| ObjectiveHelicobacter pylori(Hp)is a Gram-negative bacteria that colonizes in the surface of gastric mucosa.Hp infection is a major cause of gastritis,gastric ulcer,gastric lymphoma,and gastric cancer,and is considered as the class I carcinogen by the WHO.However,the molecular mechanism of the development of gastric cancer caused by Hp infection has not yet been elucidated.Among them,DNA damage and repair mechanism caused by Hp infection are hotspot of research in recent years.The development of Hp-related diseases is a complex process.In addition to factors such as host,diet,and environment,Hp virulence factors play an important role in disease progression.The cagA and vacA genes are important proinflammatory factors,and Gastric ulcers are closely related to the occurrence and development of gastric cancer.In this study,we will focus on the effect of Hp and its cagA EPIYA motif type and vac A genotype on the expression of DNA damage marker protein and HR repair key proteins to reveal the molecular mechanism of Hp in promoting the development of gastric cancer.Materials and Methods1.The antral mucosa of chronic superficial gastritis,chronic atrophic gastritis,intestinal metaplasia,and dysplasia were collected.HE staining and immunohistochemistry were used to determine the histopathological grade and Hp infection.2.Immunohistochemical method was used to detect the expression of DNA damage marker protein(?H2AX)and HR repair key proteins(MRE11,Rad51 and CtIP)in each tissue,and analyze the relationship between the expression of proteins and Hp infection.3.DNA of each tissue was extracted and PCR was used to amplify gene fragments of cagA 3' end and vacA gene s,m,i and d regions.Gene sequencing and agarose gel electrophoresis were used to determine the type of EPIYA motif and vacA gene subtype in each tissue and analysis of its relationship with protein expression.4.Statistical analysis: The data obtained were statistically analyzed using SPSS20.0 software.Chi-square test was used to analyze the relationship between genotype and pathological type.The Kruskal-Wallis H test was used for multiple levels of protein expression and Mann-Whitney U test was used for comparison between groups.statistically significant at p<0.05.Results1.For ?H2AX expression,Hp positive group was higher than Hp negative group in CSG,CAG and IM stages,but there was no significant difference between both in Dys stage;For MRE11 expression,Hp positive group was higher than Hp negative group in CSG and CAG stages,but there was no significant difference between both in IM and Dys stage;For Rad51 expression,Hp negative group was higher than Hp positive group in CSG and IM stages,but there was no significant difference between both in CAG and Dys stage;For CtIP expression,Hp negative group was higher than Hp positive group in CSG and IM stages,but there was no significant difference between both in CAG and Dys stage.2.The EPIYA motif of the cagA gene in our region is mainly EPIYA-ABD,and the vacA gene type is mainly s1m1i1d1.Each motif type and vacA genotype had no effect on the expression of DNA damage marker protein and HR repair key proteins.3.The expression of DNA damage marker protein in m1,i1,d1 subgroups was higher than that in m2,i2,d2 subgroups.There was no difference in the expression of HR repair key proteins in each subgroup.Conclusions1.In the early stage of gastric cancer,Hp infection causes DNA damage in human gastric epithelial cells,but it inhibits the expression of some key proteins in the HR repair pathway,which may inhibit precise HR repair and increase the risk of cell malignancy.The pathogenic effect of Hp is obvious in early inflammation.2.The EPIYA motifs of Hp strains in this area are mainly EPIYA-ABD types,and the vacA genotypes are mainly s1m1i1d1.3.The vacA gene m1,i1,d1 subtypes have a stronger effect on DNA damage,while the cagA EPIYA motif type and vacA genotype have no significant effect on the expression of HR repair key proteins. |
PDF Full Text Request