Biomedical subjects

Hiroki Sasaki

Publications and source records attributed to Hiroki Sasaki.

At least 19 recordsLinked to original sources

PubMed2006

Fully automatic diagnostic system for early- and late-onset mild Alzheimer's disease using FDG PET and 3D-SSP.

PURPOSE: The purpose of this study was to design a fully automatic computer-assisted diagnostic system for early- and late-onset mild Alzheimer's disease (AD). METHODS: Glucose metabolic images were obtained from mild AD patients and normal controls using positron emission tomography (PET) and( 18)F-fluorodeoxyglucose (FDG). Two groups of 20 mild AD patients with different ages of onset were examined. A fully automatic diagnostic system using the statistical brain mapping method was established from the early-onset (EO) and late-onset (LO) groups, with mean ages of 59.1 and 70.9 years and mean MMSE scores of 23.3 and 22.8, respectively. Aged-matched normal subjects were used as controls. We compared the diagnostic performance of visual inspection of conventional axial FDG PET images by experts and beginners with that of our fully automatic diagnostic system in another 15 EO and 15 LO AD patients (mean age 58.4 and 71.7, mean MMSE 23.6 and 23.1, respectively) and 30 age-matched normal controls. A receiver operating characteristic (ROC) analysis was performed to compare data. RESULTS: The diagnostic performance of the automatic diagnostic system was comparable with that of visual inspection by experts. The area under the ROC curve for the automatic diagnostic system was 0.967 for EO AD patients and 0.878 for LO AD patients. The mean area under the ROC curve for visual inspection by experts was 0.863 and 0.881 for the EO and LO AD patients, respectively. The mean area under the ROC curve for visual inspection by beginners was 0.828 and 0.717, respectively. CONCLUSION: The fully automatic diagnostic system for EO and LO AD was able to perform at a similar diagnostic level to visual inspection of conventional axial images by experts.

Age of Onset↗

PubMed2006

Homozygous deletion and reduced expression of the DOCK8 gene in human lung cancer.

A homozygous deletion of the DOCK8 (dedicator of cytokinesis 8) locus at chromosome 9p24 was found in a lung cancer cell line by array-CGH analysis. Cloning of the full-length DOCK8 cDNA led us to define that the DOCK8 gene encodes a protein consisting of 2,099 amino acids. DOCK8 was expressed in a variety of human organs, including the lungs, and was also expressed in type II alveolar, bronchiolar epithelial and bronchial epithelial cells, which are considered as being progenitors for lung cancer cells. DOCK8 expression was reduced in 62/71 (87%) primary lung cancers compared with normal lung tissue, and the reduction occurred irrespective of the histological type of lung cancer. 5-Aza-2'-deoxy-cytidine and/or Trichostatin A treatments induced DOCK8 expression in lung cancer cell lines with reduced DOCK8 expression. Therefore, epigenetic mechanisms, including DNA methylation and histone deacetylation, were indicated to be involved in DOCK8 down-regulation in lung cancer cells. Further screening revealed homozygous deletions of the DOCK8 gene in a gastric and a breast cancer cell line. DOCK family proteins have been shown to play roles in regulation of migration, morphology, adhesion and growth of cells. Thus, the present results suggest that genetic and epigenetic inactivation of DOCK8 is involved in the development and/or progression of lung and other cancers by disturbing such regulations.

Azacitidine↗

PubMed2005

In vivo and in vitro characterization of human fibroblasts recruited selectively into human cancer stroma.

Fibroblasts, which are a major component of cancer-induced stroma, can have a significant impact on the progression of adjacent malignant epithelia. To characterize fibroblasts recruited into cancer-induced stroma, we examined the recruitment efficiency of 9 human fibroblast cell lines into experimental tumors generated in immunodeficient mice. Green fluorescence protein (GFP)-labeled fibroblast cell lines and human pancreatic cancer cell line Capan-1 were injected i.p. at different sites; the GFP-labeled cells within xenografts were then analyzed. KM104GFP (bone marrow) and VA-13GFP (lung) were selectively recruited into cancer stroma more efficiently than the other cell lines. KM104GFP cells did not affect tumor volume; however, VA-13GFP cells increased tumor volume by about 2-fold. After 5 cyclic in vivo passages of KM104GFP in Capan-1, we selected a subpopulation with an 8.4-fold higher recruitment efficiency (KM104GFP-5G) compared to parental KM104GFP. KM104GFP-5G also exhibited higher chemotaxis and chemoinvasion activity compared to KM104GFP in response to cancer-released chemoattractant(s). Oligonucleotide microarray analysis identified 8 genes with >3-fold upregulation and 6 genes with >3-fold downregulation in KM104GFP-5G. Immunohistochemistry confirmed that fibroblasts recruited into pancreatic cancer stroma strongly expressed carbonic anhydrase IX and keratin-8, whose transcripts were upregulated in KM104GFP-5G by oligonucleotide microarray analysis, whereas their expression in fibroblasts within noncancerous pancreatic stroma were under the detection level. Our results indicate that fibroblast recruitment is not selective with respect to organ origin and that particular fibroblast subpopulations with specific phenotypic characteristics could be recruited efficiently into cancer-induced stroma.

Animals↗

PubMed2005

Probing the chromosome 9p21 region susceptible to DNA double-strand breaks in human cells in vivo by restriction enzyme transfer.

A restriction enzyme, MspI, was introduced into cultured human cells as a probe to detect genomic regions susceptible to DNA double-strand breaks (DSBs). A 2 h exposure to MspI at a concentration of 8 U/mul produced DSBs at MspI sites in more than 80% of HeLa cells. The sensitivity to digestion was examined on chromosomal DNAs for the region containing the p16 tumor suppressor gene and two other related genes, p14ARF and p15, by Southern blot hybridization analysis and linker-mediated capture of DNA fragments digested in vivo. DNAs for the promoter regions of the three genes, respectively, were sensitive to MspI digestion in HeLa cells, while DNA for the p16 promoter region was less sensitive in lung cancer cells with hypermethylation of the region. Breakpoints for interstitial 9p21 deletions removing the p16/p14ARF/p15 locus in a variety of human cancers were significantly over-represented in the three sensitive regions. The results suggest that the MspI sensitivity in vivo of each genomic region reflects its susceptibility to DSBs that trigger chromosome aberrations in human cells. This method could help us understand the pathogenic significance of differential susceptibility to DSBs among genomic regions in human carcinogenesis.

Cell Cycle Proteins↗

PubMed2005

Effect of differences in cancer cells and tumor growth sites on recruiting bone marrow-derived endothelial cells and myofibroblasts in cancer-induced stroma.

Cancer-stromal interaction is well known to play important roles during cancer progression. Recently we have demonstrated that bone marrow-derived vascular endothelial cells (BMD-VE) and myofibroblasts (BMD-MF) are recruited into the human pancreatic cancer cell line Capan-1 induced stroma. To assess the effect of the difference in cancer cell types on the recruitment of BMD-VE and BMD-MF, 10 kinds of human cancer cell line were implanted into the subcutaneous tissue of the immunodeficient mice transplanted with bone marrow of double-mutant mice (RAG-1-/- beta-gal Tg or RAG-1-/- GFP Tg). The recruitment frequency of BMD-VE (%BMD-VE) and BMD-MF (%BMD-MF), and tumor-associated parameters [tumor volume (TV), microvessel density (MVD) and stromal proportion (%St)] were measured. The correlation among them was analyzed. Although %BMD-VE and %BMD-MF varied (from 0 to 21.6%, 0 to 29.6%, respectively), depending on the cancer cell line, both parameters were significantly correlated with %St (p < 0.005). Furthermore %BMD-VE and %BMD-MF also significantly correlated (p < 0.005). In order to assess the effect of tumor growth sites on the recruitment of the cells of interest, a human pancreatic cancer cell line, Capan-1, was transplanted into 5 different sites: subcutaneous tissue, peritoneum, liver, spleen and lung. Tumors in the subcutaneous tissue and peritoneum induced desmoplastic stroma (%St = 22.7%, 19.5%, respectively) and contained BMD-VE (%BMD-VE = 21.6%, 16.5% respectively) and BMD-MF (%BMD-MF = 29.6%, 24.5%, respectively), but weak stromal induction without recruitment of BMD-VE or -MF was observed in the tumors at of the liver, spleen and lung (%St = 9.7%, 9.1%, 5.4%, respectively). cDNA microarray analysis identified the 29 genes that expression was especially up- or down-regulated in the cell line that induced an abundant stromal reaction. However they did not encoded the molecules that were directly involved in stromal cell recruitment (chemokines), differentiation (cytokines) or proliferation (growth factors). These results indicate that the recruitment of BMD-VE and -MF is required for stromal formation during cancer progression and that the cancer microenvironment is important in stromal reaction and the recruitment of BMD-VE and -MF.

Animals↗

PubMed2005

Frag1, a homolog of alternative replication factor C subunits, links replication stress surveillance with apoptosis.

We report the identification and characterization of a potent regulator of genomic integrity, mouse and human FRAG1 gene, a conserved homolog of replication factor C large subunit that is homologous to the alternative replication factor C subunits Elg1, Ctf18/Chl12, and Rad24 of budding yeast. FRAG1 was identified in a search for key caretaker genes involved in the regulation of genomic stability under conditions of replicative stress. In response to stress, Atr participates in the down-regulation of FRAG1 expression, leading to the induction of apoptosis through the release of Rad9 from damaged chromatin during the S phase of the cell cycle, allowing Rad9-Bcl2 association and induction of proapoptotic Bax protein. We propose that the Frag1 signal pathway, by linking replication stress surveillance with apoptosis induction, plays a central role in determining whether DNA damage is compatible with cell survival or whether it requires cell elimination by apoptosis.

Amino Acid Sequence↗

PubMed2005

Comparison of gray matter and metabolic reduction in mild Alzheimer's disease using FDG-PET and voxel-based morphometric MR studies.

PURPOSE: The aim of this study was to investigate regional differences between morphologic and functional changes in the same patients with mild Alzheimer's disease (AD) using statistical parametric mapping (SPM) and voxel-based morphometry (VBM). METHODS: Thirty patients with very mild AD (mean age 66.8 years, mean MMSE score 24.0) and 30 age- and sex-matched normal volunteers underwent both( 18)F-fluorodeoxyglucose (FDG) positron emission tomography (PET) and three-dimensional spoiled gradient echo (SPGR) magnetic resonance imaging (MRI). Statistical parametric mapping was used to conduct VBM analysis of the morphological data, which were compared voxel by voxel with the results of a similar analysis of the glucose metabolic data. RESULTS: In AD patients, VBM data indicated a significant gray matter volume density decrease in bilateral amygdala/hippocampus complex (p < 0.05, corrected), while FDG-PET analysis showed significant glucose metabolic reductions in the posterior cingulate gyri and the right parietal lobule, compared with those in the normal control group. CONCLUSION: In very mild AD, morphological change occurs in the medial temporal lobes, while in contrast, metabolic changes occur in the posterior cingulate gyri and parietal lobule.

Algorithms↗

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↗

PubMed2005

Mutations and deletions of the CBP gene in human lung cancer.

PURPOSE: Microarray-based comparative genomic hybridization analysis led us to detect a homozygous deletion at the cyclic AMP response element binding protein-binding protein (CBP) locus in a lung cancer cell line. Oncogenic roles of CBP had been suggested by functional and genetic studies; thus, involvement of CBP gene alterations in lung carcinogenesis was investigated by undertaking comprehensive analysis of genetic CBP alterations in human lung cancer. EXPERIMENTAL DESIGN: Fifty-nine cell lines and 95 surgical specimens of lung cancer were analyzed for mutations, homozygous and hemizygous deletions, and expression of the CBP gene. RESULTS: Homozygous CBP deletions, including two intragenic deletions, were detected in three (5.1%) lung cancer cell lines. CBP mutations, including missense, nonsense, and frame-shift mutations, were detected in six (10.2 %) cell lines and five (5.3%) surgical specimens of lung cancer. The wild-type CBP allele was retained in 9 of 11 cases with CBP mutations, and both the wild-type and mutant alleles were expressed in all the six cases with heterozygous CBP mutations examined. Three mutations with amino acid substitutions in the histone acetyltransferase domain caused significant reduction in transcription activation activity of CBP protein in vivo. CONCLUSIONS: A fraction of lung cancers carried mutations and/or deletions of the CBP gene, suggesting that genetic CBP alterations are involved in the genesis and/or progression of a subset of lung cancers.

Acetyltransferases↗

PubMed2005

Direct hepatic fate specification from mouse embryonic stem cells.

The molecules responsible for hepatic differentiation from embryonic stem (ES) cells have yet to be elucidated. Here we have identified growth factors that allow direct hepatic fate-specification from ES cells by using simple adherent monolayer culture conditions. ES cell-derived hepatocytes showed liver-specific characteristics, including several metabolic activities, suggesting that ES cells can differentiate into functional hepatocytes without the requirement for embryoid body (EB) formation, in vivo transplantation, or a coculture system. Most importantly, transplantation of ES cell-derived hepatocytes in mice with cirrhosis showed significant therapeutic effects. In conclusion, this novel system for hepatic fate specification will help elucidate the precise molecular mechanisms of hepatic differentiation in vitro and could represent an attractive approach for developing stem cell therapies for treatment of hepatic disease in humans. Supplementary material for this article can be found on the HEPATOLOGY website ( http://www.interscience.wiley.com/jpages/0270-9139/suppmat/index.html).

Animals↗

PubMed2005

Voxel-based morphometric comparison between early- and late-onset mild Alzheimer's disease and assessment of diagnostic performance of z score images.

BACKGROUND AND PURPOSE: Voxel-based morphometry (VBM), used for detecting brain atrophy, permits comparison of local gray matter concentration at every voxel in an image between two groups. We sought to delineate the specific patterns of cerebral gray matter loss with regard to onset of Alzheimer's disease (AD) by using MR imaging and VBM and to evaluate the diagnostic performance of VBM with Z score images. METHODS: Two groups of 30 patients with mild AD of different ages of onset were examined. Mean ages in the early- and late-onset groups were 60.2 +/- 5.2 and 71.5 +/- 2.6 years, respectively. Control subjects were aged-matched healthy volunteers. Regions of gray matter loss in early- and late-onset AD were examined with VBM. Diagnostic performance of Z score images obtained with the VBM method was evaluated in patients and control subjects by calculating the area under the receiver operating characteristic curve (A(z)). RESULTS: Both AD groups had significantly reduced gray matter in the bilateral medial temporal regions. In addition, the early-onset group had more severe gray matter loss in the bilateral parietal and posterior cingulate cortices and precuneus region. No difference was noted in diagnostic performance of Z score images between the early- (A(z) = 0.9435) and late-onset (A(z) = 0.9018) groups. CONCLUSION: Differences were noted in the patterns of regional gray matter loss in patients with early-onset AD versus those with late-onset AD. Parietotemporal and posterior cingulate gray matter loss was found in early-onset AD but not in late-onset AD. Z score images obtained with VBM had a great diagnostic performance for mild AD and can be applied for detecting mild AD in clinical examinations.

Aged↗

PubMed2004

Gene expression profiling identifies a set of transcripts that are up-regulated inhuman testicular seminoma.

Seminoma constitutes one subtype of human testicular germ cell tumors and is uniformly composed of cells that are morphologically similar to the primordial germ cells and/or the cells in the carcinoma in situ. We performed a genome-wide exploration of the genes that are specifically up-regulated in seminoma by oligonucleotide-based microarray analysis. This revealed 106 genes that are significantly and consistently up-regulated in the seminomas compared to the adjacent normal tissues of the testes. The microarray data were validated by semi-quantitative RT-PCR analysis. Of the 106 genes, 42 mapped to a small number of specific chromosomal regions, namely, 1q21, 2p23, 6p21-22, 7p14-15, 12pll, 12p13, 12q13-14 and 22q12-13. This list of up-regulated genes may be useful in identifying the causative oncogene(s) and/or the origin of seminoma. Furthermore, immunohistochemical analysis revealed that the seminoma cells specifically expressed the six gene products that were selected randomly from the list. These proteins include CCND2 and DNMT3A and may be useful as molecular pathological markers of seminoma.

Animals↗

PubMed2004

The mouse natural killer T cell-associated antigen recognized by U5A2-13 monoclonal antibody is intercellular adhesion molecule-1.

Natural Killer T (NKT) cells in mice are generally defined as NK1.1(+) T cells, although NK1.1 antigen is expressed only in C57BL/6 and related strains. This has precluded investigations of other strains. To find a novel NKT cell surface marker, we generated a monoclonal antibody (mAb), U5A2-13, which recognizes phenotypically and functionally similar populations to NKT cells in naïve mice irrespective of strain. Here, by using a COS-7 expressional cloning system, we molecularly cloned a cDNA encoding a protein reactive with the U5A2-13 mAb and then identified it as intercellular adhesion molecule-1 (ICAM-1). Importantly, the U5A2-13 mAb did not stain hepatic mononuclear cells from ICAM-1 gene disrupted mice. Furthermore, Pepscan method disclosed that the discontinuous epitope for U5A2-13 mAb is composed of three loops located in extracellular domain two of ICAM-1. Overall, U5A2-13, a mAb originally established for mouse NKT cells, recognizes a novel conformational epitope of ICAM-1.

Animals↗

PubMed2004

Identification of genes whose expression is upregulated in lung adenocarcinoma cells in comparison with type II alveolar cells and bronchiolar epithelial cells in vivo.

To identify genes whose expression is upregulated in lung adenocarcinoma (AdC) cells in comparison with noncancerous peripheral lung epithelial cells, type II alveolar cells and bronchiolar epithelial cells, as well as AdC cells, were isolated by laser capture microdissection, and subjected to cDNA microarray analysis of 637 human cancer-related genes. Each of the component cells was obtained from several different individuals and analysed independently. As a comparison, two lung AdC cell lines and two primarily cultured normal lung epithelial cell lines were also subjected to cDNA microarray analysis. Four genes, TOP2A, MMP15, MX2 and KOC1, were commonly upregulated in microdissected AdC cells in comparison with microdissected epithelial cells. Hierarchical clustering analysis revealed that differences in gene-expression profiles were more evident between cultured and uncultured cells than between cancerous and noncancerous cells. To further identify the common molecular targets of AdC cells in vivo, quantitative real-time RT-PCR was performed against the four genes upregulated by cDNA microarray analysis. The TOP2A, MMP15, MX2 and KOC1 genes were overexpressed in 10/10 (100%), 8/10 (80%), 5/10 (50%) and 3/10 (30%) microdissected AdC cell samples, respectively, in comparison with any of nine independently microdissected noncancerous epithelial cell samples. The TOP2A gene was commonly overexpressed in lung AdC cells, as previously reported. In addition, the MMP15 and MX2 genes were identified, for the first time, as being commonly overexpressed in lung AdC cells. These results strongly indicate that the MMP15 and MX2 genes could be novel markers for molecular diagnosis and therapy of lung AdC.

Adenocarcinoma↗

PubMed2004

Highly specific marker genes for detecting minimal gastric cancer cells in cytology negative peritoneal washings.

Peritoneal wash cytology plays a pivotal role in the decision for gastric cancer treatment because advanced gastric cancer often turns out incurable with peritoneal metastasis. Molecular detection of minimal cancer cells from peritoneal washings may overcome the sensitivity boundary of conventional cytology and contribute to the prediction of the disease outcome. To select marker candidates out of ten thousands of genes, we performed microarray analyses in 12 gastric cell lines and 8 peritoneal washings of early stage cases. With 40 candidates selected by the above expression profiling, RT-PCR in 16 representative peritoneal wash samples was performed to identify genes specific to cytology positive samples. The finally selected five genes, CK20, FABP1, MUC2, TFF1, and TFF2, were then evaluated for their utility as a marker for minimal residual disease in 99 peritoneal wash samples. Nested RT-PCR using the five genes showed positive results highly specific to incurable cases (91-100%). With a high specificity, the combination of these five genes succeeded in identifying 6 out of 20 (30%) additional patients with all types of early recurrence that could not be predicted by the conventional method. The six newly identified recurrences included four non-peritoneal ones, showing that RT-PCR using the five genes without a real-time quantitative PCR technique contributes to the detection of minimal residual disease.

Ascitic Fluid↗

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

Frequent and preferential infection of Treponema denticola, Streptococcus mitis, and Streptococcus anginosus in esophageal cancers.

Multiple cancers frequently occur in the upper digestive tract. One possible explanation is that specific bacterial infection stimulates the normal epithelium to initiate inflammation and/or promotes carcinogenesis. This study was undertaken to determine which bacterial species is predominantly associated with esophageal cancer. We examined the bacterial diversity in this type of cancer and in the saliva from healthy people by using a culture-independent molecular method. Here we report the preferential and frequent infection of the oral periodontopathic spirochete Treponema denticola (T. denticola), Streptococcus mitis (S. mitis), and Streptococus anginosus (S. anginosus) in esophageal cancer from different regions of the world, and we also describe the induction of inflammatory cytokines by infection of S. anginosus and S. mitis. Our present data suggest that these three bacteria could have significant roles in the carcinogenic process of many cases of esophageal cancer by causing inflammation and by promoting the carcinogenic process, and that eradication of these three bacteria may decrease the risk of recurrence.

Blotting, Northern↗

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↗