The genetic events of HPV-immortalized esophageal epithelium cells

We studied cytogenesis, telomere and telomerase, and c-myc, ras, bcl-2, and p53 genes of cells in the progressive process of immortal epithelial cells from embryonic esophagus induced by human papillomavirus (HPV). The SHEE c ell line, established by us, consist of immortalized epithelial cells from the embryonic esophagus induced by genes E6E7 of HPV type 18. It was in ini tial malignant transformation when cultivated over 60 passages without co-c arcinogens. Cells of the 10th, 31st, and 60th passages were represented in the progressive process within the immortal period. In these three stages o f the cell line, the modal number of chromosome and karyotypes were analyze d. The telomere length was assayed by Southern blot methods, and the telome rase activity was analyzed by hTR and hTERT assay. C-myc, p53, bcl-2, ras g enes were assayed by the multi-PCR method. The morphology of the 10th passa ge cells exhibited good differentiation, the 60th passage cells were relati vely poorly differentiated, and the 31st passage cells differentiated in tw o distinct ways. The growth characteristics of the 31st and 60th passage ce lls were weakened at contact-inhibition and anchorage-dependent growth. Kar yotypes of three cell passages belonged to hyperdiploid and hypotriploid wi th abnormal chromosomes +1, +3, +7, +9, +17, +18; del(1)(p32); der(4), t(4; ?)(q31;?); der(5),t(5;?)(q31;?); der(13),t(13;13)(p11;q11) and others. Bimo dal distribution of chromosomes with more aberrant chromosomes appeared in the 31st and 60th passage cells. Telomere length sharply shortened from nor mal fetal esophagus to the 10th and 31st passage step by step, but was stab le from the 31st to the 60th passage and the telomerase activities measured were expressed at late two passages. p53 mutant was positive in three pass ages, c-myc was positive in the 31st and the 60th passage K-ras only in the last. The results reveal that changes of chromosomes, telomere length, tel omerase activity and certain gene expressions are important events of HPV-i mmortalized esophageal epithelium cells. All of these changes occurred in d ynamic progressive process. This cell line may be useful for the elucidatio n of the genetic mechanism of cellular immortalization.