A Study On Expression And Mutation Of The PTEN Gene In Esophageal Squamous Cell Carcinoma

The PTEN gene (phosphatase and tensin homolog deleted on chromosome ten) has been identified as the first tumor suppressor located at 10q23. The gene has dual-specificity phosphatase with lipid and protein phoshpatase activation. The PTEN has the regulatory effect on a cascade of signaling pathways such as PI-3K/AKT signaling pathway. Its main functions include induction of apoptosis, inhibition of cell proliferation, cell migration and adhesion, and participating in normal embryonic development and maintenance of the immune system stability. Mutation and deletions in the PTEN locus were associated with a number of human cancers, including endometrial cancer, prostate cancer, glioblastoma, melanoma, etc, in which there are very high mutations of the PTEN gene. Current studies have confirmed that the mutations and inactivations of the PTEN gene are closely related with tumor development. Esophageal squamous cell carcinoma (ESCC) is one of the most common malignant tumors in China, and incidence and mortality of ESCC are the highest in Linzhou City of Henan province. The reports of the studies regarding molecular biology of the PTEN gene in ESCC are still rare. In this study, we investigated the expressions and mutations of the PTEN gene in three cell lines of ESCC, EC9706, ECa109 and EC1, respectively. Besides, the relationship between the PTEN gene and carcinogenesis of ESCC was discussed. The findings of this study will help to explore the roles of the PTEN gene in carcinogenesis of the esophagus and to provide new methods for gene diagnosis and therapy for ESCC.Total RNAs were extracted from EC9706, ECa109 and EC1 cells. The mRNAs of total RNAs were transcripted reversely into cDNA, respectively. The first strand cDNA was used as a template, and PTEN was amplified by polymerase chain reaction with a set of synthetic oligonucleotide primers. The amplified DNA fragment was introduced into the pMD18-T vector resulting in a new plasmid pMD18-T-PTEN, which was then transformed to E.coli JM109 cells. Subsequently, the positive colonies were picked out for sequencing, and the results of sequencing were blasted with the wild type PTEN sequence on GenBank. Furthermore, the mRNA of PTEN was investigated using RT-PCR in the three cell lines.The data was statistically analyzed by one-way analysis of variance using SPSS version 13.0. The results of statistics were expressed as the means±the standard deviations. A P value of<0.05 was considered statistically significant.1. mRNA of PTEN was expressed in all the three cell lines, and the expression of mRNA of PTEN was much higher in well differentiated ECa109 cells than that in poor differentiated EC9706 and EC1 cells (P<0.01).2. There were mutations of the PTEN gene in exons 2,5,6,8 and 9 of EC9706 cells, exons 5,8 and 9 of ECa109 cells and exons 6,8 and 9 of EC 1 cells. The mutations mentioned above were missense or nonsense mutations. The mutations of the PTEN gene of three cell lines lead the changes of amino acid sequences. 1. The expression of PTEN in ESCC cells is related with the degree of tumor differentiation. The expression of mRNA of PTEN in well differentiated ESCC cells is significantly higher than that in poorly differentiated ESCC cells.2. There exist mutations of the PTEN gene in ESCC and the hot mutation points are located at exon 5,8 which lead to the changes of amino acid sequences, suggesting that mutations of the PTEN gene may affect the structure and functions of PTEN protein.