Tumor suppressor gene alterations of spontaneously malignant transformed cells from human embryonic muscle in vitro

Abstract

Abstract.Recent research has shown that mesenchymal stem cells (MSCs) which were cultured for long time could transform malignantly, the transformation mechanism is not clear yet, it might be associated with the activation of oncogenes and inactivation of tumor suppressor genes. In our initial investigation, we found that the cells arising from human embryonic muscle could spontaneously transform into malignancy in vitro and we obtained 6 immortalized cell lines. In this study, polymerase chain reaction (PCR) was used to assay several tumor suppressor genes of these cell lines, and homozygous deletions within chromosomal band 9p2l including MTAP (methylthioadenosine phosphorylase), p16 and p15 were detected. PCR products of p53 exons 7 and 8 of these novel tumor cell lines were assayed by sequencing, and the results showed high prevalence of mutations in these regions, the mutation rate reached as high as 8% in exon 7 and 14% in exon 8, and all of them were point mutations, the intron 7 changed more significantly, including piece deletion, insertion, frameshift and point mutation, it showed almost no similarity to that of the wt p53 sequence, that was totally different from other p53 mutation data published. All the mutation sequences were identical in 6 cell lines, this suggest that there may be a common mutation mechanism and strong selective advantage in these novel tumor cell lines over long-term culture. In conclusion, our research shows that the inactivation of tumor suppressor genes may play an important role in the process of malignant transformation of embryonic muscle cells in vitro. IntroductionThe development of tumors is generally accepted to be a multistep process in which alterations in oncogenes and tumor suppressor genes play an important role (1,2). The process of carcinogenesis involves the gain of oncogene activity and the loss of tumor suppressor gene function, such as Rb, p53 and cyclin-dependent kinase inhibitor (CDKI) family (3-5). The aberrant gene could interfere in many cell funtions, such as cell proliferation, differentiation and apoptosis. The accumulative mutations constitute the base of cell malignant transformation in gene level. The acquired capabilities of tumor cells include their ability to proliferate continuously ignoring apoptosis or growth-inhibitory signals, generating their own mitogenic signals.At present, the reports on the malignant transformation of human cell in vitro are rare. Recent research has found that mesenchymal stem cells (MSCs) cultured for long time in vitro could lead to malignant transformation, but the mechanism is not very clear. The activation of oncogenes and inactivation of tumor suppressor genes are considered to be one of the mechanisms (6-9). We have found that human embryonic muscle cells which were cultured in vitro longterm could become spontaneously immortalized, and we obtained 6 novel malignant cell lines, this new type of cell lines were named human embryonic muscle-derived malignant transformed cells (hEMTCs). These hEMTCs could be pass...