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Michiie Sakamoto

Publications and source records attributed to Michiie Sakamoto.

At least 55 records · Page 3Linked to original sources
PubMed2004

Prognostic significance of dysadherin expression in esophageal squamous cell carcinoma.

OBJECTIVE: Dysadherin is a cancer-associated cell membrane glycoprotein that has been reported to downregulate E-cadherin expression and promote metastasis. To evaluate the role of dysadherin in metastasis of esophageal squamous cell carcinoma (ESCC), we examined dysadherin and E-cadherin expression in patients with this cancer. METHODS: Dysadherin and E-cadherin expression was evaluated in 117 ESCC patients (pT1, 31; pT2, 30; pT3, 39; pT4, 17) by immunohistochemistry. The findings were compared with the clinicopathological data of the patients. RESULTS: Both dysadherin and E-cadherin were localized to the cell membrane. Thirty patients (29.1%) had tumors positive for dysadherin and 41 patients (35.0%) had tumors positive for E-cadherin. Tumors showing dysadherin positivity and negative E-cadherin expression had a significantly worse prognosis than other tumors. When the patients with dysadherin- positive tumors were combined with E-cadherin-negative patients, this group had a worse prognosis (p < 0.0001). Cox multivariate analysis revealed that dysadherin expression was an independent prognostic factor for ESCC (p = 0.003), but E-cadherin expression was not. CONCLUSION: Combined analysis of dysadherin and E-cadherin expression may help to predict the prognosis of patients with ESCC. Our results suggested that expression of dysadherin by this cancer may partly explain the poor prognosis of patients with preservation of E-cadherin expression.

Aged↗
PubMed2004

Expression profiling in hepatocellular carcinoma with intrahepatic metastasis: identification of high-mobility group I(Y) protein as a molecular marker of hepatocellular carcinoma metastasis.

Hepatocellular carcinoma (HCC) is one of the most common and aggressive human malignancies. Its high mortality rate is mainly a result of intra-hepatic metastases. To investigate the detailed genetic mechanisms in cancer metastasis, we compared the expression profiles of 20 HCCs with intrahepatic metastasis and 10 HCCs without intrahepatic metastasis using an oligonucleotide array. Of the approximately 12,600 genes that were analyzed, we identified 34 genes whose expression levels were significantly correlated with intrahepatic metastasis (P<0:05). Of these genes, we further investigated the expression of high-mobility group I(Y) [HMG-I(Y)] protein. Real-time quantitative reverse transcription polymerase chain reaction (RT-PCR) confirmed that HMG-I(Y) was upregulated in HCC with intrahepatic metastasis, compared to its level in HCC without intrahepatic metastasis. Further immunohistochemical examination of HMG-I(Y) revealed a significant overexpression in HCC with intrahepatic metastasis, compared with that in HCC without intrahepatic metastasis (P<0:05). These results indicate that the molecular signatures of HCC with intrahepatic metastasis and of HCC without intrahepatic metastasis are clearly different. HMG-I(Y) expression was associated with intrahepatic metastasis and may be a predictive marker of HCC intrahepatic metastasis.

Algorithms↗
PubMed2003

Promoter methylation of the TSLC1 gene in advanced lung tumors and various cancer cell lines.

We previously identified TSLC1, a tumor suppressor gene in human nonsmall cell lung cancer (NSCLC). TSLC1 belongs to immunoglobulin superfamily molecules and is involved in cell adhesion. Loss of TSLC1 expression was strongly correlated with the promoter hypermethylation in several NSCLC cell lines. Here, we examined the methylation status of the TSLC1 gene promoter in 48 primary NSCLC tumors by bisulfite SSCP in combination with bisulfite sequencing. Six CpG sites around the promoter regions were significantly methylated in 21 of 48 primary NSCLC tumors (44%). Promoter methylation was more likely to be observed in relatively advanced tumors with TNM classification of pT2, pT3 or pT4 (19 of 33, 58%) than in those with pT1 (2 of 15, 13%), suggesting that alteration of TSLC1 would be involved in the progression of human NSCLC. Loss of TSLC1 expression was also observed in 20 of 46 (43%) human cancer cell lines, including those from esophageal (3 of 3), gastric (8 of 9), ovarian (2 of 5), endometrial (2 of 2), breast (1 of 3), colorectal (2 of 8) and small cell lung cancers (2 of 10). Combined analysis of promoter methylation and the allelic state in these cell lines indicated that the TSLC1 gene was often silenced not only by mono-allelic methylation associated with loss of the other allele but also through bi-allelic methylation. These results suggest that alteration of TSLC1 would be involved in advanced NSCLC as well as in many other human cancers.

Adenocarcinoma↗
PubMed2003

Nemo-like kinase induces apoptosis in DLD-1 human colon cancer cells.

Deregulation of Wnt/beta-catenin signaling is thought to play a critical role in human carcinogenesis. Nemo-like kinase (NLK) is an evolutionarily conserved serine/threonine kinase that suppresses beta-catenin/T-cell factor (TCF) complex transcriptional activity through phosphorylation of TCF. Since NLK may be a tumor suppressor as a negative regulator of Wnt/beta-catenin pathway, we established tetracycline-inducible NLK and its kinase-negative mutant expression in DLD-1 human colon cancer cells to analyze the effect of NLK on cell growth and viability. The induction of wild-type NLK in DLD-1 cells caused suppression of cell growth whereas the kinase-negative mutant did not. Flow cytometry indicated that NLK expression increased the number of apoptotic cells but did not induce obvious cell cycle arrest. Apoptosis induction by wild-type NLK was confirmed using TUNEL assays. Our results suggest that overexpression of NLK may have targets other than TCF for induction of apoptosis in human colon carcinoma cells.

Apoptosis↗
PubMed2003

Proteomic analysis of intestinal epithelial cells expressing stabilized beta-catenin.

Aberrant accumulation of beta-catenin protein because of mutation of either the beta-catenin or adenomatous polyposis coli gene plays an essential role in the development of colorectal carcinoma. We established previously a stable clone of the rat small intestinal epithelial cell line IEC6, which is capable of inducing stabilized beta-catenin protein lacking NH(2)-terminal glycogen synthase kinase-3beta phosphorylation site under a strict control of the tetracycline-regulatory system. This clone, IEC6-TetOFF-beta-catenin DeltaN89, shows in vitro polypoid growth on the removal of doxycycline and seems to be an appropriate model for analyzing the molecular mechanisms of early intestinal carcinogenesis. Of >2000 protein spots displayed by newly developed two-dimensional difference gel electrophoresis, 22 were found to be up- or down-regulated on the induction of stabilized beta-catenin. The majority of these proteins fell into two categories: (a) redox-status regulatory proteins and (b) cytoskeleton-associated proteins. Representatively, a key redox-status regulatory protein, manganese superoxide dismutase, up-regulated in IEC6 cells expressing stabilized beta-catenin protein, was overexpressed in adenoma and adenocarcinoma cells of familial adenomatous polyposis patients in parallel with the accumulation of beta-catenin. These results suggest that aberrant accumulation of beta-catenin might contribute to colorectal carcinogenesis by affecting redox status in the mitochondria of intestinal epithelial cells.

Adenocarcinoma↗
PubMed2003

Increased protein expression of DNA methyltransferase (DNMT) 1 is significantly correlated with the malignant potential and poor prognosis of human hepatocellular carcinomas.

Alteration of DNA methylation is one of the most consistent epigenetic changes in human cancers. DNA methyltransferase (DNMT) 1 is a major enzyme involved in establishing genomic methylation patterns. Most of the studies concerning DNMT1 expression in human cancers have been performed only at the mRNA level. To directly examine DNMT1 protein expression levels during human hepatocarcinogenesis, 16 histologically normal liver tissues, 51 noncancerous liver tissues exhibiting chronic hepatitis or cirrhosis, which are considered to be precancerous conditions, and 53 hepatocellular carcinomas (HCCs) were subjected to immunohistochemic examination. If more than 20% of the cells exhibited nuclear DNMT1 staining, the tissue sample was considered to be DNMT1-positive. DNMT1 immunoreactivity was observed in 23 (43%) of the HCCs, but in none (0%) of the histologically normal liver or noncancerous liver tissues exhibiting chronic hepatitis or cirrhosis. The incidence of increased DNMT1 protein expression in HCCs correlated significantly with poor tumor differentiation (p = 0.0006) and portal vein involvement (p = 0.0002). Moreover, the recurrence-free (p = 0.0001) and overall (p < 0.0001) survival rates of patients with HCCs exhibiting increased DNMT1 protein expression were significantly lower than those of patients with HCCs that did not exhibit increased expression. Increased DNMT1 protein expression may play a critical role in the malignant progression of HCCs and be a biologic predictor of both HCC recurrence and a poor prognosis in HCC patients.

Carcinoma, Hepatocellular↗
PubMed2003

Aberrant O-glycosylation inhibits stable expression of dysadherin, a carcinoma-associated antigen, and facilitates cell-cell adhesion.

Recently, we identified dysadherin, a novel carcinoma-associated glycoprotein, and showed that overexpression of dysadherin in human hepatocarcinoma PLC/PRF/5 cells could suppress E-cadherin-mediated cell-cell adhesion and promote tumor metastasis. The present study shows evidence that dysadherin is actually O-glycosylated. This was based on a direct carbohydrate composition analysis of a chimera protein of an extracellular domain of dysadherin fused to an Fc fragment of immunoglobulin. To assess the importance of O-glycosylation in dysadherin function, dysadherin-transfected hepatocarcinoma cells were cultured in a medium containing benzyl-alpha-GalNAc, a modulator of O-glycosylation. This treatment facilitated homotypic cell adhesion among dysadherin transfectants accompanied with morphological changes, indicating that the anti-adhesive effect of dysadherin was weakened. Modification of O-glycan synthesis also resulted in down-regulation of dysadherin expression and up-regulation of E-cadherin expression in dysadherin transfectants but did not affect E-cadherin expression in mock transfectants. Structural analysis of O-glycans released from the dysadherin chimera proteins indicated that a series of O-glycans with core 1 and 2 structures are attached to dysadherin, and their sialylation is remarkably inhibited by benzyl-alpha-GalNAc treatment. However, sialidase treatment of the cells did not affect calcium-dependent cell aggregation, which excluded the possibility that sialic acid itself is directly involved in cell-cell adhesion. We suggest that aberrant O-glycosylation in carcinoma cells inhibits stable expression of dysadherin and leads to the up-regulation of E-cadherin expression by an unknown mechanism, resulting in increased cell-cell adhesion. The carbohydrate-directed approach to the regulation of dysadherin expression might be a new strategy for cancer therapy.

Acetylgalactosamine↗
PubMed2003

Mutation of the DNA methyltransferase (DNMT) 1 gene in human colorectal cancers.

Alteration of DNA methylation is one of the most consistent epigenetic changes in human cancers. DNA methyltransferase (DNMT) 1 is a major enzyme that determines genomic methylation patterns. In order to understand the significance of mutations of the DNMT1 gene during human carcinogenesis, we performed polymerase chain reaction-single strand conformation polymorphism analysis using 46 oligonucleotide primer sets for all 40 coding exons and the 5'-flanking region (450 bp) of the DNMT1 gene in 29 colorectal cancers, 32 stomach cancers, 40 hepatocellular carcinomas (HCCs) and a corresponding sample of non-cancerous tissue from each case. Mutations in coding exons of the DNMT1 gene were detected in two (7%) of the colorectal cancers: they consisted of one-base deletion resulting in deletion of the whole catalytic domain and a point mutation resulting in a single amino acid substitution. No stomach cancers or HCCs showed mutations in the coding exons of the DNMT1 gene. No mutation in the 5'-flanking region of the DNMT1 gene was detected in any of the colorectal and stomach cancers or HCCs. These data suggest that mutational inactivation of the DNMT1 gene that potentially causes a genome-wide alteration of DNA methylation status may be a rare event during human carcinogenesis.

Aged↗
PubMed2003

Dysadherin overexpression in pancreatic ductal adenocarcinoma reflects tumor aggressiveness: relationship to e-cadherin expression.

PURPOSE: The E-cadherin-mediated cell adhesion system is frequently inactivated by multiple mechanisms and is involved in tumor progression in many types of cancer. Recently, we reported the cloning and characterization of dysadherin and showed that it downregulated E-cadherin and promoted metastasis. The aim of this study was to investigate the clinical significance of dysadherin expression and the relationship between dysadherin expression and E-cadherin expression in pancreatic ductal adenocarcinoma. PATIENTS AND METHODS: We examined dysadherin and E-cadherin expression in 125 surgically resected pancreatic ductal adenocarcinoma patients using immunohistochemistry. RESULTS: Dysadherin was expressed at the cell membrane of cancer cells, but not in nontumor duct and acinar cells. Its expression was stronger in infiltrative and poorly differentiated nests compared with well-differentiated nests. Although the correlation between the expression of dysadherin and E-cadherin was not significant, a group of patients showed reduced E-cadherin expression with dysadherin overexpression. Increased dysadherin expression was significantly correlated with distant metastasis (P =.047), high tumor grade (P =.006), positive tumor margins (P =.024), and infiltrative type of growth pattern (P =.014). A survival advantage was observed in patients with 0% to 20% dysadherin-positive cells compared with patients with 51% to 100% dysadherin-positive cells, independent of tumor-node-metastasis classification, and World Health Organization tumor grade (P =.019). A combination of increased dysadherin expression and reduced E-cadherin expression (< 90%) further worsened the prognosis. CONCLUSION: In pancreatic ductal adenocarcinoma, dysadherin expression seems to reflect tumor aggressiveness and to be a positive marker of poor prognosis when considered both alone and in combination with downregulation of E-cadherin.

Aged↗
PubMed2003

Overexpression of KIT in chromophobe renal cell carcinoma.

We analysed gene-expression profiles in 15 surgical specimens of conventional, papillary, and chromophobe renal cell carcinomas (RCCs) using high-density oligonucleotide arrays. From about 12,000 genes targeted by the array, 67 were upregulated specifically in each histological type of RCC. The oncogene KIT was one of the genes whose expression was upregulated specifically in chromophobe RCCs. Immunohistochemical analysis demonstrated the KIT gene product on the cell membrane of chromophobe RCC in all cases, although it was not detected in conventional RCCs or non-neoplastic kidneys except for weak staining in the cytoplasm of renal tubules. These results suggest that each histological subtype of RCC has a unique gene-expression profile, and in particular indicates for the first time that KIT could be a useful marker for chromophobe RCC. As overexpression of KIT might be involved in tumor growth, KIT could be a new therapeutic target in this special type of RCC.

Biomarkers, Tumor↗
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↗
PubMed2003

Evaluation of a whole-genome amplification method based on adaptor-ligation PCR of randomly sheared genomic DNA.

High-throughput genetic studies often require large quantities of DNA for a variety of analyses. Developing and assessing a whole-genome amplification method is thus important, especially with the current desire for large-scale genotyping in previously collected samples for which limited DNA is available. The method we have developed, called PRSG, is based on an adaptor-ligation-mediated PCR of randomly sheared genomic DNA. An unbiased representation was evaluated by performing PCR on 2,607 exons of 367 genes, which are randomly distributed throughout the genome, on PRSG products of hundreds of individuals. An infrequent loss (<1%) of the exon sequence on the PRSG products was found. Out of 307 microsatellites on various chromosomes, 258 (84%) were amplified in both the PRSG product and an original DNA, whereas 49 (16%) microsatellites were lost only in the PRSG product. Array CGH analysis of 287 loci for measuring the relative gene copy number demonstrated that a low bias was detected. Moreover, this method was validated on 100-1,000 laser-captured cells from paraffin-embedded tissues. These data show that PRSG can provide a sufficient amount of genomic sequence for a variety of genetic analyses as well as for long-term storage for future work.

DNA Ligases↗
PubMed2003

Beta-catenin mutations in pulmonary blastomas: association with morule formation.

To elucidate the contribution of beta-catenin gene mutation to the development of pulmonary blastomas, we analysed mutations in three well-differentiated fetal adenocarcinomas (WDFAs) and six biphasic pulmonary blastomas (BPBs). For comparison, eight clear-cell adenocarcinomas with fetal lung features were also examined. beta-Catenin gene mutations were found in all three WDFAs, two BPBs, and none of the clear-cell adenocarcinomas with fetal lung features. All tumours with mutations had a common histological feature, namely morule formation, and showed a characteristic heterogeneous beta-catenin expression pattern that was revealed by immunohistochemistry. Strong nuclear/cytoplasmic expression of beta-catenin was seen in clustered cells in the morular areas and in single cells in glands, and was associated with neuroendocrine differentiation. As beta-catenin mutations are rare among lung tumours, this distinctive genetic feature, which is also immunohistochemically detectable as overexpression with a heterogeneous pattern, has diagnostic significance. The presence of this common genetic alteration found in both WDFA and BPB implies a histogenetic linkage between these tumours.

Adenocarcinoma↗
PubMed2003

Alpha-fetoprotein-producing carcinoma of the colon: report of a case and review of the literature.

PURPOSE: We describe a rare case of an alpha-fetoprotein-producing carcinoma originating in the transverse colon of a 59-year-old Japanese male. METHODS: The patient reported an abdominal mass and weight loss. On examination, a tumor of the transverse colon and multiple masses in the liver were found. The serum alpha-fetoprotein level was extremely high (12,873 ng/ml). The patient underwent right hemicolectomy and intraoperative biopsy of a liver mass. RESULTS: Histologically, the colon cancer was composed of three different components: a well-differentiated tubular adenocarcinoma, a tubulopapillary carcinoma consisting of cells with clear cytoplasms, and a "hepatoid carcinoma." The hepatoid carcinoma was composed of large polygonal cells with abundant eosinophilic or clear cytoplasms, arranged in a trabecular or solid pattern, and showing marked vascular invasion. Immunohistochemically, alpha-fetoprotein was strongly expressed, largely in the hepatoid carcinoma and partially in the tubulopapillary carcinoma. The liver biopsy specimen showed morphologic and immunohistochemical features similar to those of the hepatoid carcinoma of the colon and was therefore diagnosed as a metastasis. The patient died of the cancer two months after surgery. CONCLUSION: Based on our experience of this patient and a review of the literature, alpha-fetoprotein-producing colorectal carcinomas are generally associated with a poor prognosis because of the frequent occurrence of blood-borne metastases.

Adenocarcinoma↗
PubMed2003

Clear cell endocrine tumor of the pancreas which is not associated with von Hippel-Lindau disease: report of a case.

We describe the case of a pancreatic endocrine tumor with marked clear cell changes. The patient, a 56-year-old woman, presented with appetite loss and nausea but no evidence of either von Hippel-Lindau disease, or a family history of the disease. A radiological examination revealed a hypervascular tumor measuring 5.2 cm in diameter located in the pancreatic head. The patient underwent a pylorus-preserving pancreatoduodenectomy. The cut surface of the tumor was yellowish and well demarcated. Histologically, the tumor was composed of cuboidal cells with clear cytoplasm, thus forming trabecular and insular nests. Immunohistochemically, the tumor cells were strongly positive for chromogranin. An electron microscopic examination revealed the tumor cells to contain neuroendocrine secretory granules, abundant lipid droplets, and some crystalloid bodies in the cytoplasm. We conclusively diagnosed it to be a pancreatic clear cell endocrine tumor. Such tumors not associated with von Hippel-Lindau disease are extremely rare.

Carcinoma, Islet Cell↗
PubMed2003

Expression profiling in ovarian clear cell carcinoma: identification of hepatocyte nuclear factor-1 beta as a molecular marker and a possible molecular target for therapy of ovarian clear cell carcinoma.

Of all of the epithelial ovarian cancers, clear cell carcinoma (CCC) of the ovary has the worst prognosis. We applied the oligonucleotide array technique to identify genes generally involved in CCC. Of the approximately 12,600 genes that were analyzed, 28 were expressed significantly differently between four CCC and seven non-CCC cell lines. Among 16 up-regulated genes in CCC, we further investigated a transcription factor, hepatocyte nuclear factor-1 beta (HNF-1 beta). We validated up-regulation of HNF-1 beta in CCC in terms of both mRNA and protein level using real-time quantitative reverse transcriptase-polymerase chain reaction and immunoblotting. Immunohistochemical analysis of 83 surgically resected ovarian cancers showed that almost all CCC specimens (21 of 22 cases) had nuclear staining for HNF-1 beta, whereas most non-CCC specimens (60 of 61 cases) showed no immunostaining or only focal and faint staining in the nucleus. Furthermore, we investigated the significance of HNF-1 beta expression in CCC using RNA interference. The reduction of HNF-1 beta expression by RNA interference induced apoptotic cell death in ovarian CCC cells, which was confirmed by terminal dUTP nick-end labeling and fluorescence-activated cell-sorting analyses. Our results suggest that HNF-1 beta is not only an excellent CCC-specific molecular marker but also a molecular target for therapy of ovarian CCC.

Adenocarcinoma, Clear Cell↗
PubMed2003

Possible involvement of tetraspanin CO-029 in hematogenous intrahepatic metastasis of liver cancer cells.

BACKGROUND AND AIM: A correlation between overexpression of tetraspanin CO-029 and the intrahepatic spread of hepatocellular carcinoma (HCC) has been observed in surgically resected specimens from humans. However, the cellular mechanisms involved in CO-029 protein modulation of the metastatic phenotype are unknown. In the present study, CO-029 cDNA was stably transfected into a non-metastatic human HCC cell line to investigate whether it could directly promote metastasis. METHODS: We constructed a human HCC cell line that stably overexpressed CO-029 and mock transfectants. Using these transfectants, we examined cell proliferation characteristics in monolayer culture and the ability to adhere to culture plates coated with laminin, fibronectin, vitronectin and collagen type-I or type-IV. Orthotopic implantation of these transfectants to SCID mice was also performed. RESULTS: Several clones of CO-029 transfectants and mock transfectants were established. The growth rates and adhesive properties to the extracellular matrix did not differ between CO-029 and mock transfectants. When orthotopically implanted, the size of the primary tumor in the liver did not differ between CO-029 and the mock transfectants. However, only CO-029 positive clones developed intrahepatic metastatic lesions. CONCLUSION: These results suggest that CO-029 might be involved in hematogenous intrahepatic metastasis, although the precise cellular mechanisms involved remain unknown.

Animals↗