Abstract: Objective: To investigate the enhancive effect of N,N'-dinitrosopiperazine (DNP) on induced carcinogenesis in nasal and/or nasopharyngeal epithelia among TgN(p53mt-LMP1)/HT transgenic mice to examine the underlying mechanism for the development of nasopharyngeal carcinoma (NPC). Methods: TgN(p53mt-LMP1)/HT transgenic mice and the same strain of C₅₇BL/6J wild-type mice both at the age of 5 months were randomly divided into 2 groups in parallel, respectively, i.e., TgN(p53mt-LMP1)/HT cancerous lesion-inducing group (TI), TgN(p53mt-LMP1)/HT control group (TC), C₅₇BL/6J cancerous lesion-inducing group (CI), and C₅₇BL/6J control group (CC). TI and CI mice were treated only with DNP for 16 weeks, twice each week, while TC and CC mice were given the same volume of saline as controls. At the end of treatment, animals were sacrificed to collect epithelial tissue samples from nasal cavity and nasopharynx for pathohistological evaluation by haematoxylin and eosin (HE) staining and for determination on the expression of TRAF2, c-Jun, and p16 by immunohistochemistry. Results: Atypical hyperplasia was more significant in the samples of TI than in those of TC, CI, and CC, with the rates of lesions being 90%, 10%, 0, and 0 (P<0.01) respectively, though DNP was used alone in a much shortened inducing period at less dosage and without the use of carcinogenic promoter 12-O-tetradecanoylphorbol-13-acetate as usual. The expressions of tumor necrosis factor (TNF) receptor-associated factor 2 (TRAF2) and c-Jun in these samples were significantly up-regulated in TI (P<0.01), while the expression of p16 was significantly lower in TI than in the other groups (P<0.01). Conclusion: TgN(p53mt-LMP1)/HT mice hold inherited constitutional defect in immune surveillance function, which can be aggravated by environmental carcinogens, such as DNP used even though in a much less strength. The enhanced carcinogenesis-inducing effect of DNP on TgN(p53mt-LMP1)/HT mice should be closely associated with abnormal signaling of activator protein-1 (AP-1) pathway, especially up-regulated expressions of TRAF2 and c-Jun, and down-regulated expression of p16.

[1] Berry, A., Goodwin, M., Moran, C.L., Chambers, T.C., 2001. AP-1 activation and altered AP-1 composition in association with increased phosphorylation and expression of specific Jun and Fos family proteins induced by vinblastine in KB-3 cells. Biochem. Pharmacol., 62(5): 581-591.

[2] Cai, H.Y., Li, B.S., Chen, N.Y., Ye, Y.L., Li, Z.Q., 1985. Experimental study on the model of precancerous lesion in nasal and nasopharyngeal mucosae and the blocking effect of vitamin A acid on the lesion. Chin. J. Cancer, 4(5):199-201 (in Chinese).

[3] Cao, Y., Hu, Z., Tao, Y.G., Tang, F.Q., Wang, H., Luo, F.J., Yi, W., 2002. EB virus-encoded latent membrane protein 1 activates the JNK signaling pathway via a mechanism involving TRADD and TRAF in nasopharyngeal carcinoma cell. Progr. Biochem. Biophys., 29(4):562-566 (in Chinese).

[4] Chang, H.W., Chan, A., Kwong, D.L., Wei, W.I., Sham, J.S., Yuen, A.P., 2003. Evaluation of hypermethylated tumor suppressor genes as tumor markers in mouth and throat rinsing fluid, nasopharyngeal swab and peripheral blood of nasopharygeal carcinoma patient. Int. J. Cancer, 105(6):851-855.

[5] He, Y.C., Tian, D.F., 2005. The structure of Epstein-Barr virus latent membrane protein 1 and its signal transduction network. J. Med. Postg., 18(9):837-840, 844 (in Chinese).

[6] He, Y.C., Tian, D.F., Tian, T., Lu, F.G., Liu, Y.Q., 2005. Expression of exogenous mutant p53 and latent membrane protein 1 genes in the tissues of the different organs of transgenic mice. Chin. J. Clin. Rehab., 9(42):99-101 (in Chinese).

[7] He, Y.C., Tian, D.F., Lu, F.G., Mao, J.F., 2006. Construction of transgenic mice containing human mutant p53 and EB virus latent membrane protein 1. J. Fourth Mil. Med. Univ., 27(1):6-9 (in Chinese).

[8] He, Y.C., Tian, D.F., Lu, F.G., Liu, H.P., 2008a. Association of cell proliferation in nasal and nasopharyngeal epithelia with expressive activity of apoptosis-related genes in TgN(p53mt-LMP1)/HT mice. Carcinogenesis, Teratogenesis & Mutagenesis (CTM), 20(5):363-366 (in Chinese).

[9] He, Y.C., Tian, D.F., Liu, H.P., Lu, F.G., Wu, X.Z., 2008b. Analysis on the biological phenotype and cell cycle distribution characteristics of nasal and/or nasopharyngeal epithelia of TgN(p53mt-LMP1)/HT mice. J. Med. Postg., 21(9):920-923, 928 (in Chinese).

[10] Johnson, A.C., Murphy, B.A., Matelis, C.M., Rubinstein, Y., Piebenga, E.C., Akers, L.M., Neta, G., Vinson, C., Birrer, M., 2000. Activator protein-1 mediates induced but not dasal epidermal growth factor receptor gene expression. Mol. Med., 6(1):17-27.

[11] Li, B.S., Cai, H.Y., 1987. Morphological observation on the precancerous lesion experimentally induced in nasal mucosa of rats. J. Sun Yatsen Univ. (Med. Sci.), 8(1): 27-30, 86 (in Chinese).

[12] Min, H.Q., Hong, M.H., Guo, X., 2003. Nasopharyngeal Carcinoma, 1st Ed. Chinese Medicinal Science and Technology Press, Beijing, China, p.116 (in Chinese).

[13] Song, X., Tao, Y.G., Tan, Y.N., Li, M.J., Deng, X.Y., Wu, Q., Cao, Y., 2004. Formation of c-Jun and JunB activating heterologous dimer by the Epstein-Barr virus-encoded latent membrane protein 1 (LMP1). Science in China (Series C), 34(4):325-334 (in Chinese).

[14] Tang, F.Q., Zhang, C.Y., Tian, D.F., Yin, C.N., 1999. Survey on the distribution of pathological physique patterns among the population at high risk of nasopharyngeal carcinoma and their possible association with the immune function of these hosts. Chin. J. Prac. TCM West Med., 12(19):1161-1162.

[15] Xue, Q., Sano, T., Kashiwabara, K., Saito, M., Oyama, T., Nakajima, T., 2002. Aberrant expression of pRb, p16, p14ARF, MDb12, p21 and p53 in stage I adenocarciormas of the lung. Pathol. Int., 52(2):103-109.