Thoracoscopic radiofrequency ablation therapy for hepatocellular carcinoma above the diaphragm associated with intractable hemothorax.
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Carcinoma, Hepatocellular↗
Biomedical subjects
M Omata
Publications and source records attributed to M Omata.
Weak response of helicobacter pylori antibody is high risk for gastric cancer: a cross-sectional study of 10,234 endoscoped Japanese.
BACKGROUND: A large number of endoscoped members of the general Japanese population were surveyed to investigate the relationship between Helicobacter pylori infection and gastric cancer. Special attention was given to antibody titer and age of the subjects. METHODS: We performed gastrointestinal endoscopy and measured serum anti-H. pylori antibody in 10,234 consecutive Japanese who participated in a health examination program. Gastric cancer, when suspected, was confirmed by histology. We graded the H. pylori antibody titer into three groups in accordance with optical density values by ELISA: 'strongly positive', 'weakly positive', and 'negative'. RESULTS: Among the 10,234 subjects (men/women, 7.021/3,213; mean age, 49.1 years), 4,909 (48%) were strongly positive, 1,750 (17%) were weakly positive, and 3,575 (35%) were negative for H. pylori antibody. Thirty-seven cases of gastric cancer were found among the 10,234 subjects (0.36%); 23/4,909 (0.47%) in the strongly positive group, 9/1,750 (0.51%) in the weakly positive group, and 5/3,575 (0.14%) in the negative group. Both the strongly and weakly positive groups showed a higher risk of gastric cancer than the negative group. In the subjects over age 60, the weakly positive group seemed to show the highest risk for gastric cancer. CONCLUSIONS: In this investigation of 10,234 Japanese, based on endoscopy results, those with serum H. pylori antibody had an increased risk for gastric cancer, while those 'weakly positive' showed a high risk, particularly in the elderly.
Antibodies, Bacterial↗
Analysis of apoptotic and antiapoptotic signalling pathways induced by Helicobacter pylori.
BACKGROUND AND AIMS: Although it is reported that Helicobacter pylori induces apoptosis on gastric epithelial cells, the mechanism remains unknown. Antiapoptotic effects generated by H pylori have not yet been evaluated. METHODS: (1) H pylori strains (type 1 wild, TN2-DeltacagE, TN2-DeltavacA) were cocultured with MKN45, TMK1, and HeLa cells, and cell viability and apoptosis were assessed by trypan blue exclusion and DNA laddering, respectively. (2) Activation of caspases-3, 7, and 8, cytochrome c release from the mitochondria, and Fas, Fas associated death domain protein (FADD), Bax, Bak, and Bcl-X expression were evaluated by immunoblot analysis. (3) To investigate whether nuclear factor kappa B (NFkappaB) activation induced by cag pathogenicity island (PAI) positive H pylori affects antiapoptosis, MKN45 cells stably expressing super-repressor Ikappabetaalpha were cocultured with H pylori, and cell viability and caspase activation were evaluated. NFkappaB regulated gene expression was also evaluated by ribonuclease protection assay. RESULTS: (1) Wild-type and DeltavacA mutant H pylori induced apoptosis more potently than the DeltacagE mutant. Inhibition of cell contact between H pylori and cancer cells and heat killing H pylori diminished cell death. (2) Caspases-3, 7, and 8 were activated time dependently by H pylori as well as by the agonist anti-Fas. Cytochrome c release from mitochondria was observed and was not inhibited by caspase-8 inhibitor. Although protein expression of Fas, FADD, Bax, Bak, and Bcl-X in the whole cell lysates was not changed by H pylori, Bax was decreased from mitochondria free cytosol suggesting that Bax was translocated into mitochondria. (3) Cell death and the activities of caspases-3 and 8 were promoted in MKN45 cells stably expressing super-repressor Ikappabetaalpha that inhibits NFkappaB activation. Antiapoptotic proteins c-IAP1 and c-IAP2 were upregulated by the wild-type strains. CONCLUSION: cag PAI positive H pylori is capable of inducing apoptotic effects mainly through the mitochondrial pathway. Antiapoptotic effects mediated by NFkappaB activation were also observed.
Adaptor Proteins, Signal Transducing↗
Effects of N-alpha-methyl-histamine on human H(2) receptors expressed in CHO cells.
BACKGROUND: Production of N-alpha-methyl-histamine (NAMH), a histamine H(3) receptor (H3R) agonist, is reportedly promoted in Helicobacter pylori infected human gastric mucosa. NAMH was suggested to act directly on histamine H(2) receptors (H2Rs) in animals to stimulate acid secretion and to be a H2R agonist. As H2Rs and H3Rs play different roles in gastric acid secretion, it is very important to verify that NAMH is a H2R agonist. AIMS: To determine whether NAMH is a H2R agonist, as well as a H3R agonist. METHODS: We used a Chinese hamster ovary (CHO) cell line expressing human H2Rs (CHO-H2R) and control CHO cells. Expression of human H2Rs was confirmed by tiotidine binding. cAMP production in CHO-H2R and control cells in response to histamine or NAMH was measured. cAMP production in response to 10(-7) M NAMH was also measured in the presence or absence of the H2R antagonist famotidine and the H3R antagonist thioperamide. RESULTS: NAMH dose dependently stimulated cAMP productions in CHO-H2R cells. This production was inhibited by famotidine but not by thioperamide. Control CHO cells were unresponsive to either histamine or NAMH. In addition, the effect of NAMH, in terms of cAMP production in CHO-H2R cells, was more potent than that of histamine-that is, with a lower EC(50) concentration and higher maximal cAMP production. Both NAMH and histamine, but not R-alpha-methyl-histamine, effectively inhibited [(3)H] tiotidine binding to CHO-H2R cells. CONCLUSIONS: NAMH, which is produced in the gastric mucosa by H pylori, is a potent H2R agonist as well as a H3R agonist.
Animals↗
Association between severity of type A hepatitis and nucleotide variations in the 5' non-translated region of hepatitis A virus RNA: strains from fulminant hepatitis have fewer nucleotide substitutions.
BACKGROUND: Type A hepatitis is still a considerable problem in both underdeveloped and developed countries. Why some patients progress to fulminant type A hepatitis and others do not is unclear. AIMS: To determine if nucleotide differences in the genome of hepatitis A virus (HAV) are responsible for the range of clinical severities, we analysed the 5' non-translated region (5'NTR) of the HAV genome, which has an internal ribosomal entry site and is important for cap independent translation of the viral message. METHODS: Serum samples from 84 Japanese patients with sporadic type A hepatitis from five distant regions of Japan, comprising 12 patients with fulminant hepatitis (FH), 13 with severe acute hepatitis (AHs), and 59 with acute hepatitis (AH), were examined for HAV RNA. The fragment between nucleotides 75 and 638 of the 5'NTR was amplified by reverse transcription-polymerase chain reaction, and the nucleotide sequence was determined by direct sequencing. RESULTS: Comparison of sequences of the 5'NTR revealed relatively fewer nucleotide substitutions in FH and AHs patients compared with the considerable sequence variations found in strains of AH. This tendency was most prominent between nucleotides 200 and 500. Strains from FH and AHs cases had fewer nucleotide substitutions (p<0.001) in this region. CONCLUSIONS: Nucleotide variations in the central portion of the 5'NTR of HAV may influence the severity of type A hepatitis.
5' Untranslated Regions↗
Analysis of apoptotic and antiapoptotic signalling pathways induced by Helicobacter pylori.
BACKGROUND AND AIMS: Although it is reported that Helicobacter pylori induces apoptosis on gastric epithelial cells, the mechanism remains unknown. Antiapoptotic effects generated by H pylori have not yet been evaluated. METHODS: (1) H pylori strains (type 1 wild, TN2-deltacagE, TN2-deltavacA) were cocultured with MKN45, TMK1, and HeLa cells, and cell viability and apoptosis were assessed by trypan blue exclusion and DNA laddering, respectively. (2) Activation of caspases-3, 7, and 8, cytochrome c release from the mitochondria, and Fas, Fas associated death domain protein (FADD), Bax, Bak, and Bcl-X expression were evaluated by immunoblot analysis. (3) To investigate whether nuclear factor kappa B (NFkappaB) activation induced by cag pathogenicity island (PAI) positive H pylori affects antiapoptosis, MKN45 cells stably expressing super-repressor IkappaBalpha were cocultured with H pylori, and cell viability and caspase activation were evaluated. NFkappaB regulated gene expression was also evaluated by ribonuclease protection assay. RESULTS: (1) Wild-type and deltavacA mutant H pylori induced apoptosis more potently than the deltacagE mutant. Inhibition of cell contact between H pylori and cancer cells and heat killing H pylori diminished cell death. (2) Caspases-3, 7, and 8 were activated time dependently by H pylori as well as by the agonist anti-Fas. Cytochrome c release from mitochondria was observed and was not inhibited by caspase-8 inhibitor. Although protein expression of Fas, FADD, Bax, Bak, and Bcl-X in the whole cell lysates was not changed by H pylori, Bax was decreased from mitochondria free cytosol suggesting that Bax was translocated into mitochondria. (3) Cell death and the activities of caspases-3 and 8 were promoted in MKN45 cells stably expressing super-repressor IkappaBalpha that inhibits NFkappaB activation. Antiapoptotic proteins c-IAP1 and c-IAP2 were upregulated by the wild-type strains. CONCLUSION: cag PAI positive H pylori is capable of inducing apoptotic effects mainly through the mitochondrial pathway. Antiapoptotic effects mediated by NFkappaB activation were also observed.
Apoptosis↗
False positive uptake of metaiodobenzylguanidine in hepatocellular carcinoma.
We present the case of a patient who showed increased accumulation of (123)I-metaiodobenzylguanidine (MIBG) in hepatocellular carcinoma, leading to a false presumptive diagnosis of intrahepatic neuroendocrine tumour. The increase in uptake was not seen on images obtained 4 h after tracer injection but was evident on those taken after 24 h, suggesting slower washout from the liver tumour than from non-tumoural liver parenchyma. Our observations indicate that a non-neuroendocrine malignant tumour may exhibit high accumulation of MIBG associated with prolonged retention.
3-Iodobenzylguanidine↗
Differential expression of the L-plastin gene in human colorectal cancer progression and metastasis.
To identify molecular alterations in the progression of colorectal carcinoma, we analyzed gene expression profiles of colon cancer cell lines derived from primary and metastatic tumors from a single patient. Of 2280 cDNAs investigated using our in-house microarray, the expression of 6 genes (tumor-associated antigen L6, L-plastin, the human homologue of yeast ribosomal protein S28, the B-cell translocation gene, mitochondrial aspartate-aminotransferase, and HLA-A) increased, while that of 2 genes (keratin 5 and phosphoglucomutase) decreased in metastatic-tumor-derived cells compared with primary-tumor-derived cells. Of these genes, we assessed the L-plastin gene, an actin-bundling protein, at the protein level using a tissue microarray consisting of 58 clinically stratified colorectal cancer specimens. Consistent with our microarray results, the expression of L-plastin was significantly correlated with the progression of cancer staging. Therefore, our results suggest that the L-plastin gene is a potential metastatic marker. In addition, combining cDNA microarrays and tissue arrays, as shown here, is thought to facilitate the rapid characterization of candidate biomarkers.
Antigens, Surface↗
Systematic analysis of the TGF-beta-Smad signaling pathway in gastrointestinal cancer cells.
The transforming growth factor-beta (TGF-beta)-Smad signaling pathway has an important role in carcinogenesis. To study the frequency and mechanism of functional impairment of this pathway in human gastrointestinal cancers, we used a reporter assay to examine the response of 38 cell lines (11 colorectal, 9 pancreatic, 10 gastric, and 8 hepatic cancers) to TGF-beta. We then analyzed TGF-beta type II receptor (T beta RII) gene, immunoblots of Smad4, and restoration of the pathway by rescuing T beta R or Smad. We observed impaired signaling in 91% of colorectal, 67% of pancreatic, and 40% of gastric cancer cell lines, but in none of the hepatic cancer cells. We suggest that this pathway does not function as a tumor suppressor in hepatic carcinogenesis. The impairment is due to inactivation of T beta RII and Smad4 in colorectal and pancreatic cancers. However, because the signal was not recovered by rescuing T beta R or Smad genes in TGF-beta-response-defective gastric cancer cell lines, we suggest that novel molecules or mechanisms are involved in the impaired pathway in some gastric cancers.
Blotting, Western↗
The NEDD8 system is essential for cell cycle progression and morphogenetic pathway in mice.
NEDD8/Rub1 is a ubiquitin (Ub)-like molecule that covalently ligates to target proteins through an enzymatic cascade analogous to ubiquitylation. This modifier is known to target all cullin (Cul) family proteins. The latter are essential components of Skp1/Cul-1/F-box protein (SCF)-like Ub ligase complexes, which play critical roles in Ub-mediated proteolysis. To determine the role of the NEDD8 system in mammals, we generated mice deficient in Uba3 gene that encodes a catalytic subunit of NEDD8-activating enzyme. Uba3(-/-) mice died in utero at the periimplantation stage. Mutant embryos showed selective apoptosis of the inner cell mass but not of trophoblastic cells. However, the mutant trophoblastic cells could not enter the S phase of the endoreduplication cycle. This cell cycle arrest was accompanied with aberrant expression of cyclin E and p57(Kip2). These results suggested that the NEDD8 system is essential for both mitotic and the endoreduplicative cell cycle progression. beta-Catenin, a mediator of the Wnt/wingless signaling pathway, which degrades continuously in the cytoplasm through SCF Ub ligase, was also accumulated in the Uba3(-/-) cytoplasm and nucleus. Thus, the NEDD8 system is essential for the regulation of protein degradation pathways involved in cell cycle progression and morphogenesis, possibly through the function of the Cul family proteins.
Animals↗
Distinct mechanism of Helicobacter pylori-mediated NF-kappa B activation between gastric cancer cells and monocytic cells.
NF-kappaB is a critical regulator of genes involved in inflammation. Gastric epithelial cells and macrophages are considered the main sources of pro-inflammatory cytokines. We investigated NF-kappaB activation by Helicobacter pylori in MKN45 gastric epithelial cells and THP-1 monocytic cells. Although, cag pathogenicity island (PAI)-positive H. pylori (wild type) activated NF-kappaB in both cells, isogenic mutant of cagE (DeltacagE) activated it only in THP-1 cells. Supernatant from the wild type culture could activate NF-kappaB in THP-1 cells but not in MKN45 cells. High density cDNA array analysis revealed that mRNA expression of NF-kappaB-regulated genes such as interleukin (IL)-8, tumor necrosis factor-alpha (TNFalpha), and IL-1beta was significantly up-regulated by the wild type in both cells, whereas it was up-regulated by DeltacagE only in THP-1 cells. Experiments using CD14-neutralizing antibody and IL-1 receptor-associated kinase (IRAK) assay showed that both wild type and DeltacagE H. pylori activated NF-kappaB through CD14 and IRAK in THP-1 cells but not in MKN45 cells. Macrophages from C3H/HeJ mice carrying point mutation in the Toll-like receptor 4 (TLR4) gene showed decreased NF-kappaB activation and TNFalpha secretion compared with C3H/HeN mouse macrophage when treated with H. pylori. In conclusion, H. pylori-induced NF-kappaB activation in epithelial cells is dependent on cag PAI and contact but does not involve CD14 and IRAK, whereas in macrophage/monocytic cells it is independent of cag PAI or contact but involves CD14 and TLR4.
Animals↗
NEDD8 recruits E2-ubiquitin to SCF E3 ligase.
NEDD8/Rub1 is a ubiquitin (Ub)-like post-translational modifier that is covalently linked to cullin (Cul)-family proteins in a manner analogous to ubiquitylation. NEDD8 is known to enhance the ubiquitylating activity of the SCF complex (composed of Skp1, Cul-1, ROC1 and F-box protein), but the mechanistic role is largely unknown. Using an in vitro reconstituted system, we report here that NEDD8 modification of Cul-1 enhances recruitment of Ub-conjugating enzyme Ubc4 (E2) to the SCF complex (E3). This recruitment requires thioester linkage of Ub to Ubc4. Our findings indicate that the NEDD8-modifying system accelerates the formation of the E2-E3 complex, which stimulates protein polyubiquitylation.
Anaphase-Promoting Complex-Cyclosome↗
PPARgamma agonists for intestinal ischaemia.
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Dose-Response Relationship, Drug↗
Reduced C-terminal Src kinase activity is correlated inversely with pp60(c-src) activity in colorectal carcinoma.
BACKGROUND: Specific kinase activity of the proto-oncogene product pp60(c-src) is reported to be elevated in patients with carcinoma of the colon, and a novel cytoplasmic protein-tyrosine kinase, C-terminal Src kinase (Csk), has been found to inactivate the members of the Src family protein-tyrosine kinase. In this study, Csk activity and pp60(c-src) activity were examined in colorectal tumors as well as in colon carcinoma cell lines. METHODS: Colorectal carcinoma tissue and adjacent nonneoplastic tissue from 24 patients, from 8 colon carcinoma cell lines, and from 1 normal colon cell line were used. The levels of pp60(c-src) and Csk in colorectal tissue and cell lines were analyzed by Western and/or Northern blot analysis, and their kinase activity levels were measured by in-gel kinase assay. RESULTS: In the samples from 24 patients with colorectal carcinoma, pp60(c-src) kinase activity and protein levels were increased by 7.8 +/- 0.55 and 2.6 +/- 0.13 times the control levels, respectively. Conversely, the Csk protein level and its kinase activity were reduced by 0.53 +/- 0.08 and 0.53 +/- 0.09 times the control levels, respectively. pp60(c-src) kinase activity was correlated inversely with Csk activity (correlation coefficient = -0.71; P < 0.0001). Of the cell lines, pp60(c-src) kinase activity and protein levels, respectively, were 7.4 +/- 1.22 and 1.86 +/- 0.28 times greater than normal control levels. Csk protein level and kinase activity, respectively, were 0.54 +/- 0.13 and 0.52 +/- 0.11 times less normal control levels and were correlated with mRNA amount. CONCLUSIONS: Csk mRNA, protein, and its kinase activity were reduced in colorectal carcinoma and were correlated with pp60(c-src) kinase activity level. The reduced activity of Csk may be involved in the transformation of a subset of colorectal carcinoma.
Aged↗
Palmitoylation of the canine histamine H2 receptor occurs at Cys(305) and is important for cell surface targeting.
To determine the presence and functional role of the histamine H2 receptor (H2R) palmitoylation, a receptor with a Cys(305) to Ala (A(305) receptor) mutation was generated. Wild-type (WT) and A(305) receptors were tagged at their N-termini with a hemagglutinin (HA) epitope. WT, but not A(305), receptors incorporated [3H]palmitate by metabolic labeling, indicating that the H2R is palmitoylated at Cys(305). Immunocytochemistry of WT and A(305) receptors expressed in COS7 cells revealed WT receptors to be distributed at the plasma membrane, while the majority of A(305) receptors were localized intracellularly with only a small portion being at the plasma membrane. However, the affinity of the A(305) receptor for tiotidine was comparable to that of the WT receptor. In addition, when the amounts of cell surface receptors as determined by anti-HA antibody binding were equivalent, A(305) receptors mediated production of more cAMP than WT receptors. Preincubation of COS7 cells expressing each receptor with 10(-5) M histamine for 30 min reduced subsequent cAMP production in response to histamine via the receptors to similar extents, indicating that palmitoylation is not necessary for desensitization. In addition, cell surface A(305) receptors were capable of being internalized from the cell surface at a rate and extent similar to those of WT receptors. Finally, CHO cell lines stably expressing either WT or A(305) receptors were incubated with 10(-5) M histamine for 1, 6, 12 and 24 h. Total amounts of WT and A(305) receptors, as determined by tiotidine binding, were reduced by incubation, indicating downregulation. Downregulation of the A(305) receptor was more extensive than that of the WT receptor. Thus, palmitoylation of the H2R might be important for targeting to the cell surface and stability.
Adenylyl Cyclases↗
cDNA microarray analysis of Helicobacter pylori-mediated alteration of gene expression in gastric cancer cells.
Helicobacter pylori infection stimulates several intracellular signaling pathways and is accompanied by increased gene expression in gastric epithelial cells. High-density cDNA microarray was used to characterize the mRNA expression profile of genes in human gastric cancer cells (MKN45, AGS) cocultured with H. pylori. Coculture with cag pathogenicity island (PAI)-positive H. pylori (wild-type) significantly up-regulated mRNA expression in 8 of 2304 genes tested. In 6 (interleukin-8, I(kappaB)alpha, A20, ERF-1, keratin K7, glutathione peroxidase) of the 8 genes, up-regulation was confirmed by RT-PCR. In coculture with isogenic cagE-negative mutant ((Delta)cagE), which encodes a type IV secretion system with other genes in the cag PAI, no significant up-regulation was found. We further analyzed the role of A20. Transfection of expression vector encoding A20 resulted in an inhibition of H. pylori-mediated NF-kappaB activation, indicating that H. pylori-mediated A20 expression could be a negative regulator of NF-kappaB activation. Taken together, these results indicate the importance of microarray technology as a tool for analyzing the complex interplay between H. pylori and the host.
Adenocarcinoma↗
Simple quantitative assay of alpha-fetoprotein mRNA in liver tissue using the real-time detection polymerase chain reaction assay - its application for clinical use.
Alpha-feto protein (AFP) mRNA levels increase in hepatocellular carcinoma (HCC) cells as compared with non-neoplastic tissue. Therefore, detection of AFP mRNA in blood nuclear cells is useful for the evaluation of treatment efficacy and prognosis of HCC. In this study, simple and reproducible methods were developed to quantify AFP mRNA using the real-time RT-PCR assay (Taq Man assay). By using in vitro synthesized AFP and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) RNA, the sensitivity and dynamic range of the RT-PCR assay were established. AFP mRNA in both HCC and non-neoplastic tissue, as well as in cell lines, were measured using this assay system. The expression of the AFP mRNA level was normalized using the GAPDH house keeping gene product as an endogenous reference. AFP and GAPDH mRNA can be quantified in the range of 10-10(8) copies when using this quantitative assay. Among HCC cell lines, Huh 7 and HepG2 cells, respectively, represented 1.5x10(6) and 6.0x10(5) AFP mRNA/10(6) GAPDH mRNA, in contrast to 6, 23 and 230 AFP mRNA/10(6) GAPDH mRNA for HLE, HLF and PLC/PRF/5 cells, respectively. Other cell lines derived from stomach, pancreas, and colon cancers have 10 AFP mRNA copies/10(6) GAPDH mRNA. In liver tissue from patients with chronic hepatitis, and the non-neoplastic portion of the liver from HCC patients, AFP mRNA distributes from 2.5x10(3) to 5.8x10(4)/10(6) GAPDH transcripts. In contrast, AFP mRNA in tumor cells were more than 100-fold higher than that found in corresponding non-neoplastic portions in two patients who had a high level of AFP in serum. The establishment of the TaqMan quantifying system for AFP mRNA may have important clinical implications.
Journal Article↗
Target gene therapy for alpha-fetoprotein-producing hepatocellular carcinoma by E1B55k-attenuated adenovirus.
Gene therapy using replication-competent adenovirus that selectively propagates in tumor cells may be an effective treatment for cancer. We developed an adenovirus that would be replication specific for hepatocellular carcinoma (HCC). Based on our finding that the E1B55k-deficient adenovirus was able to replicate in human primary hepatocytes, we therefore designed an adenovirus carrying E1A and attenuated E1B gene driven by the alpha-fetoprotein promoter (Adv-AFP-E1AdB), thus restricting the replication specificity in AFP-producing HCC. Replication of Adv-AFP-E1AdB in primary hepatocytes was practically negligible 4 days after infection. Although Adv-AFP-E1AdB replicated slowly in AFP-producing HCC, it efficiently destroyed HCC cells independent of their p53 status. Experiments were conducted in vivo using systemic administration of Adv-AFP-E1AdB and we observed tumor size reduction in nude mice having liver cancer. The use of replication-competent adenovirus deficient of the E1B gene coupled to an AFP-targeting strategy may be a safe and efficacious treatment for HCC.
Adenoviridae↗