The Effect Of Helicobacter Pylori Infection To The Histological Occurrence Of Gastric Cancer And Its Mechanism

BackgroundHelicobacter pylori(H.pylori) is separated and then identified from gastric mucous specimens of chronic gastritis patients by the Australian scholars Warren and Marshall in 1983,and is concerned by much researchers all over the world.In recent years,some studies have shown that Helicobacter pylori infection is an important risk factor induced gastric cancer(GC),and it have been classified as carcinogen category I for human gastric cancer in 1994 by the World Health Organization's affiliated International Agency for Research on Cancer(IARC).Gastric cancer is a common malignant tumor,and its histologic types varied from each other for everyone with its own characteristics.Lauren's classification is a simple and effective method of classification which is related to the organizational structure and cell morphology of gastric cancer,it divides GC into intestinal type and diffuse type.The cells of intestinal type GC generally arrange in a clear tubular-like structure,they are columnar or cubic structure which is similar to intestinal cancer.The cells of diffuse-type GC grow diffusely,generally do not form a tubular-like structure,is lack of cell junction and poorly differentiate.Then how does intestinal type and diffuse type GC happen? How about the incident and development process? It was not clear so far.At present,some scholars considere that intestinal type GC originates the mucosa of gastric intestinal metaplasia,and diffuse-type GC originates the gastric inherent mucosa.However,other scholars suggest that GC come from stem cells in the neck of gastric glands.Such stem cells possess two kinds of genes which were from the stomach and intestinal epithelial cells respectively according to the theory of gene activation.In the process of stem cells cancerization,if the gene possesses intestinal epithelial characteristics is activated,it would form intestinal type GC;if the gene possesses gastric-type epithelial characteristics is activated,it would form diffuse-type GC. What does H.pylori infection impact on the occurrence of gastric cancer,what types of GC does H.pylori lead to,and what mechanism would it be? Most scholars think H.pylori infection primarily be related to the occurrence of intestinal type GC. Some other scholars also think it is related to both intestinal type and diffuse-type of GC,or think it is not related to Lauren's classification of GC.The occurrent mechanism of intestinal type GC reported in the literature is different from diffuse-type of GC.The cancerization of intestinal-type GC begins from the mucosal surface,the process is from gastritis-mucosal epithelium and glandular atrophy-intestinal metaplasia -atypical hyperplasia-gastric cancer,and as H.pylori parasitising on the mucosal surface,it may be one of the reasons for intestinal type GC is closely related to H.pylori infection.The diffuse-type of GC is not going through the process above,it comes from the regeneration of gastric epithelial dysplasia,and begins in the deep mucosa,this is due to genetic factors.Then why GC caused by H.pylori infection is not all intestinal type, but also diffuse-type GC? This may be related to the different genes carried by different individuals as well as the changes of different genes caused by H.pylori infection.What mechanisms does H.pylori infection cause the different histological occurrence of gastric cancer? The current study shows that,H.pylori infection can caused the occurrence of GC through different signal transduction pathways.The signal transduction pathway Wnt/β-catenin plays an important role in the regulation of cell differentiation,migration,proliferation and polarity.This pathway is activated by the Wnt protein upstream,throughβ-catenin accumulation in the cytoplasm,translocation into the nucleus,combination with TCF/LEF,increasing downstream target genes,thus leading to cell proliferation.Saitoh T and his collegues found that when H.pylori infected gastric mucosa,IFNγ,TNFαlevels were caused increasing,and TNFαincreased the expression of Wnt10B,and then caused the occurrence of gastric cancer throughβ-catenin/TCF signaling pathways.The current study showed that, APC/β-catenin/TCF played an important role in the occurrence of gastric cancer, especially in intestinal type GC.Whether H.pylori infection can be related to the occurrence of histological type of GC through Wnt/β-catenin pathways? It was not clear.AimsThe purpose of this study is the effect of H.pylori infection to histological occurrence of gastric cancer,the mechanisms of different histological types of gastric cancer caused by H.pylori infection,providing a theoretical basis on the prevention and treatment of gastric cancer.MethodsThis study selected the gastric mucosa specimens derived by endoscopy from the Hospitals in Zhuanghe Area in Liaoning Province,the detection of H.pylori infection using HE and immunohistochemical method,the detection of E-cadherin,β-catenin, TCF4 protein expression using immunohistochemical staining.The transformation model of H.pylori infection in vitro used the techniques of cell culture,bacterial culture and bacterial and cell co-culture,the observation of cell morphological changes using phase-contrast microscopy and ordinary microscope and electron microscope;the detecion of proliferation and transformation situation using MTT,Ki67 immunohistochemistry,flat cell cloning assay;the E-cadherin,β-catenin, TCF4 protein expression in GES-1 cells infected by H.pylori using double-labeled immunofluorescence staining;the E -cadherin,β-catenin,TCF4 expression at mRNA level in GES-1 cells infected by H.pylori using RT-PCR method.The statistical analysis usedχ~2 test,Fisher's Exact Test for the comparison of the count data between groups,Mann-Whitney Test for the comparison of the measurement data between groups,Spearman correlation analysis for the relevance of each factors.Results1.H.pylori infection rate in the patients with intestinal type gastric cancer was higher than that of the diffuse-type GC(χ~2=6.784,P=0.009).2.The expression rates of E-cadherin,β-catenin,TCF4 protein were higher in the intestinal GC than that of the diffuse-type one.the cell membrane,cytoplasm and nucleus ofβ-catenin were higher in the intestinal GC than that of the diffuse-type(P <0.01).3.In intestinal type GC,there was positively correlation between in the cell membrane and in the cytoplasm ofβ-catenin(r=0.747,P=0.000),between in the cytoplasm and in the nucleus(r=0.347,P=0.000);and there was no correlation between E-cadherin protein and in the cell membrane ofβ-catenin(r=0.012,P=0.906),betweenβ-catenin protein and TCF4 in the nucleus(r=0.146,P=0.143).In diffuse-type GC there was positively correlation between E-cadherin protein and in the cell membrane ofβ-catenin(r=0.225,P=0.025),between in the cell membrane and in the cytoplasm ofβ-catenin(r=0.697,P=0.000),between in the cytoplasm and the nucleus ofβ-catenin(r=0.306,P=0.002);and there was no correlation between in the nuclear ofβ-catenin and TCF4 protein expression(r=0.118,P=0.241).4.In intestinal type GC,β-catenin protein in the cell cytoplasm and TCF4 protein expression rates in H.pylori-positive group were higher than the rates in H.pylori negative group(P<0.05).In diffuse-type GC,E-cadherin protein expression rate in H.pylori-positive group was higher than that of in the H.pylori-negative group(P<0.05),β-catenin protein expression rate in the cytoplasm H.pylori positive group was lower than the H.pylori- negative group(P<0.05).5.After co-culturing H.pylori and GES-1 cell 45 days,there emerged morphological changes,reflecting as the cells different in size,the cell shape irregular, and the cell size increasing,occasionally appearring giant cells;karyokinesis increasing and occasionally appearring pathological karyokinesis.In the transmission electron microscope,the cells microvilli increasing;the cell nucleus increasing,the cell shape irregular,the chromatin increasing which was located under the nuclear membrane,and the nuclear membrane thickening.6.After co-culturing H.pylori and GES-1 cell 45 days,the GES-1 cells MTT OD value,the cells expressed Ki67 and in the plate cloning experiment,GES-1 cells affected by H.pylori higher than that of the GES-1 control cells.7.The results by immunofluorescence showed that,E-cadherin,β-catenin,TCF4 expression in GES-1 control cells was significantly lower than GES-1 cells affected by H.pylori;theβ-catenin expression location changed.8.The results by by RT-PCR showed that,β-catenin,TCF4 expression in GES-1 control cells was significantly lower than GES-1 cells affected by H.pylori;E-cadherin higher than GES-1 cells affected by H.pylori.Conclusion1.H.pylori infection rate in intestinal type GC is higher than the diffuse-type GC.2.The expression rates of E-cadherin,β-catenin and TCF4 protein are higher in the intestinal GC than that of the diffuse-type one.In intestinal type GC H.pylori infection increasesβ-catenin and TCF4 expression,in diffuse-type GC H.pylori infection increases E-cadherin expression and reduces the cytoplasmic expression ofβ-catenin.3.H.pylori can induce the transformation of GES-1 cells,and appear the characteristics of intestinal type GC.4.The transformation of GES-1 cells induced by H.pylori may accomplish throughβ-catenin/TCF4 signal pathway.