein 70 and P53 proteins in attenuation of UVC-induced apoptosis by thermal stress in hepatocellular carcinoma cells. Photochem Photobiol, 1999, 70(1): 78-86

[27] CaliniV, UraniC, CamatiniM. Overexpression of HSP70 is induced by ionizing radiation in C3H 10T1/2 cells and protects from DNA damage. Toxicol In Vitro, 2003, 17(5–6): 561-566

[28] LiS, ZhengJ, CarmichaelST. Increased oxidative protein and DNA damage but decreased stress response in the aged brain following experimental stroke. Neurobiol Dis, 2005, 18(3): 432-440

[29] HamzehM, SunaharaGI. In vitro cytotoxicity and genotoxicity studies of titanium dioxide (TiO(2)) nanoparticles in Chinese hamster lung fibroblast cells. Toxicol In Vitro, 2012, 27(2): 864-873

[30] TennenRI, HayeJE, WijayatilakeHD, et al.. Cell-cycle and DNA damage regulation of the DNA mismatch repair protein Msh2 occurs at the transcriptional and post-transcriptional level. DNA Repair (Amst), 2013, 12(2): 97-109