[The promotive effects of N-nitrosopiperidine on the malignant transformation of the immortalized esophageal epithelium induced by human papillomavirus].

BACKGROUND

Study on the promotive effects of N-nitrosopiperidine on carcinogenesis process was performed, based on the immortalization of human fetal esophageal epithelium induced by human papillomavirus (HPV) 18E6E7 genes.

METHODS

The immortalized esophageal epithelium SHEE was induced by HPV18E6E7. The cells at 17th passages were cultured in 50 ml flasks. The N-nitrosopiperidine (NPIP) 0, 2, 4, 8 mmol/L added to the cultured medium of SHEE cells for 3 weeks. The morphology, proliferation and apoptosis of the cells were studied by phase contrast microscopy and flow cytometry. Modal number of chromosomes was analyzed by standard method. Tumorigenicity of the cells was assessed by soft agar colony formation and by transplantation of cells into nude mice. Expression of HPV was detected by Western blot.

RESULTS

When cells were exposed to high concentration (8 mmol/L) of NPIP, cell death was increased, leaving a few live cells. In normal cultural medium instead of NPIP proliferative status of the cells restored after 4 weeks and the cells progressed to the proliferation stage with continuous replication and atypical hyperplasia. At the end of the 8th week, the cells appeared with large colonies in soft-agar and tumor formation in transplanted nude mice. When the cells were cultured in 2, 4 mmol/L NPIP the doubling passage was delayed and without tumor formation in transplanted nude mice. Modal number of chromosomes was 61-65, in 8 mmol/L NPIP group and control group, 56-61. Expression of HPV18 appeared in experimental and control groups.

CONCLUSION

NPIP promotes malignant change of the immortalized esophageal epithelial cells induced by HPV18E6E7. HPV18E6E7 synergy with NPIP will accelerate malignant transformation in esophageal epithelium.