Methylation and Expression of Retinoblastoma and Transforming Growth Factor-β1 Genes in Epstein-Barr Virus-Associated and -Negative Gastric Carcinomas

Abstract

Background. Retinoblastoma (RB) and transforming growth factor-β1 (TGF-β1) are important tumor-related factors. Methods. A series of 30 EBV-associated gastric carcinoma (EBVaGC) and 38 matched EBV-negative gastric carcinoma (EBVnGC) tissues were examined for the promoter methylation of RB by methylation-specific PCR (MSP) method. The expression of RB and TGF-β1 in gastric carcinoma tissues was detected by immunohistochemistry. Results. The methylation rate of RB gene in EBVaGC and EBVnGC was 80.0% (24/30) and 50.0% (19/38), respectively. The difference of RB methylation rate between EBVaGC and EBVnGC was significant (χ(2) = 6.490, P = 0.011). There was no significant difference for RB expression between EBVaGC (43.3%, 13/30) and EBVnGC (63.2%, 24/38), and also for TGF-β1 between EBVaGC (56.7%, 17/30) and EBVnGC (63.2%, 24/38). RB methylation was not reversely correlated with RB expression in gastric carcinoma tissues (χ(2) = 2.943, P = 0.086, r = 0.208). RB methylation, loss expression of RB, and TGF-β1 expression were significantly associated with tumor invasion and lymph node metastasis (P < 0.05), but was not associated with sex, age, histological subtype (differentiation status) and tumor location. Conclusions. Methylation of RB is a common event in gastric carcinomas and EBV induces methylation of RB in EBVaGC, which may contribute to the development of gastric carcinomas. EBV has no significant effect on induction of TGF-β1 expression. Detection of RB methylation, RB expression, and TGF-β1 expression may be helpful to judge the status of tumor invasion and lymph node metastasis in gastric carcinomas.

Figure 1

RB promoter methylation in EBVaGC, EBVnGC, and matched paracarcinoma tissues. (a) Representative RB promoter methylation in EBVaGC and EBVnGC by MSP. U, PCR product from the MSP assay using primers specific for the unmethylated allele; M, PCR product from the MSP assay using primers specific for the methylated allele. (b) Representative RB MSP assay results for matched paracarcinoma tissues.

Figure 2

The protein expression of RB and TGF-β1 by immunohistochemistry (magnification ×100). (a) Positive immunohistochemistry result of RB in paraffin section. Expression of RB was found in nuclei of gastric carcinoma cells. (b) Positive immunohistochemistry result of TGF-β1 in paraffin section. Expression of TGF-β1 was found in cytoplasm of gastric carcinoma cells. (c) Negative immunohistochemistry result of RB in paraffin section. (d) Negative immunohistochemistry result of TGF-β1 in paraffin section.