Biomedical subjects

Michiko Nishigaki

Publications and source records attributed to Michiko Nishigaki.

6 recordsLinked to original sources

PubMed2005

Discovery of aberrant expression of R-RAS by cancer-linked DNA hypomethylation in gastric cancer using microarrays.

Although hypomethylation was the originally identified epigenetic change in cancer, it was overlooked for many years in preference to hypermethylation. Recently, gene activation by cancer-linked hypomethylation has been rediscovered. However, in gastric cancer, genome-wide screening of the activated genes has not been found. By using microarrays, we identified 1,383 gene candidates reactivated in at least one cell line of eight gastric cancer cell lines after treatment with 5-aza-2'deoxycytidine and trichostatin A. Of the 1,383 genes, 159 genes, including oncogenes ELK1, FRAT2, R-RAS, RHOB, and RHO6, were further selected as gene candidates that are silenced by DNA methylation in normal stomach mucosa but are activated by DNA demethylation in a subset of gastric cancers. Next, we showed that demethylation of specific CpG sites within the first intron of R-RAS causes activation in more than half of gastric cancers. Introduction of siRNA into R-RAS-expressing cells resulted in the disappearance of the adhered cells, suggesting that functional blocking of the R-RAS-signaling pathway has great potential for gastric cancer therapy. Our extensive gene list provides other candidates for this class of oncogene.

Azacitidine↗

PubMed2004

Alternative mechanisms of gene amplification in human cancers.

Gene amplification is a common phenomenon in cancer. Cytogenetic analyses have indicated that breakage-fusion-bridge (BFB) cycles drive intrachromosomal amplification of some oncogenes in a head-to-head manner in human cancers. However, the complex structures of an amplified sequence found in cancers are not always explained by the BFB model. At the 17q21 locus, which is not linked to common fragile sites, we discovered a recombination hot spot harboring amplicon repeats in tandem in a head-to-tail orientation, with the interamplicon junctions in each cancer cell being homogeneous. These findings clearly show the presence of alternative mechanisms other than BFB cycles in oncogene amplification.

Base Sequence↗

PubMed2004

Using a customized DNA microarray for expression profiling of the estrogen-responsive genes to evaluate estrogen activity among natural estrogens and industrial chemicals.

We developed a DNA microarray to evaluate the estrogen activity of natural estrogens and industrial chemicals. Using MCF-7 cells, we conducted a comprehensive analysis of estrogen-responsive genes among approximately 20,000 human genes. On the basis of reproducible and reliable responses of the genes to estrogen, we selected 172 genes to be used for developing a customized DNA microarray. Using this DNA microarray, we examined estrogen activity among natural estrogens (17beta-estradiol, estriol, estrone, genistein), industrial chemicals (diethylstilbestrol, bisphenol A, nonylphenol, methoxychlor), and dioxin. We obtained results identical to those for other bioassays that are used for detecting estrogen activity. On the basis of statistical correlations analysis, these bioassays have shown more sensitivity for dioxin and methoxychlor.

Biological Assay↗

PubMed2003

A faithful method for PCR-mediated global mRNA amplification and its integration into microarray analysis on laser-captured cells.

Quantitative and qualitative analyses of mRNAs from a small number of cells are extremely important for studies on gene expression in various physiological and pathological conditions in multicellular organisms. We present here an effective method for high-fidelity global mRNA amplification for in vivo gene expression profiling of as few as 100 cells obtained by laser-captured microdissection (LCM). This method, called TALPAT, is based on T7 RNA polymerase-mediated transcription, adaptor ligation, and PCR amplification followed by T7-transcription. More than 80% of genes were commonly identified as a more than 3-fold changed gene among three gastric cancer cell lines using cRNA amplified by both TALPAT and the ordinary in vitro T7-transcription. The reproducibility of TALPAT was validated by microarray analysis on 100 breast cancer cells obtained by LCM. For the application of the LCM-TALPAT method, we successfully obtained expression profiles of gastric cancer cells and the mesenchymal cells, enabling us to understand in vivo cell-to-cell cross-talk in the microenvironment.

Breast Neoplasms↗

PubMed2002

Identification of the CAB2/hCOS16 gene required for the repair of DNA double-strand breaks on a core amplified region of the 17q12 locus in breast and gastric cancers.

We previously reported that CAB1 and c-ERBB-2 genes were found to be located in a core amplified region of the 17q12 locus, which is frequently amplified in various cancers. During identification of this core region, CAB2, a human homologue of the yeast COS16 required for the repair of DNA double-strand breaks was cloned. Autofluorescence analysis of cells transfected with its GFP fusion protein demonstrated that CAB2 translocates into vesicles, suggesting that overexpression of CAB2 may decrease intercellular Mn2+ by accumulating it in the vesicles, in the same way as yeast COS16. This is the first report identifying all of the genes on the core amplified region of the 17q12 locus in breast and gastric cancers.

Amino Acid Sequence↗

PubMedDate not supplied

A reporter gene assay for evaluation of tissue-specific responses to estrogens based on the differential use of promoters A to F of the human estrogen receptor alpha gene.

INTRODUCTION: Reporter gene assays are useful means for monitoring cellular responses. We report here a reporter gene assay for evaluating and monitoring estrogen activities by estrogen-like compounds and xenoestrogens, which is based on the promoters from the human estrogen receptor alpha (ERalpha) gene. METHODS: The reporter gene constructs contained a proximal promoter region (containing promoters A and B: ProAB) or either of promoters C to F (ProC, ProD, ProE, and ProF) or fused minor promoters (ProCDEF). These constructs were first used to evaluate promoter activity in cell lines derived from breast (MCF-7 and T-47D cells), ovary (SK-OV-3 cells), endometrium (Ishikawa cells), and stomach (MKN-28 cells). RESULTS: Besides very high levels of activity by ProAB in all of the cell lines tested, moderate levels were detected for ProD in the breast and endometrium cell lines and for ProF in the ovary and endometrium cell lines. A moderate level of activity by ProE was detected only in the stomach cells. Differences in estrogen-like activity between ProAB and ProD were observed for tamoxifen and bisphenol A (BPA) in MCF-7 cells. DISCUSSION: The assay proposed here might provide expression profiles of cancer cells of various origins for evaluating the estrogen responsiveness and for identifying tissue- or cancer cell-specific transcription factors.

Benzhydryl Compounds↗