Abstract

Among short-term in vitro genotoxicity assays, micronucleus assays are rapid, inexpensive, and less labor-intensive system. We have undertaken a comparative study of sensitivity of cigarette smoke condensate(CSC) by general micronucleus(MN) assay and cytokinesis-block micronucleus(CBMN) assay. In this study, V79 Chinese hamster cells were employed to evaluate and compare the genotoxicity of CSC of Kentucky Reference Cigarette 2R4F by 2 kinds of in vitro MN assay methods. To determine the optimum concentration of cytochalasin B(CYB) to obtain the maximal number of binucleated cells for CBMN assay, triplicate cultures of growing cells were treated with CYB for 15 h. CYB treatments caused a concentration-dependent increase in cytotoxicity($1\~4{\mu}g/mL$) and proportion($0.25\~1\;{\mu}g/mL$) of binucleated cells. These data suggested that 1 ug/mL of CYB is as an optimum dose for CBMN assay in binucleated V79 cells. Short treatment(4 h) of CSC induced a micronucleated cells with a concentration-dependent response in the presence or absence of CYB, but CSC-induced MNs were weakened when S9 was present. Long treatments(19 h) of CSC also induced a significant increase MN formation with a concentration-dependent response. At a concentration of 75 ${mu}g/mL$, the MN cell frequencies of general MN assay and CBMN assay were $6.5\%\;and\;11.7\%$, respectively. Linear regression analysis revealed a good correlation in CBMN assay between a concentration of CSC and MN cell frequency. All these data indicated that CBMN assay is more sensitive to the induction CSC-induced MN than general MN assay.

주제어

#cigarette smoke condensate   #V79 cells   #genotoxicity assay   #micronucleus  

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참고문헌 (12)

1. Bombick, B. R., Murli, H., Avalos, J. T., Bombick, D. W., Morgan, W. T., Putnam, K.P. and Doolittle, D. J. (1997) Chemical and biological studies of a new cigarette that primarily heats tobacco. Part 2. In vitrotoxicity of mainstream smoke condensate. Food Chemical Toxicol. 36: 183-190 
2. Borenfreund, E. and Puerner, J. A. (1985)Toxicity determined in vitro by morphological alterations and neutral red absorption. Toxicol. Lett. 24: 119-124 
3. DeMarini, D. M. (1983) Genotoxicity of tobacco smoke and tobacco smoke condensate. Mutat. Res. 114: 59-89 
4. Doolittle, D. J., Lee, C. K., Ivett, J. L., Mirsalis, J. C., Riccio, E., Rudd, C. J., Burger, G. T.and Hayes A. W. (1990) Comparative studies on the genotixic activity of mainstream smoke condendsate from cigarettes which burn or only heat tobacco. Enviromentat and Motecutar Mutagenesis 15: 93-105 
5. Fenech, M. (1993) The cytokinesis block micro nucleus technique : A detailed description of the method and its application to geno toxicity studies in human polulations. Mutat. Res. 285: 35-44 
6. Kirsch Volders, M., Sofuni, T., Aartema, S.A., Albertini, A., Eastmond, D., Fenech, M., Ishidata, jr, M., Lorge, E., Norppa, H.,Surralles, J., Hude, W. and Wakata, A.(2000) Report from the in vitro micronucleus assay working group. Environmental and Molecular Mutagenesis 35: 167-172 
7. Roemer, E., Tewes, F. J., Meisgen, T. J., Veltel,D. J. and Carmines, E. L. (2002) Evaluation of the potential effects of ingredients added to cigarettes. Part 3: In vitro genotoxidty and cytotoxicity. Food Chem, Toxicol. 40: 105-111 
8. Veltel, D. and Hoheneder, A. (1996) Characterization of cigarette smoke induced micronucleic in vitro. Exp. Toxic. Pathot. 48:548-550 
9. Fenech, M. (2000) The in vitro micronucleus technique. Mutat. Res. 455: 81-95 
10. Ellard, S., Mohammed, Y., Dogra, S., Wolfel, C.,Doehmer, J. and Parry, J. M. (1991) The use of genetically engineered V79 Chinese hamster cultures expressing rat liver CYP1A1, 1A2and 2B1 cDNAs in micronucleus assays. Mutagenesis 6: 461-470 
11. Kirsch Volders, M., Elhajouji, A., Cundari, E. and VanHummelen, P. (1997) The in vitro micronucleus test : a multipoint assay to detect simutaneously mitotic delay, apoptosis, chromosome breakage, chromosome loss andnon disjunction. Mutat. Res. 392: 19-30 
12. Krishna, G., Kropko, M. L. and Theiss, J. C.(1989) Use of the cytokinesis block method for the analysis of micronuclei in V79 Chinese hamster lung cells : results with mitomycin C and cyclophosphamide. Mutat. Res. 222: 63-69 

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