复发性胃癌中p53、p-gp、GST-π、Topo-Ⅱ的表达及其相关性研究

目的探讨p53、p-gp、GST-π、Topo-Ⅱ在胃癌复发对化疗药物耐药率增加过程中的表达意义及其相关性。方法应用S-P免疫组化方法,分析156例胃癌组织中复发前后p53、p-gp、GST-π、Topo-Ⅱ蛋白的阳性表达情况。结果156例胃癌组织中,低分化腺癌患者复发前后p-gp、p-53均显著增加(P<0.05),其中p53与p-gp呈正相关(P<0.05);印戒细胞癌患者复发前后p-gp、p53、GST-π显著增加(p<0.05);粘液腺癌患者复发前后GST-π显著增加(p<0.05);管状腺癌患者复发前后p53、p-gp、GST-π、Topo-Ⅱ均无明显变化。结论低分化腺癌复发后耐药率增加,MDR基因突变可能是一个重要因素,而p53在其中起重要的介导作用;粘液腺癌患者复发后肿瘤细胞对化疗药物耐药性增强可能与GST-π表达增强有关;印戒细胞癌患者复发后肿瘤耐药率增加过程中MDR基因过度表达和GST-π表达增强可能共同起作用。胃癌除上述三种耐药机制外可能还存在其他的作用机制。

The Effect of Helicobacter Pylori Infection on Expression of hMSH2, hMLH1 and p53 in Gastric Carcinogenesis

Objective： To detect the expression of hMSH2, hMLH1 and p53 in gastric epithelial cells with and without Helicobactcr pylori （H. pylori） infection and investigate the relationship between H. pylori infection and these genes in gastric carcinogenesis. Methods： H. pylori infection was detected by rapid urease tests. Expression of hMSH2, hMLHland p53 in gastric cancer （GC） tissue, its adjacent mucosa, gastritic mucosa and normal mucosa was assessed by immunohistochemistry SP method. Results： Positive expression rate of hMSH2 in GC tissue （62.7%） was higher than those in adjacent mucosa （29.4%）, gastritic mucosa （32.4%） and normal mucosa （30.0%） （P〈0.001）. Positive rate of hMSH2 in poorly differentiated adenocarcinoma （76.4%） was higher than those in other carcinomas （54.3%, 53.1%） （P〈0.05）. Positive expression rate of hMLH1 in GC tissue （64.3%） mucosa （82.4%） and normal mucosa （80.0%） was lower than those in adjacent mucosa （84.4%）, gastritic （P〈0.01）. Positive rate of hMLH1 in mucoid carcinoma （43.7%） was lower than those in other carcinomas （78.2%, 64.7%） （P〈0.01）. Positive expression rate of p53 in GC tissue （51.9%） was higher than those in adjacent mucosa （3.1%）, gastritic mucosa （0.0%） and normal mucosa （0.0%） （P〈0.001）. Positive rate of p53 in well differentiated adenocarcinoma （32.6%） was lower than those in other carcinomas （58.8%, 68.7%） （P〈0.01）. Positive rates of hMSH2 and hMLH1 in GC with H. pylori infection were lower than those without the infection, respectively （P〈0.05）. Positive rate of p53 in GC with H. pylori infection （61.4%） was higher than that without the infection （40.6%） （P〈0.05）. Conclusion： Gastric carcinogenesis may be associated with abnormal expression of hMSH2, hMLH1 and p53; H. pylori infection affecting expression of these genes may be one of its carcinogenic mechanisms.

The Relationship between the Expression of p27^(Kip1),p53 and the Infiltration,Metastasis and Prognosis in Gastric Carcinoma

Objective: To study the relationship between the expression of p27Kip1 and p53, and the infiltration, metastasis and prognosis in gastric carcinoma. Methods: The expression of p27Kip1 and p53 at protein level was determined by immunohistochemical assay (two-step method) in 100 cases of gastric carcinoma. Results: Of the 100 cases, the positive rate of p27Kip1 and p53 expressions were 44% and 49%, respectively. In the group of gastric carcinomas with deep infiltration (infiltration group), lymph nodes metastasis group (metastasis group) and death-within-5-years group (death group), the expression of p27Kip1 was statistically lower (_P<0.05). In the metastasis group and death group, the expression of p53 was significantly higher (P<0.05). The results of the monovariate analysis revealed that the 5-year survival rate of the high p27Kip1 expression group was 70.59%, which is higher than those of the low p27Kip1 expression group (54.55%) and the negative experession group (26%). The 5-year survival rate of the high p53 expression group was 19.23%, which was lower than those of the p53 low expression group (43.75%) and the negative group (53.19%). Cox multivariate analysis showed that p27Kip1, like p53, was an independent prognostic index. But p27Klpl protein expression was a stronger independent survival predictor (RR=3.06) than p53 expression (RR=2.33). Conclusion: The low expression of p27Kip1 and the high expression of p53 reflected the more frequent invasion and metastasis, which resulted in the reduced survival of patients. As an independent markers of the gastric carcinoma, the expression of p27Kip1 is more useful than that of p53 in the prognosis prediction of gastric carcinoma.

Expression of nitric oxide synthase in human gastric carcinoma and its relation to p53, PCNA

AIM: To investigate the expression of NOS in gastric carcinoma, and to explore the relationship between the expression of nitric oxide synthases (NOS) and p53, PCNA,pathological features and clinical staging of gastric cancer.METHODS: The activity of NOS protein was investigated in 85 samples of human gastric carcinoma and 25 samples of normal gastric mucosal tissue by biochemical assay. We then examined the expression of NOS, p53, PCNA in 85 samples of human gastric cancer was examined by immunohistochemistry, and NOS mRNA expression in 85 gastric cancer tissue specimens by in situ hybridization.RESULTS: Biochemical assay showed that the activity of NOS was significantly higher in gastric carcinoma than in normal gastric mucosal tissues (t = 0.4161, P＜0.01).Immunohistochemistry revealed that endothelial nitric oxide synthase (eNOS) expressed in all samples of normal gastric mucosa, but only 6 cases of 85 gastric cancer specimens showed weak positive immunohistochemical reactions to eNOS (20%). Inducible nitric oxide synthase (iNOS) was expressed strongly in human gastric carcinoma (81.2%). In situ hybridization analysis showed that iNOS mRNA expression was significantly stronger than eNOS mRNA expression in gastric cancer tissue (x2 = 10.23, P＜0.01). The expression of iNOS in gastric cancer was associated with differentiation, clinical stages or lymph node metastases (r= 0.3426, P＜0.05). However,iNOS expression did not correlate with histological classifications and morphological types. The expression of iNOS was significantly correlated with p53 or PCNA expression (r = 0.3612, P＜0.05). The expression of neuronal nitric oxide synthase (nNOS) was not examined by immunohistochemistry and in situ hybridization in gastric cancer specimens and normal gastric mucosa.CONCLUSION: In human gastric cancer, there is an enhanced expression of iNOS, but not of eNOS. NOS promotes the proliferation of tumor cells and plays an important role in gastric cancer spread. Inactivation of antioncogene p53 and overexpression of iNOS might play a synergetic role in the process of carcinogenesis of human gastric carcinoma.

Role of Δ133p53 isoform in NF-κB inhibitor PDTC-mediated growth inhibition of MKN45 gastric cancer cells

AIM To investigate the role of Δ133p53 isoform in nuclear factor-κB(NF-κB) inhibitor pyrrolidine dithiocarbamate(PDTC)-mediated growth inhibition of MKN45 gastric cancer cells.METHODS The growth rate of MKN45 cells after treatment with different concentrations of only PDTC or PTDC in combination with cisplatin was detected by the CCK-8 assay. m RNA expression levels of Δ133p53, p53β, and the NF-κB p65 subunit and p65 protein levels were detected by reverse transcription-polymerase chain reaction(RT-PCR) and immunofluorescence, respectively. Growth of MKN45 cells was significantly inhibited by PDTC alone in a dose-dependent manner(P < 0.01). Moreover, the inhibitory effect of cisplatin was remarkably enhanced in a dose-dependent manner by co-treatment with PDTC(P < 0.01).RESULTS RT-PCR analysis revealed that m RNA expression of p65 was curbed significantly in a dose-dependent manner by treatment with only PDTC(P < 0.01), and this suppressive effect was further enhanced when co-treated with cisplatin(P < 0.01). With respect to the other p53 isoforms, m RNA level of Δ133p53 was significantly reduced in a dose-dependent manner by treatment with only PDTC or PTDC in combination with cisplatin(P < 0.01), whereas p53β m RNA expression was not altered by PDTC treatment(P > 0.05). A similar tendency of change in p65 protein expression, as observed for the corresponding m RNA, was detected by immunofluorescence analysis(P < 0.01). Pearson correlation analysis demonstrated that Δ133p53 and p65 m RNA expression levels were positively related, while no significant relationship was observed between those of p65 and p53β(r = 0.076, P > 0.01).CONCLUSIONΔ133p53 isoform(not p53β) is required in PDTCinduced inhibition of MKN45 gastric cancer cells, indicating that disturbance in the cross-talk between p53 and NF-κB pathways is a promising target in pharmaceutical research for the development of treatment strategies for gastric cancer.

H pylori (CagA) and Epstein-Barr virus infection in gastric carcinomas:Correlation with p53 mutation and c-Myc,Bcl-2 and Bax expression

AIM: To investigate the interrelationship between H pylori and Epstein-Barr virus (EBV) infection in the gastric carcinogenesis having in focus the p53 mutation and the c-Myc, Bcl-2 and Bax expression. METHODS: seventy-one gastric carcinoma tissues were assessed by polymerase chain reaction (PCR) for H pylori and in situ hybridization for EBV. c-Myc, Bcl-2 and Bax expression were detected by immunohistochemistry and single-stranded conformational polymorphism (SSCP) for p53 mutation. RESULTS: The positivity rates for H pylori and EBV were 94.4% and 8.45%, respectively. The majority of the cases displayed only the H pylori presence. All EBV positive cases were also H pylori positive. None infectious agent was observed in 5.55% of the cases. The intestinal type tumor was more frequent in the co-infected and non-infected groups. The female predominated in the non-infected group showing statistical significance (70.4% vs 29.6%, P=0.039). The Bcl-2 was only detected in the group exclusively infected by H pylori. However, c-Myc and Bax were detected in the three groups but with a low frequency in the co-infected group. Mutation of p53 was present in all groups, with the highest frequencies in the H pylori positive groups. CONCLUSION: The frequency of H pylori infection in gastric carcinomas was high. The presented data indicated that gastric carcinogenesis has different pathways depending of the presence of the two investigated infectious agents, suggesting a possible involvement of H pylori with apoptotic process. The low expression of c-Myc and Bax in the EBV-positive groups suggests that EBV may inhibit the expression of these proteins. Nevertheless, p53 mutation shows to be a relevant alteration, independent of both infectious agents.

Changes of NF-kB,p53,Bcl-2 and caspase in apoptosis induced by JTE-522 in human gastric adenocarcinoma cell line AGS cells:role of reactive oxygen species

AIM: To identify whether JTE-522 can induce apoptosis in AGS cells and ROS also involved in the process, and to investigate the changes in NF-kB, p53, bcl-2 and caspase in the apoptosis process. METHODS: Cell culture, MTT, Electromicroscopy, agarose gel electrophoresis, lucigenin, Western blot and electrophoretic mobility shift assay (EMSA) analysis were employed to investigate the effect of JTE-522 on cell proliferation and apoptosis in AGS cells and related molecular mechanisms. RESULTS: JTE-522 inhibited the growth of AGS cells and induced the apoptosis. Lucigenin assay showed the generation of ROS in cells under incubation with JTE-522. The increased ROS generation might contribute to the induction of AGS cells to apoptosis. EMSA and Western blot revealed that NF-kB activity was almost completely inhibited by preventing the degradation of IkBalpha. Additionally, by using Western blot we confirmed that the level of bcl-2 was decreased, whereas p53 showed a great increase following JTE-522 treatment. Their changes were in a dose-dependent manner. CONCLUSION: These findings suggest that reactive oxygen species, NF-kB, p53, bcl-2 and caspase-3 may play an important role in the induction of apoptosis in AGS cells after treatment with JTE-522.

DNA PLOIDY，EXPRESSION OF p53 PROTEIN AND METASTATIC BEHAVIOUR OF GASTRIC CARCINOMA

DNA ploidy of 57 gastric carcinomas with metastases(12 liver,1 adrenal,4 ovary and 48 lymph node) were measured by flow cytometry.DNA anueploidy was significantly related to liver metastases:9 out of 12 gastric carcinomas with liver metastases were anueploid(75%) as compared to 13 out of 45(28.8%) of cases without liver metastases(P<0.01);the one gastric carcinoma with adrenal metastasis was also anueploid.DNA ploidy was not related to ovarian or lymph node metastases.Another interesting finding was that all of 3 gastric carcinomas with liver metastases which showed a diploid DNA pattern,expressed p53 protein, while all of 3 carcinomas with liver metastases but no p53 protein expression were anueploid.The expression of p53 protein was not related to ovarian metastases.The results suggested that an anueploid DNA pattern and the expression of p53 protein are both objective markers valuable in predicting high risk potential of metastases to the liver,and that the combined detection of these markers can be a most useful method in the follow-up of Patients with gastric carcinoma in detecting those at high risk of developing metastases following surgical resection.Also the poorer prognosis of Patients with gastric carcinoma showing an anueploid DNA pattern may be related to the development of distant organ metastases through the blood vascular system.Furthermore,the clone of gastric carcinoma cells which accumulate p53protein or show an anueploid DNA pattern may have a causative role in the development of liver(&.adrenal) metastases.

The Effect of Helicobacter pylori Infection on hMSH2 and P53 Proteins in Gastric Carcinogenesis

OBJECTIVE To investigate the effect of Helicobacter pylori （H pylon） infection on expression of hMSH2 and P53 and its possible carcinogenic mechanism. METHODS H pylori infection was detected using a rapid urease test and haematoxylin and eosin （H＆E） staining. The hMSH2 and P53 proteins in gastric cancer （GC） tissue, its adjacent mucosa, gastritic mucosa and normal gastric mucosa were detected by the immunohistochemical SP method. RESULTS （1） The positive rate of hMSH2 expression in GC tissue （62.7%）was higher than those in non-cancer tissues （P〈0.001）; the positive rate of hMSH2 expression in poorly differentiated adenocarcinoma （76.4%） was higher than that in other carcinomas （P〈0.05）. The positive rate of P53 expression in GC tissue （51.9%） was higher than that in noncancer tissues （P〈0.001）; the positive rate of P53 expression in well-differentiated adenocarcinoma （32.6 %） was lower than that in other carcinomas （P〈0.01）. （2） The positive rate of hMSH2 expression in GC tissue with H pylori infection was lower than that without the infection （52.8% vs. 74.5%; P〈0.05）. The positive rate of P53 expression in GC tissue with H pylori infection was higher than that without the infection （61.4% vs. 40.6%; P〈0.05）.（3） The expression of hMSH2 and P53 in GC tissue correlated positively （r=0.457, P〈0.01）. CONCLUSION High expression of hMSH2 and P53 as well as their interaction may be involved in gastric carcinogenesis; H pylori infection affecting expression of hMSH2 and P53 may be one of its carcinogenic mechanisms.

Significance and expression of Bax, Survivin and p53 in gastric carcinoma and precancerous lesions using tissue microarray

Objective： To explore the relationship between expressions of apoptosis-related protein Bax, Survivin and p53 and the molecular mechanisms of carcinogenesis and progression of gastric carcinoma. Methods： Tissue microarray and immunohistochemistry were used in this study. Results： The positive rate of Bax protein in gastric cancer （17.7%, 17/96） was significantly lower than those in adjacent normal mucosa （51%）, intestinal metaplasia （69.2%） and dysplasia （75%）, P 〈 0.01. The positive rate of Survivin expression in gastric cancer （80.6%, 89/98） was significantly higher than that in adjacent normal mucosa （3.9%）, P 〈 0.01. The positive rates of Survivin expression in tumors with different organ metastases （in lymph node metastasis 86.2%, liver 100% and ovarian 100%） were statistically higher than in tumors without metastasis （64.3%）, P 〈 0.05. Bax expression was correlated with Survivin but not with rap53 that was closely related to Survivin expression （P 〈 0.05） in gastric cancer. Conclusion： The abnormal expressions of Bax, Survivin and rap53 were correlated with the tumorigenesis and progression of gastric carcinoma. P53 and Survivin genes may share the similar mechanism in regulating cell apoptosis, and because of the mutation, p53 gene may lower its down-regulation to Survivin expression.

Effects of folic acid on epithelial apoptosis and expression of Bcl-2 and p53 in premalignant gastric lesions

AIM: To evaluate the effects of folic acid on epithelial apoptosis and expression of Bcl-2 and p53 in the tissues of premalignant gastric lesions. METHODS: Thirty-eight patients, with premalignant gastric lesions including 18 colonic-type intestinal metaplasia (IM) and 20 mild or moderate dysplasia, were randomly divided into a treatment group (n = 19) receiving folic acid 10 mg thrice daily and a control group (n = 19) receiving sucralfate 1 000 mg thrice daily for 3 mo. All patients underwent endoscopies and four biopsies were taken prior to treatment and repeated after concluding therapy. Folate concentrations in gastric mucosa were measured with chemiluminescent enzyme immunoassay. Epithelial apoptosis and the expression of Bcl-2 and p53 protein in gastric mucosa were detected with flow cytometric assay. RESULTS: The mean of folate concentration in gastric mucosa was 9.03±3.37 μg/g wet wt in the folic acid treatment group, which was significantly higher than 6.83±3.02 μg/g wet wt in the control group. Both the epithelial apoptosis rate and the tumor suppressor p53 expression in gastric mucosa significantly increased after folic acid treatment. In contrast, the expression of Bcl-2 oncogene protein decreased after folic acid therapy. CONCLUSION: These data indicate that folic acid may play an important role in the chemoprevention of gastric carcinogenesis by enhancing gastric epithelial apoptosis in the patients with premalignant lesions.

Effect of Helicobacter pylori infection on p53 expression of gastric mucosa and adenocarcinoma with microsatellite instability

AIM： To investigate the relationship between Helicobacter pyiori（H pylori） infection, microsatellite instability and the expressions of the p53 in gastritis, intestinal metaplasia and gastric adenocarcinoma and to elucidate the mechanism of gastric carcinogenesis relating to Hpyloriinfection. METHODS： One hundred and eight endoscopic biopsies and gastric adenocarcinoma were available for the study including 33 cases of normal, 45 cases of gastritis, 30 cases of intestinal metaplasia, and 46 cases of gastric adenocardnoma. Peripheral blood samples of these patients were also collected. Hpyloriinfection and p53 expressions were detected by means of streptavidin-peroxidase （SP）immunohistochemical method. Microsatellite loci were studied by PCR-SSCP-CE using the markers BAT-26,D17S261, D3S1283, D2S123, and D3S1611. NSI was defined as the peak shift in the DNA of the gastric tissue compared with that of the peripheral blood samples. Based on the number of mutated MSI markers, specimens were charac-terized as high MSI （MSI-H） if they manifested instability at two or more markers, low MSI （MSI-L） if unstable at only one marker, and microsatellite stable （MSS） if they showed no instability at any marker.RESULTS： H pylori infection was detected in the samples of gastritis, intestinal metaptasia, and gastric adenocarcinoma and the infection frequencies were 84.4%, 76.7%, and 65.2%, respectively, whereas no Hpyloriinfection was detected in the samples of normal control. There was as ignificant difference in the infection rates between gastritis and carcinoma samples （P = 0.035）. No MSI was detected in gasbitis samples, one MSI-H and two MSI-L were detected among the 30 intestinal metaplasia samples, and 12 MSI-H and 3 MSI-L were detected in the 46 gastric carcinomas. In those gastric cardnomas, the MSI-H frequency in Hpylori-positive group was significantly higher than that in Hpylori-negative group. No p53 expression was detected in the normal and gastritis samples from dyspeptic patients. P53-positive immunohistochemical staining was detected in 13.3% of intestinal metaplasia samples and in 43.5% of gastric carcinoma samples. The levels of p53 in Hpylori-positive samples were higher than those in the negative group when the carcinoma samples were subdivided into H pylori-positive and -negative groups （P = 0.013）. Eight samples were detected with positive p53 expression out of the 11 MSI-H carcinomas with Hpyloriinfection and no p53 expression could be seen in the Hpylori-negativesamples.CONCLUSION： H py/ori affect the p53 pattern in gastric mucosa when MMR system fails to work. Mutations of the p53 gene seem to be an early event in gastric carcinogenesis.

rAd-p53 enhances the sensitivity of human gastric cancer cells to chemotherapy

AIM： To investigate potential antitumor effects of rAd-p53 by determining if it enhanced sensitivity of gastric cancer cells to chemotherapy. METHODS： Three gastric cancer cell lines with distinct levels of differentiation were treated with various doses of rAd-p53 alone, oxaliplatin （OXA） alone, or a combination of both. Cell growth was assessed with an 3-（4,5）-dimethylthiahiazo （-z-yl）-3,5-diphenytetrazoli- umromide assay and the expression levels of p53, Bax and Bcl-2 were determined by immunohistochemistry. The presence of apoptosis and the expression of caspase-3 were determined using flow cytometry. RESULTS： Treatment with rAd-p53 or OXA alone inhibited gastric cancer cell growth in a time- and dose- dependent manner; moreover, significant synergistic effects were observed when these treatments were combined. Immunohistochemical analysis demonstrated that treatment with rAd-p53 alone, OXA alone or combined treatment led to decreased Bcl-2 expression and increased Bax expression in gastric cancer cells.Furthermore, flow cytometry showed that rAd-p53 alone, OXA alone or combination treatment induced apoptosis of gastric cancer cells, which was accompa- nied by increased expression of caspase-3. CONCLUSION： rAd-p53 enhances the sensitivity of gastric cancer cells to chemotherapy by promoting apoptosis. Thus, our results suggest that p53 gene therapy combined with chemotherapy represents a novel avenue for gastric cancer treatment.

Correlation of p53 over-expression and alteration in p53 gene detected by polymerase chain reaction-single strand conformation polymorphism in adenocarcinoma of gastric cancer patients from India

AIM： To study the alterations in p53 gene among Indian gastric cancer patients and to correlate them with the various clinicopathological parameters. METHODS： A total of 103 gastric cancer patients were included in this study. The p53 alterations were studied by both immunohistochemical method as well as polymerase chain reaction （PCR）-single strand conformation polymorphism （SSCP） analysis. We only studied four （exon 5, 6, 7, and 8） of the 11 ,p53 exons. The alterations in p53 were also correlated with respect to various clinicopathological parameters. RESULTS： Among 103 cases, p53 over-expression and alteration were detected in 37 （35.92%） and 19 （18.44%） cases, respectively. Most of the ,p53 alterations were found at exon 5 （31.54%）, followed by exon 6 （26.31%）, exon 7 （21.04%） and exon 8 （21.04%）. A significant correlation of p53 overexpression was found with p53 alteration （P = 0.000）. Concordance between ,p53 alteration （as detected by SSCP） and over-expression [as detected by immunohistochemistry （IHC）] was found in 75% cases. We found that IHC-positive/SSCP-negative cases accounted for 21% of cases and IHC-negative/SSCP- positive cases accounted for remaining 4% cases. CONCLUSION： Our results show that p53 gene mutations are significantly correlated with p53 protein over-expression, with 75% concordance in over-expression and alteration in the p53 gene, but 25% disconcordance also cautions against the assumption that p53 over-expression is always associated with a gene mutation. There may be other mechanisms responsible for stabilization and accumulation of p53 protein with no evidence of gene mutation that reflect an accumulation of a non-mutated protein, or a false negative SSCP result.

Checkpoint with forkhead-associated and ring finger promoter hypermethylation correlates with microsatellite instability in gastric cancer

AIM:To examine the methylation status of the promoter region of the checkpoint with forkhead-associated and ring fi nger(CHFR) and microsatellite mutator status in 59 primary gastric cancers.METHODS:We investigated the promoter methylation of CHFR in 59 cases of gastric cancer using methylation-specifi c PCR.Five microsatellite loci were analyzed using high-intensity microsatellite analysis reported previously, and p53 gene mutations were investigated by direct sequencing.RESULTS:Twenty cases(33.9%) showed promoter methylation and no relation was observed with the clinicopathological factors.We found that the promoter methylation of CHFR was frequently accompanied with microsatellite instability(MIN).Seven of 20(35.0%) cases showed MIN in hypermethylation of the CHFR tumor, while three of 39(7.7%) cases showed MIN in the non-methylated CHFR tumor(P < 0.01).However, we failed to fi nd any relationship between CHFR methylation and p53 mutation status.CONCLUSION:The coordinated loss of both the mitotic check point function and mismatch repair system suggests the potential to overcome the cell cycle check point, which may lead to an accumulation of mutations.However, the p53 mutation was not related to hypermethylation of the CHFR promoter and MIN, which indicates that an abnormality in p53 occurs as an independent process from the mismatch repair deficiency in carcinogenesis.

Alterations of tumor suppressor and tumor-related genes in the development and progression of gastric cancer

The development and progression of gastric cancer involves a number of genetic and epigenetic alterations of tumor suppressor and tumor-related genes. The majority of differentiated carcinomas arise from intestinal metaplastic mucosa and exhibit structurally altered tumor suppressor genes, typified by p53, which is inactivated via the classic two-hit mechanism, i.e. loss of heterozygosity （LOH） and mutation of the remaining allele. LOH at certain chromosomal loci accumulates during tumor progression. Approximately 20% of differentiated carcinomas show evidence of mutator pathway tumorigenesis due to hMLH1 inactivation via hypermethylation of promoter CpG islands, and exhibit high-frequency microsatellite instability. In contrast, undifferentiated carcinomas rarely exhibit structurally altered tumor suppressor genes. For instance, while methylation of E-cadherin is often observed in undifferentiated carcinomas, mutation of this gene is generally associated with the progression from differentiated to undifferentiated carcinomas. Hypermethylation of tumor suppressor and tumor-related genes, including APC, CHFR, DAP- kinase, DCC, E-cadherin, GSTP1, hMLH1, p16, PTEN, RASSF1A, RUNX3, and TSLC1, can be detected in both differentiated and undifferentiated carcinomas at varying frequencies. However, the significance of the hypermethylation varies according to the analyzed genomic region, and hypermethylation of these genes can also be present in non-neoplastic gastric epithelia. Promoter demethylation of specific genes, such as MAGE and synudein y, can occur during the progressive stages of both histological types, and is associated with patient prognosis. Thus, while the molecular pathways of gastric carcinogenesis are dependent on histological background, specific genetic alterations can still be used for risk assessment, diagnosis, and prognosis.

Apoptotic cell death and its relationship to gastric carcinogenesis

AIM: To investigate the apoptotic process of cells with in the intestinal metaplasia areas co-localizing with chronic gastritis and gastric carcinomas and to analyze the involvement of proteins regulating apoptosis in the process of intestinal metaplasia related gastric carcinogenesis. METHODS: Forty-two gastric carcinoma and seventeen chronic gastritis cases were included in this study. All cases were examined for the existence of intestinal metaplasia. Ten cases randomly selected from each group were processed for TUNEL assay. TUNEL positive cells within the intestinal metaplasia areas, co-localizing either to gastric carcinoma or chronic gastritis, were counted and converted to apoptotic indices. In addition, p53, bcl-2 and bax expression patterns within these tissues were analyzed on the basis of immunohistochemistry. RESULTS: Twenty-eight of the cases were intestinal and 14 of the cases were diffuse type adenocarcinomas. 64% (27/42) of the gastric carcinoma cases had intestinal metaplasia. Intestinal metaplasia co-localized more with intestinal type carcinomas compared with diffuse type carcinomas [75% (21/28) vs 42% (6/14), respectively; P ≤0.05]. The mean apoptotic index in tumor cells was 0.70±0.08. The mean apoptotic index in intestinal metaplasias co-localizing to tumors was significantly higher than that of intestinal metaplasias co-localizing to chronic gastritis (0.70±0.03 vs 0.09±0.01, respectively; P≤0.05). p53 positivity was not observed in areas of intestinal metaplasia adjacent to tumors or chronic gastritis. Intestinal metaplasia areas adjacent to tumors showed lower cytoplasmic bcl-2 positivity compared to intestinal metaplasia areas adjacent to chronic gastritis [55.5% (15/27) vs 70.5% (12/17), respectively]. On the other hand, intestinal metaplasia areas adjacent to tumors showed significantly higher cytoplasmic bax positivity compared to intestinal metaplasia areas adjacent to chronic gastritis [44.4% (12/27) vs 11.7% (2/17), respectively; P ≤0.05]. CONCLUSION: Existence of apoptotic cells on the basis of TUNEL positivity is shown in intestinal metaplasias co-localizing to both diffuse and intestinal type gastric cancers in this study. Our results also suggested bax expression dependent induction of apoptosis especially in intestinal metaplasia areas adjacent to tumors. These findings strongly support the involvement of apoptotic mechanisms in the process of gastric carcinogenesis especially in the transition from intestinal metaplasia to gastric cancer. It may be suggested that induction of apoptosis in intestinal metaplasia areas adjacent to tumors may involve different mechanisms than induction by chronic inflammation.