Background:SEMA3B is a candidate tumor suppressor gene,which plays an essential role in tumorigenesis and progression of a wide variety of cancer. Aims:To explore preliminarily the influence of DNA methylation on regulation of SEMA3B gene expression in gastric cancer. Methods:Real-time PCR was used to determine the expression of SEMA3B mRNA in six human gastric cancer cell lines(SGC7901,AGS,MGC803,BGC823,MKN45,and HGC27),one gastric epithelial cell line(GES-1),and 41 gastric cancer tissue and paired adjacent noncancerous tissue specimens. Methylation specific PCR was used to detect the promoter methylation of SEMA3B gene,and correlation between methylation of SEMA3B gene and clinicopathological characteristics of gastric cancer was analyzed. After treated with a demethylation agent,5-Aza-dC(10μmol/L),for 72 hours,the methylation and mRNA expression of SEMA3B gene were re-examined in above-mentioned cell lines. Results:SEMA3B mRNA expression was significantly lower in six gastric cancer cell lines and gastric cancer tissues than that in GES-1 cells(P <0. 001)and paired adjacent noncancerous tissues(P <0. 01). Promoter methylation of SEMA3B gene could be detected in all six gastric cancer cell lines but not GES-1 cells. Frequency of SEMA3B gene methylation was significantly higher in gastric cancer tissues than in paired adjacent noncancerous tissues (67. 6% vs. 32. 4%,P <0. 01),and the frequency was correlated with differentiation and lymph node metastasis of gastric cancer(P<0. 05). After treated with 5-Aza-dC,hypermethylation of SEMA3B gene in 5 gastric cancer cell lines was decreased,accompanied by up-regulation of SEMA3B mRNA expression. Conclusions:Hypermethylation in promoter of SEMA3B gene might be one of the mechanisms accounting for the reduced expression of SEMA3B,and being involved in tumorigenesis and progression of gastric cancer. SEMA3B might be a potential biomarker for diagnosis and prognostic assessment for gastric cancer and a promising epigenetic therapeutic target.