A solid tumor of donor cell-origin after allogeneic peripheral blood stem cell transplantation.
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Adenocarcinoma↗
Biomedical subjects
H Kawamata
Publications and source records attributed to H Kawamata.
Frequent downregulation of 14-3-3 sigma protein and hypermethylation of 14-3-3 sigma gene in salivary gland adenoid cystic carcinoma.
14-3-3 sigma:, a target gene of the p53 tumour suppressor protein, has been shown to regulate the cell cycle at the G2/M checkpoint. Recent studies have demonstrated that 14-3-3 sigma is downregulated by hypermethylation of the CpG island in several types of cancer. In this study, we investigated the expression and methylation status of 14-3-3 sigma in human salivary gland adenoid cystic carcinoma (ACC) and mucoepidermoid carcinoma (MEC). Immunohistochemical analysis revealed that the positive expression rate of 14-3-3 sigma in ACC (one out of 14) was markedly lower than that in MEC (ten out of 10). Since most of the ACCs carried the wild-type p53 protein, downregulation of 14-3-3 sigma in ACC may not be due to the dysfunction of p53 pathway. Microdissection-methylation-specific PCR revealed that frequent hypermethylation of the 14-3-3 sigma gene was observed in ACC when compared to that in MEC. In cultured-ACC cells, we confirmed the downregulation of 14-3-3 sigma via hemimethylation of the gene by sequencing analysis after sodium bisulphite treatment. Furthermore, re-expression of 14-3-3 sigma in the ACC cells was induced by the treatment with DNA demethylating agent, 5-aza-2'-deoxycytidine. Irradiation apparently induced the enhanced expression of 14-3-3 sigma and G2/M arrest in normal salivary gland cells; however, in the ACC cells, neither induction of 14-3-3 sigma nor G2/M arrest was induced by irradiation. These results suggest that downregulation of 14-3-3 sigma might play critical roles in the neoplastic development and radiosensitivity of ACC.
14-3-3 Proteins↗
Development of colonic neoplasia in p53 deficient mice with experimental colitis induced by dextran sulphate sodium.
BACKGROUND: Several animal models for human ulcerative colitis (UC) associated neoplasia have been reported. However, most neoplasias developed in these models have morphological and genetic characteristics different from UC associated neoplasia. AIMS: To establish a new colitis associated neoplasia model in p53 deficient mice by treatment with dextran sulphate sodium (DSS). METHODS: DSS colitis was induced in homozygous p53 deficient mice (p53(-/-)-DSS), heterozygous p53 deficient mice (p53(+/-)-DSS) and wild-type mice (p53+/+-DSS) by treatment with 4% DSS. Numbers of developed neoplasias were compared among the experimental groups, and macroscopic and microscopic features of the neoplasias were analysed. Furthermore, K-ras mutation and beta-catenin expression were assessed. RESULTS: p53(-/-)-DSS mice showed 100% incidence of neoplasias whereas the incidences in p53(+/-)-DSS and p53+/+-DSS mice were 46.2% and 13.3%, respectively. No neoplasias were observed in the control groups. The mean numbers of total neoplasias per mouse were 5.0 (p53(-/-)-DSS), 0.62 (p53(+/-)-DSS), and 0.2 (p53+/+-DSS). The number of neoplasias per mouse in the p53(-/-)-DSS group was significantly higher than that in the other DSS groups. The incidences of superficial type neoplasias were 91.7% in p53(-/-)-DSS mice, 75.0% in p53(+/-)-DSS mice, and 33.3% in p53+/+-DSS mice. The K-ras mutation was not detected in any of the neoplasias tested. Translocation of beta-catenin from the cell membrane to the cytoplasm or nucleus was observed in 19 of 23 (82.6%) neoplasias. CONCLUSIONS: The p53(-/-)-DSS mice is an excellent animal model of UC associated neoplasia because the morphological features and molecular genetics are similar to those of UC associated neoplasia. Therefore, this model will contribute to the analysis of tumorigenesis related to human UC associated neoplasia and the development of chemopreventive agents.
Animals↗
TSC-22 (TGF-beta stimulated clone-22): a novel molecular target for differentiation-inducing therapy in salivary gland cancer.
TSC-22 (Transforming growth factor-beta stimulated clone-22) was originally isolated as a TGF-beta-inducible gene in mouse osteoblastic cells. TSC-22 encodes a putative transcriptional regulator containing a leucine zipper-like structure. Several differentiation-inducing stimuli up-regulate the TSC-22 gene. Furthermore, TSC-22 acts as an effector that integrates multiple extracellular signals during embryogenesis of Drosophila and mouse. Separately, we identified TSC-22 cDNA as an anti-cancer drug (vesnarinone)-inducible gene in a human salivary gland cancer cell line, TYS. Vesnarinone is known to have a differentiation-inducing activity in several cell types. We showed that TSC-22 negatively regulated the growth of TYS cells, and that down-regulation of TSC-22 played a major role in the salivary gland tumorigenesis. Subsequently, we found that artificial overexpression of TSC-22 enhanced chemosensitivity and radiation-sensitivity by inducing apoptosis in TYS cells. Recently, we isolated TSC-22 genomic DNA and analyzed the transcriptional and post-transcriptional regulation of the TSC-22 gene. Then, we confirmed by the luciferase reporter assay that several differentiation-inducing stimuli directly activated the promoter region of TSC-22 gene. Now we are investigating the chemical compounds, which could enhance the transcription of the TSC-22 gene. Thus, because TSC-22 is a key molecule for differentiation of several cells, it can be used as a molecular target for cancer differentiation therapy in salivary gland cancer.
Amino Acid Sequence↗
A MEK inhibitor (U0126) markedly inhibits direct liver invasion of orthotopically inoculated human gallbladder cancer cells in nude mice.
Primary cancer of the gallbladder is not unusual. Most cases of gallbladder cancer are found at an advanced stage, accompanied by the invasion to the liver, metastases to the lymph nodes and distant organs, and peritoneal dissemination. In this study, we first examined the effect of mitogen-activated protein kinase kinase (MEK) inhibitors on the production of matrix metalloproteinases (MMPs), urokinase-type plasminogen activator (uPA), and tissue inhibitors of metalloproteinases (TIMPs) in a human gallbladder cancer cell line, NOZ cells in vitro. MEK inhibitors (PD98059 and U0126) inhibited the production of MMP-2, MMP-9 and high MW uPA, and upregulated TIMPs (TIMP-1, TIMP-2 and TIMP-3). Subsequently, we examined the effect of U0126 on invasion and metastasis of orthotopically inoculated NOZ cells in nude mice. Direct liver invasion by cancer cells was detected in all of the mice in the control group, but in only one mouse in the U0126-treated group. Most of the primary tumors in the U0126-treated group expanded to the liver, but did not invade into the liver. Vessel invasion in the liver was evident in 4 out of 5 mice in the control group, but in only one mouse in the U0126-treated group. Lymph node metastases and peritoneal dissemination were recognized in all of the mice in both groups. All 5 mice in the U0126-treated group, and 4 out of 5 mice in the vehicle control group, had metastases in the lungs. The present results suggest that a MEK inhibitor, U0126, prolonged the survival of the mice with NOZ tumor by inhibiting direct liver invasion and vessel invasion of the cancer cells via down-regulation of the matrix degrading ability of the cancer cells.
Animals↗
Transcriptional activation of cyclin-dependent kinase inhibitor, p21waf1 gene by treatment with a differentiation inducing agent, vesnarinone in a human salivary gland cancer cell line.
Recently, a new concept for cancer therapy termed "tumor dormancy therapy" has been proposed. The concept of this therapy is to prolong the survival time of cancer patients while maintaining their quality of life. We have been developing a differentiation-inducing therapy, which is included in the tumor dormancy therapy, for salivary gland cancer. In this study, we examined the effect of a differentiation-inducing drug, Vesnarinone on the growth of several cancer cells, and examined the molecular mechanism by which Vesnarinone induces the cyclin dependent kinase inhibitor, p21waf1 in the cancer cells. Vesnarinone significantly suppressed the growth of TYS (salivary gland cancer cells), PC3 (prostate cancer cells), and A431 (squamous cell cancer cells). Furthermore, Vesnarinone dose-dependently enhanced the expression of p21waf1 mRNA in TYS cells. Using the luciferase reporter assay it was found that the enhancement of p21waf1 mRNA expression by Vesnarinone was through direct transcriptional activation of the p21waf1 promoter. Thus, analyzing the molecular mechanisms of differentiation inducing drugs may lead to the development of a new therapeutic strategy for several human malignancies, including salivary gland cancer.
Antineoplastic Agents↗
Dedifferentiation and decreased expression of adhesion molecules, E-cadherin and ZO-1, in colorectal cancer are closely related to liver metastasis.
Carcinoma cells with high metastatic potential often show a dedifferentiated phenotype at the primary site. In this study, a total of 48 cases (24 primary tumors of colorectal cancer (Pr-CRC) with liver metastasis, 24 without) were examined for E-cadherin and ZO-1 expression by immunohistochemical staining, and for their dedifferentiated phenotype. The expression levels of E-cadherin and ZO-1 were markedly decreased in the cancer cells of tumors with liver metastasis. Moreover, dedifferentiation of cancer cells, which was evaluated by the modified Gleason score, was also related to liver metastasis. However, none of the conventional clinicopathologic parameters of invasion, except lymph node metastasis, showed any relationship with liver metastasis. These results indicate that dedifferentiation and a decreased expression of E-cadherin and ZO-1 are closely related to liver metastasis.
Cadherins↗
Vesnarinone, a differentiation inducing drug, directly activates p21(waf1) gene promoter via Sp1 sites in a human salivary gland cancer cell line.
We previously demonstrated that a differentiation inducing drug, vesnarinone induced the growth arrest and p21(waf1) gene expression in a human salivary gland cancer cell line, TYS. In the present study, we investigated the mechanism of the induction of p21(waf1) gene by vesnarinone in TYS cells. We constructed several reporter plasmids containing the p21(waf1) promoter, and attempted to identify vesnarinone-responsive elements in the p21(waf1) promoter. By the luciferase reporter assay, we identified the minimal vesnarinone-responsive element in the p21(waf1) promoter at -124 to -61 relative to the transcription start site. Moreover, we demonstrated by electrophoretic mobility shift assay that Sp1 and Sp3 transcription factors bound to the vesnarinone-responsive element. Furthermore, we found that vesnarinone induced the histone hyperacetylation in TYS cells. These results suggest that vesnarinone directly activates p21(waf1) promoter via the activation of Sp1 and Sp3 transcription factors and the histone hyperacetylation in TYS cells.
Antineoplastic Agents↗
Down-regulated p16 expression predicts poor prognosis in patients with extrahepatic biliary tract carcinomas.
The prognosis of extrahepatic biliary tract cancer (EBT) patients is generally accepted to be poor. We immunohistochemically evaluated expression of p16, a cyclin-depend kinase inhibitor, in tumor specimens surgically removed from 99 EBT patients. We also examined whether there was any relationship between expression of p16 and biological malignancy of the tumor by comparing its clinicopathological factors. Consequently, we found that there were three types of p16 expression in the tumor cells; diffuse, heterogeneous and negative types, the percentages of which were 19, 41 and 39%, respectively. Heterogeneous and negative types, whose expression of p16 was considered to be down-regulated, showed scirrhous (p=0.022) and infiltrating growth (p=0.002). In addition, we found that the proportion of down-regulated expression of p16 was different, depending on the location of the tumor. We also observed that the down-regulated p16 expression was the highest in a proportion of patients with the extrahepatic bile duct carcinoma. In contrast, the proportion of down-regulated p16 expression was the least among the patients in the region of the ampulla of Vater with better prognosis, and we showed that the prognosis of patients with down-regulated expression of p16 was the poorest in terms of the cancer location where it is limited to the region of ampulla of Vater. These findings suggest that down-regulated p16 expression is evaluated as a factor of poorer prognosis and also that immunohistochemical pattern of p16 expression becomes a marker reflecting the biological malignancy of EBT patients.
Ampulla of Vater↗
Role of HGF/c-met system in invasion and metastasis of oral squamous cell carcinoma cells in vitro and its clinical significance.
We examined the role of the hepatocyte growth factor (HGF)/c-met system on invasion and metastasis of oral squamous cell carcinoma (SCC) cells. In monolayer culture, exogenous HGF marginally affected the growth of oral SCC cells (BHY, HN, IH) and human gingival epithelial cells (GE). In type I collagen matrix, however, HGF significantly enhanced the invasive growth of the cancer cells (p < 0.05). We detected the expression of c-met (HGF receptor) mRNA in all of the cancer cells, but not in human gingival fibroblasts (GF). Oral SCC cells did not secret HGF protein into the medium, but GF secreted a large amount of HGF protein (15 ng/ml). Furthermore, HGF markedly enhanced the migration of cancer cells in a Transwell invasion chamber. Then, we examined the serum levels of HGF in oral SCC patients, or HGF concentrations in oral cancer tissues. Serum levels of HGF in the patients were significantly higher than those in healthy volunteers (p < 0.05). After initial treatment, all of the tumor-free survivors showed a marked decline in the serum HGF levels. Furthermore, HGF concentrations in metastatic cancer tissues were significantly higher than those of non-metastatic cancer tissues and normal gingiva (p < 0.01). These results suggest that HGF plays an important role in invasion and metastasis of oral SCC cells as a paracrine factor, and an elevated HGF level in the cancer tissue can be a predictive marker for metastasis formation in patients with oral SCC.
Animals↗
Low-dose retinoic acid enhances in vitro invasiveness of human oral squamous-cell-carcinoma cell lines.
Retinoids inhibit the proliferation of several types of tumour cells, and are used for patients with several malignant tumours. In this study, we examined the effect of retinoic acids (RAs) on the invasive potentials of the oral squamous cell carcinoma (SCC) cells, BHY and HNt. BHY cells expressed all of retinoid nuclear receptors (RARalpha, beta, gamma, and RXRalpha) and cytoplasmic retinoic acid binding proteins (CRABP1 and CRABP2). HNt cells lacked the expression of RARbeta, but expressed other nuclear receptors and CRABPs. All-trans retinoic acid (ATRA) and 13-cis retinoic acid (13-cisRA) (10(-6)and 10(-7)M) inhibited the growth of the cells, but low-dose ATRA and 13-cisRA (10(-8)M) marginally affected the growth of the cells. Surprisingly, low-dose RAs enhanced the activity of tissue-type plasminogen activator (tPA), and activated pro-matrix metalloproteinases (proMMP2 and proMMP9). Activation of proMMP2 and proMMP9 was inhibited by aprotinin, a serine-proteinase, tPA inhibitor. Furthermore, low-dose RAs enhanced the in vitro invasiveness of BHY cells. These results indicate that low-dose RAs enhances the in vitro invasiveness of oral SCC cells via an activation of proMMP2 and proMMP9 probably mediated by the induction of tPA.
Antineoplastic Agents↗
Alpha-synuclein has an altered conformation and shows a tight intermolecular interaction with ubiquitin in Lewy bodies.
Alpha-synuclein, a protein in which two mutations have been identified that cause autosomal dominant Parkinson's disease, is thought to serve as a nidus for the development of a Lewy body. We hypothesized that alpha-synuclein would display different intra- and intermolecular associations in Lewy bodies than it does in its normal intracellular compartments. Using sensitive fluorescence resonance energy transfer (FRET) techniques, we found evidence that alpha-synuclein is more compact and in closer association with other alpha-synuclein molecules in Lewy bodies than it is in the neuropil. In addition, we found evidence of a close, direct intermolecular interaction between the N terminus of alpha-synuclein and ubiquitin. These observations provide support for the hypothesis that in Lewy bodies alpha-synuclein adopts an altered three-dimensional structure and undergoes N-terminal ubiquitination.
Hippocampus↗
Intracytoplasmic localization of cathepsin D reflects the invasive potential of gastric carcinoma.
BACKGROUND: The present study was undertaken to investigate the role of cathepsin D in the invasiveness of human gastric cancer. METHODS: Immunohistochemical cathepsin D staining was performed in samples from 29 early gastric adenocarcinomas (papillary or tubular adenocarcinoma) and 15 gastric adenomas, and their adjacent nonneoplastic gastric mucosa. We classified the patterns of cathepsin D immunostaining into four types; type A, fine granular staining in the apical portion: type B, intense coarse granular staining in the apical portion; type C, fine granular staining in the basal portion; and type D, diffuse granular staining throughout the cytoplasm. RESULTS: All of the nonneoplastic mucosa showed an apical cytoplasmic distribution pattern (type A or type B). However, 20% (2/10) of low-grade gastric adenomas and 60% (3/5) of high-grade gastric adenomas showed an abnormal staining pattern. i.e., types C and D. Moreover, 82% (9/11) definite intramucosal gastric adenocarcinomas, and the majority of gastric adenocarcinomas with submucosal invasion [83% (15/18) of those in the mucosal part and 100% (14/ 14) of those in the invasive submucosal part] showed an abnormal staining pattern (types C and D). Interestingly, most of the carcinoma cells invading the stroma and lymphatics showed the type D staining pattern. CONCLUSIONS: These results indicate that an abnormal cytoplasmic staining pattern of cathepsin D may reflect the invasive potential of gastric carcinoma cells.
Adenocarcinoma↗
Precancerous lesions of the colorectum.
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Adenocarcinoma↗
Genetic analysis of a local recurrent tumor after colonic polypectomy.
After polypectomy, the cut end of the polyp is usually examined by light microscopy to assess the risk of recurrent cancer. Here, we report a recurrent tumor that appeared in the colon 6 years after polypectomy, although cancer cells were not observed in hematoxylin and eosin-stained sections of the cut end of the primary polyp. Retrospectively, the primary polyp and the recurrent tumor were analyzed for mutations of the p53 gene. We detected p53 mutations in the primary polyp, even in the cut end of the polyp. The same set of two p53 mutations was detected in the recurrent tumor. These observations indicate a common origin of the primary tumor and the recurrent tumor. We conclude that it is important to analyze p53 mutations in colonic polyps, especially when the cut end of the polyp is difficult to evaluate histologically, in order to predict recurrence.
Adenocarcinoma↗
Development of rotational digital angiography system--clinical value in acute pulmonary thromboembolism.
A rotational digital angiography system was newly developed by authors. During the rotation, a pulse exposure is done at every 1.25 degrees by computer control, so that 288 different projectional images can be acquired following a single injection of contrast medium. Because rotational image data can be easily reconstructed to 3-D images, more useful diagnostic informations are obtained. Rotational digital angiography was performed for 15 patients with acute pulmonary thromboembolism, and massive thromboemboli were revealed in all patients. The diagnostic usefulness of this new system has also proven to be improved in the interventional procedures. At the appropriate angle, the thrombosed vessels were clearly distinguished. Thus, as the tip of the catheter was advanced to the target site, the thrombosed lesion could be exclusively treated by interventional techniques. The device has, therefore, been found to be of great value in the diagnosis and intervention of acute pulmonary thromboembolism.
Acute Disease↗
Alpha-synuclein-enhanced green fluorescent protein fusion proteins form proteasome sensitive inclusions in primary neurons.
Alpha-synuclein accumulates in the brains of sporadic Parkinson's disease patients as a major component of Lewy bodies, and mutations in alpha-synuclein are associated with familial forms of Parkinson's disease. The pathogenic mechanisms that precede and promote the aggregation of alpha-synuclein into Lewy bodies in neurons remain to be determined. Here, we constructed a series of alpha-synuclein-enhanced green fluorescent protein (alpha-synucleinEGFP, SynEGFP) fusion proteins to address whether the Parkinson's disease-associated mutations alter the subcellular distribution of alpha-synuclein, and to use as a tool for experimental manipulations to induce aggregate formation. When transfected into mouse cultured primary neurons, the 49-kDa alpha-synucleinEGFP fusion proteins are partially truncated to a approximately 27-kDa form. This non-fluorescent carboxy-terminally modified fusion protein spontaneously forms inclusions in the neuronal cytoplasm. A marked increase in the accumulation of inclusions is detected following treatment with each of three proteasome inhibitors, n-acetyl-leu-leu-norleucinal, lactacystin and MG132. Interestingly, Ala30Pro alpha-synucleinEGFP does not form the cytoplasmic inclusions that are characteristic of wild-type and Ala53Thr alpha-synucleinEGFP, supporting the idea that the Ala30Pro alpha-synuclein protein conformation differs from wild-type alpha-synuclein. Similar inclusions are formed if alpha-synuclein carboxy-terminus is modified by the addition of a V5/6xHistidine epitope tag. By contrast, overexpression of unmodified alpha-synuclein does not lead to aggregate formation. Furthermore, synphilin-1, an alpha-synuclein interacting protein also found in Lewy bodies, colocalizes with the carboxy-terminally truncated alpha-synuclein fusion protein in discrete cytoplasmic inclusions.Our finding that manipulations of the carboxy-terminus of alpha-synuclein lead to inclusion formation may provide a model for studies of the pathogenic mechanisms of alpha-synuclein aggregation in Lewy bodies.
Animals↗
Interaction of alpha-synuclein and synphilin-1: effect of Parkinson's disease-associated mutations.
alpha-Synuclein is a major component of Lewy bodies, a neuropathological feature of Parkinson's disease. Two alpha-synuclein mutations, Ala53Thr and Ala30Pro, are associated with early onset, familial forms of the disease. Recently, synphilin-1, a protein found to interact with alpha-synuclein by yeast two hybrid techniques, was detected in Lewy bodies. In this study we report the interaction of alpha-synuclein and synphilin-1 in human neuroglioma cells using a sensitive fluorescence resonance energy transfer technique. We demonstrate that the C-terminus of alpha-synuclein is closely associated with the C-terminus of synphilin-1. A weak interaction occurs between the N-terminus of alpha-synuclein and synphilin-1. The familial Parkinson's disease associated mutations of alpha-synuclein (Ala53Thr and Ala30Pro) also demonstrate a strong interaction between their C-terminal regions and synphilin-1. However, compared with wild-type alpha-synuclein, significantly less energy transfer occurs between the C-terminus of Ala53Thr alpha-synuclein and synphilin-1, suggesting that the Ala53Thr mutation alters the conformation of alpha-synuclein in relation to synphilin-1.
Blotting, Western↗