Project description:We proposed that the frequency of EBV reactivation may be crucial for its pathogenic role and highly recurrent EBV reactivations exert a profound influence on genome instability. Recurrent reactivations were induced in the EBV-latently infected NPC cells (NA cells) by treating with 12-o-tetradecanoylphorbol-13-acetate (TPA) and sodium n-butyrate (SB) once per passage periodically. Genome-wild oligoarray-based comparative genomic hybridization (CGH) technology was applied to investigate the copy-number aberrations in response for different frequencies of EBV reactivation. The results showed that the NR15 cells, NA cells with the highest frequency (15 times) of reactivation, exhibit extensive genomic copy-number alterations (CNAs) mostly involving chromosome 3p, 3q, 8p, 8q, 9p and 9q, whereas limited number of CNAs were observed both in NR1 cells where only the initial reactivation take place and NP15 cells which were culture in parallel with NR15 cells without EBV-reactivation. We concluded that it is the highly recurrent reactivations of EBV, but neither just a primary reactivation nor EBV-latent infection, may intimately involve in carcinogenesis of nasopharyngeal epithelial cells with the progressive genome instabilities and the accumulation of genetic mutations. Keywords: array-based comparative genomic hybridization, reactivation of EBV, incidence, frequency, recurrent, genome instability, copy-number alterations, carcinogenesis, and nasopharyngeal carcinoma (NPC).
Project description:We proposed that the frequency of EBV reactivation may be crucial for its pathogenic role and highly recurrent EBV reactivations exert a profound influence on genome instability. Recurrent reactivations were induced in the EBV-latently infected NPC cells (NA cells) by treating with 12-o-tetradecanoylphorbol-13-acetate (TPA) and sodium n-butyrate (SB) once per passage periodically. Genome-wild oligoarray-based comparative genomic hybridization (CGH) technology was applied to investigate the copy-number aberrations in response for different frequencies of EBV reactivation. The results showed that the NR15 cells, NA cells with the highest frequency (15 times) of reactivation, exhibit extensive genomic copy-number alterations (CNAs) mostly involving chromosome 3p, 3q, 8p, 8q, 9p and 9q, whereas limited number of CNAs were observed both in NR1 cells where only the initial reactivation take place and NP15 cells which were culture in parallel with NR15 cells without EBV-reactivation. We concluded that it is the highly recurrent reactivations of EBV, but neither just a primary reactivation nor EBV-latent infection, may intimately involve in carcinogenesis of nasopharyngeal epithelial cells with the progressive genome instabilities and the accumulation of genetic mutations. Experiment Overall Design: Samples of four sources were analyzed. The NP1 cells denoted the EBV-latently infected nasopharyngeal carcinoma cell lines, NA cells, with low-passage-number and mock-treated. The NR1 denoted the NA cells harboring one time of EBV reactivation. The NR15 cells denoted the NA cells harboring fifteen times of EBV reactivation. The NP15 cells denoted the NA cells cultured in parallel with NR15 (experienced 15 times of passages) without EBV-reactivation. Genomic DNAs extracted from NP15, NR1 and NR15 cells were used as experimental samples and subjected to Cy5-labeling reactions. Genomic DNAs extracted from NP1 cells were used as the common reference samples subjected to Cy3-labeling reactions. All three experiments were verified by dye-swap design where the targets labeling conditions were exchanged, NP1 labeled with Cy5, and NP15, NR1 or NR15 labeled with Cy3.
Project description:Seroepidemiological studies imply a correlation between Epstein-Barr virus (EBV) reactivation and the development of nasopharyngeal carcinoma (NPC). Phorbol esters, butyrates and N-nitroso compounds are known chemical carcinogens in foodstuffs and cigarettes that have been implicated as risk factors contributing to the development of NPC. We have demonstrated previously that low dose N-methyl-N'-nitro-N-nitrosoguanidine (MNNG, 0.1 microg/ml) had a synergistic effect with 12-O-tetradecanoylphorbol-13-acetate (TPA) and sodium butyrate (SB) in enhancing EBV reactivation (Chem Biol Interact 188: 623-634). Since residents of areas with a high risk of NPC are reported to contact with these carcinogens (TPA, SB or nitrosamines) frequently, we sought to determine the consequence of repeated exposure of EBV-harboring nasopharyngeal cells to these carcinogens in a long-term, low dose, repeated manner. An NPC cell line latently infected with EBV, NA, was periodically treated with TPA/SB combined with MNNG for recurrent EBV reactivation. After 10 times of chemically-induced recurrent reactivation of EBV, the expression profile analysis indicates that many carcinogenesis-related genes were altered in recurrent reactivated NA cells when compared to the parental NA cells. The expression profile was analyzed in the recurrent EBV reactivated NA cells and the parental NA cells.
Project description:Recurrent karyotypic abnormalities are a characteristic feature of cervical cancer (CC) cells, which may result in deregulated expression of important genes that contribute to tumor initiation and progression. To examine the role of genomic copy number alterations, we surveyed genetic lesions in CC utilizing single nucleotide polymorphism (SNP) array. We identified specific genetic alterations associated with CC. These data will be useful in identification of target altered genes, novel markers for predicting high risk precancerous lesions to invasive cancer, comparison of copy number alterations with gene expression changes can provide gene targets for pharmacologic intervention. We demonstrate specific regions of gene amplification (e.g., 11q22), copy number gains (e.g., 3q, 5p, and 20q), and deletions (e.g., 2q, 11q23) in the present study, which forma a framework for identification of critical genes in CC tumorigenesis. Keywords: Cervical cancer, copy number alterations, HPV type, gene amplification