Changes in Bcl-2 and Bax Expression in Rat Tongue during 4-Nitroquinoline 1-oxide-induced Carcinogenesis

Abstract

Bax is considered as a main effector of apoptosis. Bax forms homodimers and also heterodimers with Bcl-2. The function of the Bax-Bax dimer in active cell death is antagonized by Bax-Bcl-2 heterodimers. Thus, the ratio of Bcl-2 and Bax should control the susceptibility of cells to those stimuli that induce apoptotic cell death. An increase in apoptotic change has been shown in many carcinomas. In the present study, the changes in Bcl-2 and Bax expression in the tissue during carcinogenic transformation were examined immunohistochemically by means of the 4-nitroquinoline 1-oxide (4NQO)-induced carcinoma model. Animals were divided into 7 groups of 10 rats each, and given 50 ppm 4NQO solution as drinking water for 4, 8, 12, 16, 20, or 24 weeks. Ten animals were used as controls. Gradual increases in the numbers of Bcl-2- and Bax-positive cells were shown corresponding to the progression of experimental carcinogenesis. Statistically significant differences in Bcl-2 and Bax expression were demonstrated between control and four-week treatment groups (p < 0.01), and between control and eight-week treatment groups (p < 0.05), respectively. Levels of both proteins remained high after the period of dysplastic change of the epithelium. In conclusion, Bcl-2 and Bax are involved in the progression of 4NQO-induced carcinoma.

Keywords

Bcl-2,Bax 4-nitroquinoline 1-oxide (4NQO)experimental carcinogenesis squamous cell carcinoma

References

Abiko Y. , Ohuchi T. , Nakahata H. , Sadaoka T. , Kanda M. , Kaku T. ( 1994). In situ labeling of nuclear DNA fragmentation in normal oral epithelia and squamous cell carcinomas. Jpn J Oral Biol 36:67-70.

Akao Y. , Otsuki Y. , Kataoka S. , Ito Y. , Tsujimoto Y. (1994). Multiple subcellular localization of bcl-2: detection in nuclear outer membrane, endoplasmic reticulum membrane, and mitochondrial membranes. Cancer Res 54:2468-2471.

Apolinario RM , van der Valk P. , de Jong JS , Deville W. , van Ark-Otte J. , Dingemans AM , et al. (1997). Prognostic value of the expression of p53, bcl-2, and bax oncoproteins, and neovascularization in patients with radically resected non-small-cell lung cancer. J Clin Oncol 15:2456-2466.

Arends MJ , Wyllie AH (1991). Apoptosis: mechanisms and roles in pathology. Int Rev Exp Pathol 32:223-254.

Bakhshi A. , Jensen JP , Goldman P. , Wright JJ , McBride OW , Epstein AL , et al. (1985). Cloning the chromosomal breakpoint of t(14;18) human lymphomas: clustering around JH on chromosome 14 and near a transcriptional unit on 18. Cell 41:899-906.

Chen X. , Ko LJ , Jayaraman L. , Prives C. (1996). p53 levels, functional domains, and DNA damage determine the extent of the apoptotic response of tumor cells. Genes Dev 10 :2438-2451.

Eguchi Y. , Ewert DL , Tsujimoto Y. (1992). Isolation and characterization of the chicken bcl-2 gene: expression in a variety of tissues including lymphoid and neuronal organs in adult and embryo. Nucleic Acids Res 20:4187-4192.

Hockenbery DM , Zutter M. , Hickey W. , Nahm M. , Korsmeyer SJ (1991). Bcl2 protein is topographically restricted in tissues characterized by apoptotic cell death. Proc Natl Acad Sci USA 88:6961-6965.

Ikegaki N. , Katsumata M. , Minna J. , Tsujimoto Y. (1994). Expression of bcl-2 in small cell lung carcinoma cells. Cancer Res 54:6-8.

10.

Jordan RC , Catzavelos GC , Barrett AW , Speight PM (1996). Differential expression of bcl-2 and bax in squamous cell carcinomas of the oral cavity. Oral Oncol, Eur J Cancer 32(B) :394-400.

11.

Krajewski S. , Tanaka S. , Takayama S. , Schibler MJ , Fenton W. , Reed JC ( 1993). Investigation of the subcellular distribution of the bcl-2 oncoprotein: residence in the nuclear envelope, endoplasmic reticulum, and outer mitochondrial membranes. Cancer Res 53:4701-4714.

12.

Laemmli UK (1970). Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227 :680-685.

13.

Leek RD , Kaklamanis L. , Pezzella F. , Gatter KC , Harris AL (1994). bcl-2 in normal human breast and carcinoma, association with oestrogen receptor-positive, epidermal growth factor receptor-negative tumours and in situ cancer. Br J Cancer 69:135-139.

14.

Maltzman W. , Czyzyk L. (1984). UV irradiation stimulates levels of p53 cellular tumor antigen in nontransformed mouse cells. Mol Cell Biol 4:1689-1694.

15.

Miyashita T. , Reed JC ( 1995). Tumor suppressor p53 is a direct transcriptional activator of the human bax gene. Cell 80 :293-299.

16.

Miyashita T. , Krajewski S. , Krajewska M. , Wang HG , Lin HK , Liebermann DA , et al. (1994). Tumor suppressor p53 is a regulator of bcl-2 and bax gene expression in vitro and in vivo. Oncogene 9:1799-1805.

17.

Nakagawa K. , Yamamura K. , Maeda S. , Ichihashi M. (1994). bcl-2 expression in epidermal keratinocytic diseases . Cancer 74:1720-1724.

18.

Negrini M. , Silini E. , Kozak C. , Tsujimoto Y. , Croce CM (1987). Molecular analysis of mbcl-2: structure and expression of the murine gene homologous to the human gene involved in follicular lymphoma . Cell 49:455-463.

19.

Oltvai ZN , Milliman CL , Korsmeyer SJ (1993). Bcl-2 heterodimerizes in vivo with a conserved homolog, Bax, that accelerates programmed cell death. Cell 74:609-619.

20.

Osugi Y. (1996). p53 expression in various stages of 4-nitroquinoline 1-oxide induced carcinoma in the rat tongue. J Osaka Dent Univ 30:29-35.

21.

Pezzella F. , Turley H. , Kuzu I. , Tungekar MF , Dunnill MS , Pierce CB , et al. (1993). bcl-2 protein in non-small-cell lung carcinoma . N Engl J Med 329:690-694.

22.

Pilotti S. , Collini P. , Rilke F. , Cattoretti G. , Del Bo R. , Pierotti MA (1994). Bcl-2 protein expression in carcinomas originating from the follicular epithelium of the thyroid gland. J Pathol 172:337-342.

23.

Polyak K. , Waldman T. , He T-C. , Kinzler KW , Vogelstein B. (1996). Genetic determinants of p53-induced apoptosis and growth arrest. Genes Dev 10:1945-1952.

24.

Saegusa M. , Takano Y. , Hashimura M. , Shoji Y. , Okayasu I. (1995). The possible role of bcl-2 expression in the progression of tumors of the uterine cervix. Cancer 76:2297-2303.

25.

Silvestrini R. , Veneroni S. , Daidone MG , Benini E. , Boracchi P. , Mezzetti M. , et al. (1994). The Bcl-2 protein: a prognostic indicator strongly related to p53 protein in lymph node-negative breast cancer patients . J Natl Cancer Inst 86:499-504.

26.

Stenman G. (1981). Malignancy of 4NQO-induced oral squamous cell carcinomas in the rat. Acta Otolaryngol 92:557-561.

27.

Tahara E. (1993). Molecular mechanism of stomach carcinogenesis . J Cancer Res Clin Oncol 119:265-272.

28.

Tsujimoto Y. , Croce CM (1986). Analysis of the structure, transcripts, and protein products of bcl-2, the gene involved in human follicular lymphoma. Proc Natl Acad Sci USA 83 :5214-5218.

29.

Tsujimoto Y. , Cossman J. , Jaffe E. , Croce CM (1985). Involvement of the bcl-2 gene in human follicular lymphoma. Science 228:1440-1443.

30.

Vaux DL , Cory S. , Adams JM (1988). Bcl-2 gene promotes haemopoietic cell survival and cooperates with c-myc to immortalize pre-B cells. Nature 335:440-442.

31.

Yin X.-M. , Oltvai ZN , Korsmeyer SJ (1994). BH1 and BH2 domains of Bcl-2 are required for inhibition of apoptosis and heterodimerization with Bax. Nature 369:321-323.