Four functional genotoxic marker genes (Bax, Btg2, Ccng1, and Cdkn1a) discriminate genotoxic hepatocarcinogens from non-genotoxic hepatocarcinogens and non-genotoxic non-hepatocarcinogens in rat public toxicogenomics data, Open TG-GATEs.

Background: Previously, Japanese Environmental Mutagen and Genome Society/Mammalian Mutagenicity Study Group/Toxicogenomics Study Group (JEMS/MMS toxicogenomic study group) proposed 12 genotoxic marker genes (Aen, Bax, Btg2, Ccnf, Ccng1, Cdkn1a, Gdf15, Lrp1, Mbd1, Phlda3, Plk2, and Tubb4b) to discriminate genotoxic hepatocarcinogens (GTHCs) from non-genotoxic hepatocarcinogens (NGTHCs) and non-genotoxic non-hepatocarcinogens (NGTNHCs) in mouse and rat liver using qPCR and RNA-Seq and confirmed in public rat toxicogenomics data, Open TG-GATEs, by principal component analysis (PCA). On the other hand, the U.S.

Postural control exercise without using the upper limbs improves activities of daily living in patients with stroke.

Introduction: Stroke is one of the most common neurological disorders worldwide. Stroke survivors have restricted activities of daily living (ADL) and lower functional independence measures (FIM) after disease onset. Recovery of postural control abilities in patients with stroke is one of the most important therapeutic goals.

Short-term in vivo testing to discriminate genotoxic carcinogens from non-genotoxic carcinogens and non-carcinogens using next-generation RNA sequencing, DNA microarray, and qPCR.

Next-generation RNA sequencing (RNA-Seq) has identified more differentially expressed protein-coding genes (DEGs) and provided a wider quantitative range of expression level changes than conventional DNA microarrays. JEMS·MMS·Toxicogenomics group studied DEGs with targeted RNA-Seq on freshly frozen rat liver tissues and on formalin-fixed paraffin-embedded (FFPE) rat liver tissues after 28 days of treatment with chemicals and quantitative real-time PCR (qPCR) on rat and mouse liver tissues after 4 to 48 h treatment with chemicals and analyzed by principal component analysis (PCA) as statics. Analysis of rat public DNA microarray data (Open TG-GATEs) was also performed.

Human gastric cancer risk screening: From rat pepsinogen studies to the ABC method.

We examined the development of gastric cancer risk screening, from rat pepsinogen studies in an experimental rat gastric carcinogenesis model induced with N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and human pepsinogen studies in the 1970s and 1980s to the recent "ABC method" for human gastric cancer risk screening. First, decreased expression or absence of a major pepsinogen isozyme, PG1, was observed in the rat gastric mucosa from the early stages of gastric carcinogenesis to adenocarcinomas following treatment with MNNG. In the 1980s, decreases in PGI in the human gastric mucosa and serum were identified as markers of atrophic gastritis.

Using FFPE RNA-Seq with 12 marker genes to evaluate genotoxic and non-genotoxic rat hepatocarcinogens.

Introduction: Various challenges have been overcome with regard to applying 'omics technologies for chemical risk assessments. Previously we published results detailing targeted mRNA sequencing (RNA-Seq) on a next generation sequencer using intact RNA derived from freshly frozen rat liver tissues. We successfully discriminated genotoxic hepatocarcinogens (GTHCs) from non-genotoxic hepatocarcinogens (NGTHCs) using 11 selected marker genes.

Preparation of the standard cell lines for reference mutations in cancer gene-panels by genome editing in HEK 293 T/17 cells.

Background: Next Generation Sequencer (NGS) is a powerful tool for a high-throughput sequencing of human genome. It is important to ensure reliability and sensitivity of the sequence data for a clinical use of the NGS. Various cancer-related gene panels such as Oncomine™ or NCC OncoPanel have been developed and used for clinical studies.

Evaluation of 12 mouse marker genes in rat toxicogenomics public data, Open TG-GATEs: Discrimination of genotoxic from non-genotoxic hepatocarcinogens.

Previously, we proposed 12 marker genes (Aen, Bax, Btg2, Ccnf, Ccng1, Cdkn1a, Gdf15, Lrp1, Mbd1, Phlda3, Plk2 and Tubb4b) to discriminate mouse genotoxic hepatocarcinogens (GTHC) from non-genotoxic hepatocarcinogens (NGTHC). This was determined by qPCR and principal component analysis (PCA), as the aim of an in vivo short-term screening for genotoxic hepatocarcinogens. For this paper, we conducted an application study of the 12 mouse marker genes to rat data, Open TG-GATEs (public data).

Using RNA-Seq with 11 marker genes to evaluate 1,4-dioxane compared with typical genotoxic and non-genotoxic rat hepatocarcinogens.

It has long been unclear whether 1,4-dioxane (DO) is a genotoxic hepatocarcinogen (GTHC). Therefore, the present study aimed to evaluate rat GTHCs and non-genotoxic hepatocarcinogens (NGTHCs) via selected gene expression patterns in the liver, as determined by next generation sequencing-targeted mRNA sequencing (RNA-Seq) and principal component analysis (PCA). Previously, we selected 11 marker genes (Aen, Bax, Btg2, Ccnf, Ccng1, Cdkn1a, Lrp1, Mbd1, Phlda3, Plk2, and Tubb4b) to discriminate GTHCs and NGTHCs.

Collaborative studies in toxicogenomics in rodent liver in JEMS·MMS; a useful application of principal component analysis on toxicogenomics.

Toxicogenomics is a rapidly developing discipline focused on the elucidation of the molecular and cellular effects of chemicals on biological systems. As a collaborative study group of Toxicogenomics/JEMS·MMS, we conducted studies on hepatocarcinogens in rodent liver in which 100 candidate marker genes were selected to discriminate genotoxic hepatocarcinogens from non-genotoxic hepatocarcinogens. Differential gene expression induced by 13 chemicals were examined using DNA microarray and quantitative real-time PCR (qPCR), including eight genotoxic hepatocarcinogens [o-aminoazotoluene, chrysene, dibenzo[a,l]pyrene, diethylnitrosamine (DEN), 7,12-dimethylbenz[a]anthracene, dimethylnitrosamine, dipropylnitrosamine and ethylnitrosourea (ENU)], four non-genotoxic hepatocarcinogens [carbon tetrachloride, di(2-ethylhexyl)phthalate (DEHP), phenobarbital and trichloroethylene] and a non-genotoxic non-hepatocarcinogen [ethanol].

An active alternative splicing isoform of human mitochondrial 8-oxoguanine DNA glycosylase (OGG1).

Eight alternatively spliced isoforms of human 8-oxoguanine DNA glycosylase (OGG1) (OGG1-1a, -1b, -1c, -2a, -2b, -2c, -2d and -2e) are registered at the National Center for Biotechnology Information (NCBI). OGG1-1a is present in the nucleus, whereas the other seven isoforms are present in the mitochondria. Recombinant OGG1-1a has been purified and enzyme kinetics determined.

Enzyme kinetics of an alternative splicing isoform of mitochondrial 8-oxoguanine DNA glycosylase, ogg1-1b, and compared with the nuclear ogg1-1a.

Eight alternatively spliced isoforms of human 8-oxoguanine DNA glycosylase (OGG1) (OGG1-1a to -1c and -2a to -2e) are registered in the National Center for Biotechnology Information. OGG1(s) in mitochondria have not yet been fully characterized biochemically. In this study, we purified mitochondrial recombinant OGG1-1b protein and compared its activity with nuclear OGG1-1a protein.

Differential gene expression profiling between genotoxic and non-genotoxic hepatocarcinogens in young rat liver determined by quantitative real-time PCR and principal component analysis.

We recently successfully discriminated mouse genotoxic hepatocarcinogens from non-genotoxic hepatocarcinogens via selected gene expression profiling in the mouse liver based on quantitative real-time PCR (qPCR) and statistical analysis using principal component analysis (PCA). In the present study, we applied these candidate marker genes to rat hepatocarcinogens in the rat liver. qPCR analysis of 33 genes was conducted on liver samples from groups of 4 male 4-week-old F344 rats at 4 and 48 h after a single oral administration of chemicals [2 genotoxic hepatocarcinogens: diethylnitrosamine and 2,6-dinitrotoluene; a non-genotoxic hepatocarcinogen: di(2-ethylhexyl)phthalate; and a non-genotoxic non-hepatocarcinogen: phenacetin].

Discrimination of genotoxic and non-genotoxic hepatocarcinogens by statistical analysis based on gene expression profiling in the mouse liver as determined by quantitative real-time PCR.

The general aim of the present study is to discriminate between mouse genotoxic and non-genotoxic hepatocarcinogens via selected gene expression patterns in the liver as analyzed by quantitative real-time PCR (qPCR) and statistical analysis. qPCR was conducted on liver samples from groups of 5 male, 9-week-old B6C3F(1) mice, at 4 and 48h following a single intraperitoneal administration of chemicals. We quantified 35 genes selected from our previous DNA microarray studies using 12 different chemicals: 8 genotoxic hepatocarcinogens (2-acetylaminofluorene, 2,4-diaminotoluene, diisopropanolnitrosamine, 4-dimethylaminoazobenzene, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone, N-nitrosomorpholine, quinoline and urethane) and 4 non-genotoxic hepatocarcinogens (1,4-dichlorobenzene, dichlorodiphenyltrichloroethane, di(2-ethylhexyl)phthalate and furan).

Identification of BC005512 as a DNA damage responsive murine endogenous retrovirus of GLN family involved in cell growth regulation.

Genotoxicity assessment is of great significance in drug safety evaluation, and microarray is a useful tool widely used to identify genotoxic stress responsive genes. In the present work, by using oligonucleotide microarray in an in vivo model, we identified an unknown gene BC005512 (abbreviated as BC, official full name: cDNA sequence BC005512), whose expression in mouse liver was specifically induced by seven well-known genotoxins (GTXs), but not by non-genotoxins (NGTXs). Bioinformatics revealed that BC was a member of the GLN family of murine endogenous retrovirus (ERV).

Genome-wide gene expression profile of jejunal pouch mucosa as a gastric substitute.

Background/aims: Gastric cancer is the most common cancer in Japan. Genome-wide gene expression in the jejunal pouch mucosa was examined using a DNA microarray and quantitative real-time PCR (qPCR) to evaluate the safety, especially with regard to carcinogenic changes, of the jejunal pouch in patients who showed long-term survival.

Methodology: Biopsy samples of jejunal pouch and jejunal conduit were collected from four patients who had undergone gastrectomy 9 to 13 years previously.

Discrimination analysis of human lung cancer cells associated with histological type and malignancy using Raman spectroscopy.

The Raman spectroscopic technique enables the observation of intracellular molecules without fixation or labeling procedures in situ. Raman spectroscopy is a promising technology for diagnosing cancers-especially lung cancer, one of the most common cancers in humans-and other diseases. The purpose of this study was to find an effective marker for the identification of cancer cells and their malignancy using Raman spectroscopy.

Comparison of lung cancer cell lines representing four histopathological subtypes with gene expression profiling using quantitative real-time PCR.

Background: Lung cancers are the most common type of human malignancy and are intractable. Lung cancers are generally classified into four histopathological subtypes: adenocarcinoma (AD), squamous cell carcinoma (SQ), large cell carcinoma (LC), and small cell carcinoma (SC). Molecular biological characterization of these subtypes has been performed mainly using DNA microarrays.

Light sheet direct Raman imaging technique for observation of mixing of solvents.

The light sheet direct Raman (LSDR) imaging technique is used to obtain wide scope, simultaneous images of samples emitting Raman scattered light, without mapping their point-to-point Raman scattering intensities. A prototype system consisting of a background-free electronically tuned Ti:sapphire laser (BF-ETL), band-pass (BP) filters, and a charge-coupled device (CCD) detector is developed in the present study. The LS excitation method enables us to obtain a wide field of Raman view.

Gastric carcinogenesis by N-Methyl-N-nitrosourea is enhanced in db/db diabetic mice.

In 2005, a Japanese epidemiological study showed that increase in plasma glucose levels is a risk factor for gastric cancer. However, no animal model has hitherto shown any association between diabetes mellitus and neoplasia in the stomach. Diabetic (db/db) mice have obese and diabetic phenotypes, including hyperglycemia, because of disruption of the leptin receptor.

Dose-dependent alterations in gene expression in mouse liver induced by diethylnitrosamine and ethylnitrosourea and determined by quantitative real-time PCR.

We examined the dose-dependency of gene expression changes for 51 genes in mouse liver treated with two N-nitroso genotoxic hepatocarcinogens, diethylnitrosamine (DEN) and ethylnitrosourea (ENU) by quantitative real-time PCR (qPCR). DEN (3, 9, 27 and 80mg/kg bw) or ENU (6, 17, 50 and 150mg/kg bw) was injected intraperitoneally into groups of five male 9-week-old B6C3F(1) mice and the livers were dissected after 4h and 28 days. Total RNA from pooled livers was reverse-transcribed to cDNA and the amount of each gene was quantified by qPCR.

Fluorescence-suppressed Raman technique for quantitative analysis of protein solution using a micro-Raman probe, the shifted excitation method, and partial least squares regression analysis.

A practical Raman analyzing technique with suppression of the strong fluorescent background in order to obtain quantitative information is proposed in the present study. The technique is based on the shifted excitation method and partial least squares regression (PLSR) analysis. The Raman system consists of a single Raman spectrometer, a background-free electrically tunable Ti:Sapphire laser (BF-ETL), and a micro-Raman probe (MRP).

Dendritic cell appearance and differentiation during early and late stages of rat stomach carcinogenesis.

Dendritic cell appearance and differentiation during early and late stages of rat stomach carcinogenesis were studied in the pyloric mucosa. Young male rats were given drinking water with or without N-methyl-N'-nitro-N-nitrosoguanidine (MNNG; 100 mg/liter) for 14 days. Use of competitive RT-PCR and northern blotting showed that MNNG exposure induced 3- to 4-fold greater expression of the genes for integrin beta7 and integrin alphaE2 (identical with antigen OX-62, a dendritic cell marker), as well as three cytokines, IL-4, GM-CSF and TNFalpha, in the stomach pyloric mucosa of resistant Buffalo rats compared to sensitive ACI rats.

Overexpression of NP95 mRNA by tumor promoters in the promotion phase of a two-stage BALB/3T3 cell transformation assay.

We studied altered gene expressions in BALB/3T3 cells treated by different tumor promoters in the promotion phase of a transformation assay, an in vitro model of a two-stage carcinogenicity test, using fluorescent mRNA differential display analysis. Expression of the NP95 gene, which was previously found to be the gene of a murine nuclear protein associated with cell proliferation, was increased in the cultures treated by 12-O-tetradecanoylphorbol-13-acetate (TPA), okadaic acid, and orthovanadate. The upregulation of NP95 mRNA was confirmed by reverse transcription-PCR, and Northern blot.

Appearance of osteonectin-expressing fibroblastic cells in early rat stomach carcinogenesis and stomach tumors induced with N-methyl-N'-nitro-N-nitrosoguanidine.

The present study was designed to define molecular alterations in the initiation stage of rat stomach carcinogenesis. Groups of male Lewis rats, 6 weeks old, were given drinking water with or without N-methyl-N'-nitro-N-nitrosoguanidine (MNNG; 100 mg/liter). Total RNA was isolated from the stomach pyloric mucosa, and fluorescent differential display analysis was performed.