Human Gastric Carcinogenesis Research Articles

Expression of pituitary homeobox 1 gene in human gastric carcinogenesis and its clinicopathological significance

To investigate the effect of pituitary homeobox 1 (PITX1) expression in cases of human gastric cancer on cancer differentiation and progression, and carcinogenesis. Using polyclonal PITX1 antibodies, we studied the expression of PITX1 in normal gastric mucosa, atypical hyperplasia, intestinal metaplasia, and cancer tissue samples from 83 gastric cancer patients by immunohistochemistry. Moreover, semi-reverse transcription polymerase chain reaction (semi-RT-PCR) was performed to detect the mRNA level of PITX1 in three gastric cancer cell lines and a normal gastric epithelial cell line. Subsequently, somatic mutations of the PITX1 gene in 71 gastric cancer patients were analyzed by a combination of denaturing high performance liquid chromatography (DHPLC) and DNA sequencing. Immunohistochemistry showed that PITX1 was strongly or moderately expressed in the parietal cells of normal gastric mucosa (100%), while 55 (66.3%) out of 83 samples of gastric cancers showed decreased PITX1 expression. Moreover, PITX1 expression was reduced in 20 out of 28 cases (71.5%) of intestinal metaplasia, but in only 1 out of 9 cases (11%) of atypical hyperplasia. More importantly, PITX1 expression was significantly associated with the differentiation, position and invasion depth of gastric cancers (r = -0.316, P < 0.01; r = 0.213, P < 0.05; r = -0.259, P < 0.05, respectively). Similarly, levels of PITX1 mRNA were significantly decreased in 2 gastric cancer cell lines, BGC-823 and SGC-7901, compared with the normal gastric epithelial cell line GES-1 (0.306 +/- 0.060 vs 0.722 +/- 0.102, P < 0.05; 0.356 +/- 0.081 vs 0.722 +/- 0.102, P < 0.05, respectively). Nevertheless, no somatic mutation of PITX1 gene was found in 71 samples of gastric cancer by DHPLC analysis followed by sequencing. Down-regulation of PITX1 may be a frequent molecular event in gastric carcinogenesis. Aberrant levels of PITX1 expression may be closely correlated with the progression and differentiation of gastric cancer.

Isolation and characterization of human gastric cell lines with stem cell phenotypes

The aim of this study was to develop an in vitro human gastric stem and/or progenitor cell model that may be used to study the mechanism of gastric carcinogenesis induced by Helicobacter pylori infection. Human gastric biopsy was minced and digested with collagenase and dispase and cultured in a low-calcium medium (serum-free keratinocyte medium; keratinocyte-SFM) supplemented with N-acetyl-L-cysteine and L-ascorbic acid 2-phosphate. Actively proliferating epithelial colonies with sustained growth were isolated and characterized for karyotype and phenotypes related to stem cell characteristics including proliferation and differentiation potential, ability of anchorage-independent growth (AIG), gap junctional intercellular communication (GJIC) and the expression of Oct-4, a transcription factor previously shown to be expressed in embryonic stem cells, adult stem cells and undifferentiated tumor cells. To study the carcinogenic effect of H. pylori infection, gastric stem and/or progenitor cells were incubated with H. pylori culture products and/or N-methyl-N-nitro-N-nitrosoguanidine (MNNG), a chemical carcinogen, to see the telomerase activation. Multiple cell lines with stem cell features were isolated by this new cell culture method. The results based on detailed characterization of one cell clone, KMU-GI2, revealed stem cell features of these cells. The initial clone contained mostly undifferentiated epithelial-like cells, which, upon subculture and propagation, gave rise to a heterogeneous cell population. Single cell-derived subclones, similar to the parental population, retained high differentiation potential and were capable of giving rise to many morphologically different cell types (i.e. epithelial-like, glial or neuron-like, round and various peculiar-shaped cells). Although these cells were normal in karyotype and competent in GJIC, they had the ability to grow in soft agar. Cells expressing epithelial membrane antigen (EMA), mucin 5AC, glial fibrillary acidic protein (GFAP), cytokeratin-18 (CK-18), trefoil factor 1 (TFF-1) and Oct-4 were found in the cell culture, but not E-cadherin-, gastrin- or telomerase-expressing cells. Furthermore, spontaneously immortalized non-tumorigenic clones could be derived from the cell population. After treating these cell cultures with the chemical carcinogen, MNNG and H. pylori culture products for 5 days, telomerase activity and telomerase mRNA expression were significantly elevated, while treatment with either of them showed no effect. The new cell culture method can be used to develop gastric epithelial cell clones with sustained growth from endoscopic biopsy. The gastric cell clone showed several stem and/or progenitor cell phenotypes (i.e. the ability of AIG, high differentiation capacity, high susceptibility to spontaneous immortalization and the expression of Oct-4). The telomerase expression in these gastric stem and/or progenitor cells can be upregulated by exposure to H. pylori culture products and MNNG, an important step in neoplastic transformation. These results show that putative human gastric stem and/or progenitor cell clones can be developed by our method and these cells could be useful for studying the mechanisms of human gastric carcinogenesis including the mechanism of action of H. pylori, as well as the regulation of the proliferation and differentiation of human gastric mucosa.

Prognostic Implications of Cyclin B1, p34cdc2, p27Kip1and p53 Expression in Gastric Cancer

PurposeCell cycle progression is regulated by interactions of specific cyclins and cyclin dependent kinases (CDKs) at the G1-S and G2-M checkpoints and cell cycle deregulation plays a major role in carcinogenesis of human cancers.Patients and MethodsTo investigate the role of cell cycle regulators in the pathogenesis and progression of human gastric cancers, 23 cases of gastric carcinomas were examined for the expression of cyclin B1, p34cdc2, p27Kip1 and p53 by immunohistochemical methods, and gene expression was correlated with various clinicopathologic findings.ResultsOut of 23 cases studied, cyclin B1 was diffusely expressed in 20 cases (87.0%), p34cdc2 in 14 cases (60.9%) and p53 in 12 cases (52.2%), whereas in normal gastric tissues, cyclin B1 and p34cdc2 were weakly expressed and p53 was not expressed. In contrast, p27Kip1 was expressed in only 8.7% of gastric carcinomas compared with 78.3% of normal gastric tissues. There was correlation between the expression of cyclin B1 and expression of p34cdc2 (p = 0.002), between the expression of cyclin B1 and loss of p27Kip1 (p = 0.025), and between the expression of p34cdc2 and loss of p27Kip1 (p = 0.065). In addition, expression of cyclin B1 was correlated with regional lymph node metastasis (p = 0.032).ConclusionOur results indicate that cyclin B1 and p34cdc2 are involved in the genesis or progression of gastric cancers. Furthermore, overexpression of cyclin B1 may play an important role in lymph node metastatic potential of gastric cancer. Thus, abnormal expression of cyclin B1 and CDKs, overexpression of p53 and loss of p27Kip1 expression may play important roles in human gastric carcinogenesis.

High salt diets dose‐dependently promote gastric chemical carcinogenesis in Helicobacter pylori‐infected Mongolian gerbils associated with a shift in mucin production from glandular to surface mucous cells

Intake of salt and salty food is known as a risk factor for gastric carcinogenesis. To examine the dose-dependence and the mechanisms underlying enhancing effects, Mongolian gerbils were treated with N-methyl-N-nitrosourea (MNU), Helicobacter pylori and food containing various concentrations of salt, and were sacrificed after 50 weeks. Among gerbils treated with MNU and H. pylori, the incidences of glandular stomach cancers were 15% in the normal diet group and 33%, 36% and 63% in the 2.5%, 5% and 10% NaCl diet groups, showing dose-dependent increase (p < 0.01). Intermittent intragastric injection of saturated NaCl solution, in contrast, did not promote gastric carcinogenesis. In gerbils infected with H. pylori, a high salt diet was associated with elevation of anti-H. pylori antibody titers, serum gastrin levels and inflammatory cell infiltration in a dose-dependent fashion. Ten percent NaCl diet upregulated the amount of surface mucous cell mucin (p < 0.05), suitable for H. pylori colonization, despite no increment of MUC5AC mRNA, while H. pylori infection itself had an opposing effect, stimulating transcription of MUC6 and increasing the amount of gland mucous cell mucin (GMCM). High salt diet, in turn, decreased the amount of GMCM, which acts against H. pylori infection. In conclusion, the present study demonstrated dose-dependent enhancing effects of salt in gastric chemical carcinogenesis in H. pylori-infected Mongolian gerbils associated with alteration of the mucous microenvironment. Reduction of salt intake could thus be one of the most important chemopreventive methods for human gastric carcinogenesis.

P27kip1 Deficiency Confers Susceptibility to Gastric Carcinogenesis in Helicobacter pylori–Infected Mice

Determining how Helicobacter pylori promotes gastric cancer and whether H pylori eradication decreases cancer risk would be helped by suitable murine models. Mice lacking the cyclin-dependent kinase inhibitor p27kip1 are susceptible to carcinogen-induced tumors. Furthermore, p27 stimulates gastric epithelial apoptosis and inhibits proliferation, expression is decreased by H pylori, and low levels are associated with a poor prognosis in gastric cancer. We therefore evaluated p27-deficient mice as a model for H pylori-associated gastric cancer. Wild-type and p27-/- C57BL/6 mice were infected with H pylori mouse-adapted Sydney strain at 6-8 weeks of age and 6-10 mice of each type were euthanized 15, 30, 45, 60, and 75 weeks later. Uninfected p27-/- mice developed gastric hyperplasia. H pylori-infected p27-/- mice frequently developed intestinal metaplasia (40% at 30 weeks, 67% at 45 weeks), and after 60 weeks 7 of 12 mice developed significant dysplasia and gastric cancer, recapitulating human intestinal-type gastric carcinogenesis. Wild-type mice developed intestinal metaplasia only after 75 weeks of infection; significant gastric dysplasia was observed in 1 animal (P < .05 for each comparison with p27-/- mice). No disease developed in uninfected mice. H pylori infection in p27-/- mice was associated with significantly decreased apoptosis and increased epithelial proliferation, inflammation, and H pylori density compared with infection in wild-type mice. p27 loss and H pylori colonization cooperate to produce gastric cancer. The p27-deficient mouse affords opportunities to examine the pathogenesis of H pylori in gastric carcinogenesis and to test eradication and chemopreventive strategies.

Possible Involvement of RUNX3 Silencing in the Peritoneal Metastases of Gastric Cancers

Our previous results suggested that a lack of RUNX3 function contributed to human gastric carcinogenesis, but the role of RUNX3 in progression and metastasis remains unclear. We examined RUNX3 expression in clinical samples of peritoneal metastases in gastric cancers. Changes in metastatic potential were assessed in animal experiments using stable RUNX3 transfectants of gastric cancer cells. Finally, global expression changes were analyzed using a cDNA microarray. Significant down-regulation of RUNX3 through methylation on the promoter region was observed in primary tumors (75%) as well as in all clinical peritoneal metastases of gastric cancers (100%) compared with normal gastric mucosa. Stable transfection of RUNX3 inhibited cell proliferation slightly, and modest transforming growth factor-beta (TGF-beta)-induced antiproliferative and apoptotic effects were observed. Interestingly, it strongly inhibited peritoneal metastases of gastric cancers in animal model (P < 0.01). Furthermore, we did globally analyzed expression profiles of approximately 21,000 genes in parent cells and stable transfectant of RUNX3 using a cDNA microarray. Microarray analysis identified approximately 28 candidate genes under the possible downstream control of RUNX3, some of these genes were considered to be possibly involved in peritoneal metastases, which were related to signal transduction (vav3, TOLL-like receptor, MAPKK, MET, S1 00A1 1, and cathepsin E), apoptosis (caspase 9), immune responses (CD55 and TLR1O), and cell adhesion (sialyltransferase 1 and galectin 4). Some of the genes are involved in the TGF-beta signaling pathway. These results indicate that silencing of RUNX3 affects expression of important genes involved in aspects of metastasis including cell adhesion, proliferation, apoptosis, and promoting the peritoneal metastasis of gastric cancer. Identification of such genes could suggest new therapeutic modalities and therapeutic targets.

Proteomic identification of differentially‐expressed genes in human gastric carcinomas

Although genetic alterations in proto-oncogenes, tumor-suppressor genes, cell cycle regulators, and cell growth factors have been implicated in the process of human gastric carcinogenesis, the principle carcinogenic mechanisms are not fully understood. In this study, we used a proteomic approach to search for genes that may be involved in gastric carcinogenesis and that might serve as diagnostic markers. We identified nine proteins with increased expression and 13 proteins with decreased expression in gastric carcinomas. The two most notable groups included proteins involved in mitotic checkpoint (MAD1L1 and EB1) and mitochondrial functions (CLPP, COX5A, and ECH1). This suggested that there are links between dysfunctions in these processes and gastric carcinogenesis. We also observed the differential expression of HSP27 and CYR61 proteins in gastric carcinoma, whose expression is known to be altered in other types of tumors. Furthermore, the study identified proteins whose function in gastric carcinomas was previously unsuspected and that may serve as new molecular markers for gastric carcinomas. Importantly, immunohistochemical analyses confirmed that the levels of expression of MAD1L1, HSP27, and CYR61 were altered in gastric carcinoma tissues. Therefore, our study suggested not only that the proteins identified in this study can be useful diagnostic markers but also that a proteomics-based approach is useful for developing a more complete picture of the pathogenesis and function of gastric carcinomas.

Smad4 expression in gastric adenoma and adenocarcinoma: frequent loss of expression in diffuse type of gastric adenocarcinoma.

Smads are signal transducers for the members of the TGF-beta superfamily. Of these Smads, Smad4 is essential for TGF-beta signaling. The purpose of this study was to elucidate Smad4 expression and localization in 65 gastric adenomas, 49 intestinal-type and 39 diffuse type of gastric adenocarcinomas (including 12 cases of fresh frozen tissue) using Real-time RT-PCR and immunohistochemistry. Real-time RT-PCR showed that intestinal type gastric adenocarcinomas have higher Smad4 mRNA expression than diffuse type gastric adenocarcinomas. Immunohistochemical stain for Smad4 revealed that expression of Smad4 was significantly lower in diffuse-type gastric adenocarcinoma than intestinal-type gastric adenocarcinomas. Also, higher Smad4 protein expression in intestinal type gastric adenocarcinomas than overall gastric adenoma was noted. The rate of reduced Smad4 expression was higher in advanced gastric cancer than early gastric cancer. These results suggest that Smad4 might play different roles in human gastric carcinogenesis, especially between intestinal type and diffuse type of gastric adenocarcinoma.

The aetiology and clinical relevance of gastric intestinal metaplasia

Helicobacter pylori causes chronic gastritis in virtually all subjects colonised with this bacterium. This can ultimately lead to a loss of gastric glands [ [1] Kuipers E.J Uyterlinde A.M Peña A.S Roosendaal R Pals G Nelis G.F et al. Long term sequelae of Helicobacter pylori gastritis. Lancet. 1995; 345: 1525-1528 PubMed Scopus (720) Google Scholar ]. The resulting atrophic gastritis and intestinal metaplasia increase the risk of dysplasia and gastric cancer [ [2] Ekstrom A.M Held M Hansson L.-E Engstrand L Nyren O Helicobacter pylori in gastric cancer established by CagA immunoblot as a marker of past infection. Gastroenterology. 2001; 121: 784-791 Abstract Full Text Full Text PDF PubMed Scopus (370) Google Scholar ]. Most cases of gastric cancer occur on a background of H. pylori infection and chronic gastritis with gland loss [ 3 Correa P Human gastric carcinogenesis: a multistep and multifactorial process. Cancer Res. 1992; 52: 6735-6740 PubMed Google Scholar , 4 Sipponen P Kekki M Haapakoski J Ihamäki T Siurala M Gastric cancer risk in chronic atrophic gastritis: statistical calculations of cross-sectional data. Int. J. Cancer. 1985; 35: 173-177 Crossref PubMed Scopus (331) Google Scholar , 5 Uemura N Okamoto S Yamamoto S Mutsumura N Yamaguchi S Yamakido M et al. Helicobacter pylori infection and the development of gastric cancer. N. Engl. J. Med. 2001; 345: 784-789 Crossref PubMed Scopus (3613) Google Scholar ]. The rate of progression to atrophic gastritis is slow and dependent on the severity and distribution of gastritis.

Aberrant methylation of integrin alpha4 gene in human gastric cancer cells.

Integrins are adhesion receptors that mediate both cell-extracellular matrix and cell-cell interactions. It has also been reported that the loss of integrin alpha4 expression might be associated with metastasis in several cancers. However, the molecular mechanism for loss of their expression in cancers has not been explored. In the present study, we found that the integrin alpha4 expression is lost in human gastric cancer cell lines and that this is recovered by treatment with DNA methyltransferase inhibitor, implying transcriptional silencing by DNA methylation. Methylation-specific PCR (MSP) and bisulfite genomic DNA sequencing demonstrated the CpG methylation-dependent silencing of integrin alpha4 expression in eight of nine (88.8%) gastric cancer cell lines and in 84.7% of 46 primary tumors. We also investigated whether the restoration of integrin alpha4 in integrin alpha4-inactivated cells affects their ability to invade extracellular matrix, using matrigel assays. Interestingly, integrin alpha4-stable transfectants had markedly less invasive ability than the parental cells. Taken together, these results suggest that the transcriptional repression of the integrin alpha4 gene is caused by aberrant DNA methylation, and that this may play an important role in human gastric carcinogenesis.

Helicobacter pylori and gastric malignancies.

Despite decreasing incidence during the last 50 years, gastric cancer still ranks as one of the most frequent cancers. A multifactorial model of human gastric carcinogenesis is currently accepted in which different dietary and nondietary factors, including genetic susceptibility of the host and Helicobacter pylori infection are involved at different stages in the cancer process. On the molecular level, at least two phenotypes, associated with distinct pathways of genome destabilization, have been identified. However, applying new technologies such as cDNA microarrays a new era in the analysis of molecular markers has started. This molecular technology may open the path towards novel treatment modalities, i.e. gene therapy. Epidemiological, biological, and molecular genetic studies have also implicated the role of H. pylori in lymphomagenesis. Knowledge of pathogenesis and therapy is increasing while good epidemiological data are rare. Many studies have demonstrated that MALT-type lymphomas develop along two different pathways: t(11;18)-positive cases, and t(11;18)-negative cases. Meanwhile, a third translocation could be detected, the t(14;18), opening the discussion of a possible third pathway of lymphoma development.

No involvement of β- catenin gene mutation in gastric carcinomas induced by N-methyl- N-nitrosourea in male F344 rats

The agent N-methyl- N-nitrosourea (MNU) is a direct acting carcinogen and induces well-differentiated adenocarcinoma on the rat gastric mucosa. In this study, 27 histopathologically verified gastric carcinomas induced in male F344 rats were analyzed for mutations in the N-terminal phosphorylation sites (codons 1–51) of the β -catenin gene by using polymerase chain reaction (PCR)-single strand conformation polymorphism (SSCP) assays. In parallel studies, the specific localization of the β-catenin protein was also examined by immunohistochemical analysis. No mutations in the β -catenin gene were found in any of 27 gastric carcinomas induced by MNU. Immunohistochemical analysis resulted in the β-catenin protein to be localized in the plasma membrane but cytoplasmic and/or nuclear accumulation of β-catenin was not identified in any of these carcinomas. These results suggest that mutations in the β -catenin gene are less contributory to the development of rat gastric carcinomas induced by MNU. This animal model may provide a system for evaluating the mechanism of human gastric carcinogenesis that is not associated with β -catenin gene mutations.

Molecular cloning of p53 cDNA of Mongolian gerbil and establishment of yeast p53 functional assay system.

Epidemiological studies have shown a correlation between Helicobacter pylori infection and human gastric carcinogenesis. A Mongolian gerbil model has demonstrated that H. pylori infection induced gastric carcinoma. However, the disadvantage of this animal model is a lack of information regarding the cellular genes involved in oncogenesis. Mutation of the p53 gene is one of the most common steps in gastric carcinogenesis. In this study, we aimed to clone the p53 gene of the Mongolian gerbil and detect the functional mutations in H. pylori-infected animals. The p53 complementary DNA (cDNA) of Mongolian gerbil was cloned by the methods of reverse-transcribed polymerase chain reaction and rapid amplification of cDNA ends. The p53 cDNA of Mongolian gerbil has a 78.8% homology to that of humans. A novel yeast p53 assay system was established and enabled to detect the functional mutations of the p53 gene in the stomach of the Mongolian gerbil. This is the first report of the complete sequence of wild-type p53 cDNA of the Mongolian gerbil. This genetic information and an assay system designed to detect the functional mutations of the p53 gene are useful for further investigations of gastric oncogenesis in this animal model.

Tributyrin inhibits human gastric cancer SGC-7901 cell growth by inducing apoptosis and DNA synthesis arrest.

To evaluate the effects of tributyrin, a pro-drug of natural butyrate and a neutral short-chain fatty acid triglyceride, on the growth inhibition of human gastric cancer SGC-7901 cell. Human gastric cancer SGC-7901 cells were exposed to tributyrin at 0.5, 1, 2, 5, 10 and 50 mmol/L(-1) for 24-72 h. MTT assay was applied to detect the cell proliferation. [(3)H]-TdR uptake was measured to determine DNA synthesis. Apoptotic morphology was observed by electron microscopy and Hoechst-33258 staining. Flow cytometry and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay were performed to detect tributyrin-triggered apoptosis. The expressions of PARP, Bcl-2 and Bax were examined by Western blot assay. Tributyrin could initiate growth inhibition of SGC-7901 cell in a dose- and time-dependent manner. [(3)H]-TdR uptake by SGC-7901 cells was reduced to 33.6 % after 48 h treatment with 2 mmol/L(-1) tributyrin, compared with the control (P<0.05). Apoptotic morphology was detected by TUNEL assay. Flow cytometry revealed that tributyrin could induce apoptosis of SGC-7901 cells in dose-dependent manner. After 48 hours incubation with tributyrin at 2 mmol/L(-1), the level of Bcl-2 protein was lowered, and the level of Bax protein was increased in SGC-7901, accompanied by PARP cleavage. Tributyrin could inhibit the growth of gastric cancer cells effectively in vitro by inhibiting DNA synthesis and inducing apoptosis, which was associated with the down-regulated Bcl-2 expression and the up-regulated Bax expression. Therefore, tributyrin might be a promising chemopreventive and chemotherapeutic agent against human gastric carcinogenesis.

Inhibition of growth of mouse gastric cancer cells by Runx3, a novel tumor suppressor.

We reported recently that the silencing of RUNX3 is causally related to gastric cancer in humans. Here we report that in three of four cell lines derived from N-methyl-N-nitrosourea-induced mouse glandular stomach carcinomas, Runx3 is silenced due to hypermethylation of CpG islands in the promoter region, as we also observed for human gastric cancer cells. Although two of the sites we tested in the promoter of the fourth line were not methylated, in all four cases the silencing of Runx3 could be reversed by treatment of the cells with 5'-azacytidine and trichostatin A. Interestingly, the exogenous expression of RUNX3 in cell lines that do not express the endogenous gene caused an inhibition of growth in soft agar, suggesting that anchorage-independent growth could be used as an assay of RUNX3 activity in vitro. These observations suggest that the mouse system described here may be useful as a model for the study of human gastric carcinogenesis.

幽门螺杆菌感染与不同胃粘膜病变中PCNA、bcl—2表达的关系

Objective To study the relationship between Hp infection and gastric epithelial cell proliferation as well as apoptosis in different gastric mucosa lesions.Methods Hp infection?PCNA and bcl\|2 expression?DNA content and aneuploidy of CAG?IM?GED and GC in 312 gastric specimens were detected by means of SP immunohischemical staining and image analysis.Results In Hp positive cases, positive rates and overexpression rates of PCNA and bcl\|2 expressions in gastric carcinogenesis were progressively increased?Overexpression rates of PCNA in IMIII?GEDII\|III and intestinal type GC were higher than those in IMI\|II?GEDI and IMIII?GEDII\|III respectively ( P 0.05, P 0.01).The detection results of DNA content and aneuploidy status were in keeping with expressions of PCNA. In Hp negative cases, expressions of PCNA?bcl\|2 and detection results of DNA content?aneuploidy status were different from Hp positive cases.Conclusions In the gastric carcinogenesis, Hp infection might promote proliferation and decrease apoptosis of gastric epithelial cells.The severe imbalance of cell proliferative and apoptotic processes was an important mechanism of human gastric carcinogenesis implicated in Hp infection.

Frequent mtDNA mutations and its role in gastric carcinogenesis

In order to disclose the relationship between mutations of mitochondrial DNA (mtDNA) and gastric carcinogenesis, we screened the entire mtDNA sequence in 30 cases of human gastric cancer and matched normal tissues by using denaturing high-performance liquid chromatography (DHPLC) and DNA sequencing. Our data showed that high frequency (66.7%, 20÷30) of mitochondrial genome mutation occurred in gastric cancer. Among these variants, 17 cases (56.7%, 17÷30) were identified to be somatic mutation. High level mutant frequency was found inND4, ND5 coding genes and D-loop control region, which was 36.7%, 26.7% and 30% respectively. Comparing with complexes III, IV and Vof the electron transport chain, we found that variants appeared to be more frequent in the subunit genes of complex I. Most of mutations were base substitutions (85.4%, 41÷48). Our results suggested that mutations of subunit genes encoding complex I, especiallyND3, ND4 andND5 genes, might contribute to human gastric carcinogenesis.

Alteration of P73 and P51 genes and its significance in human gastric carcinogenesis

OBJECTIVE: To evaluate the relatinship between the expression of P51, P73 and the oncogenesis and development of human gastric carcinoma. METHODS: The expression of P73 mRNA were detected both in 32 human gastric carcinoma tissues and adjacent normal gastric tissues by reverse transcriptase polymerase chain reaction (RT-PCR). RESULTS: Overexpressions of P73 mRNA were found in 17/32 gastric carcinoma tissues,in 2/32 adjacent normal gastric tissues.The positive expression rate of P73 mRNA in gastric carcinooma tissues was significantly higher than in adjacent normal gastric tissues( P<0.01). However, a significant correlation was found between the positive expression rate of P73 mRNA in gastric carcinoma tissues and the TNM staging(P<0.05). THe low expressions of P51A mRNA and P51B were found in all gastric carcinoma tissues and adjacent normal gastric tissues. The expression of P51A in gastric carcinoma tissues were much higher than adjacent normal gastric tissues (P<0.05). The expression of P51B is no significant correlation was observed between gastirc carcinoma tissues and adjacent normal gastric tissues. CONCLUSION: The results suggest that there is an overexpression odf P73 and P51A mRNA in gastric cancer tissues, and their expressions is relationship with oncogenesis and developnment of gastric carcinoma.

Helicobacter pylori-stimulated EGF receptor transactivation requires metalloprotease cleavage of HB-EGF

Helicobacter pylori has a major aetiological role in human gastric carcinogenesis but the cellular and molecular pathways by which infection promotes transformation remain to be resolved. This study demonstrates that H. pylori exposure to MKN-1, ST42, and MKN-28 gastric epithelial tumour cells results in the activation of HB-EGF gene expression and EGFR tyrosine phosphorylation. These cell responses are induced by both cagPAI positive and cagPAI negative H. pylori strains and are dependent on cell surface expression of the HB-EGF precursor. The induction of HB-EGF gene transcription by H. pylori requires metalloprotease-, EGFR-, and Mek1-activities, indicating the involvement of the “triple membrane passing signal” (TMPS) for EGFR transactivation. Moreover, the release of the inflammatory cytokine IL-8 by cells exposed to H. pylori is significantly impaired by inhibitors of TMPS pathway elements. Our findings support a model in which H. pylori triggers constitutive EGFR signal activation, which enhances IL-8 production, and initiates neoplastic transformation of gastric epithelial cells.

Genomic structure of the human β-PIX gene and its alteration in gastric cancer

β-PIX, a newly identified p21-activated kinase (PAK)-interacting exchange factors (PIX), encodes a guanine nucleotide exchange factor for Rho guanosine triphosphatases. Characterization of β-PIX gene was performed using the BAC Library method. The β-PIX gene has 17 exons and an A/T polymorphism at the 32nd base upstream of the intron/exon junction of exon 7. The frequencies of genotypes A/T, A/A and T/T were 23.6% (13/55), 72.7% (40/55) and 3.6% (2/55), respectively; these frequencies are in Hardy–Weinberg equilibrium. Two out of 14 informative tumors (14.3%) were shown to have lost their heterozygosity at this locus, but no mutations in the remaining alleles were detected. In addition, we examined the gene-expression profile in another set of 30 gastric samples, but no significant over-expression of either the β-PIX gene or the α-PIX gene was found. Though the β-PIX gene has been speculated to potentially have tumor-related biological characteristics, the findings of the present study suggest that the involvement of β-PIX gene in human gastric carcinogenesis is minimal.