Relationship between Epstein-Barr virus-encoded proteins with cell proliferation, apoptosis, and apoptosis-related proteins in gastric carcinoma

Abstract

Aim: To investigate the interrelationship between Epstein-Barr virus (EBV)-encoded proteins and cell proliferation, apoptosis and apoptosis-related proteins in gastric carcinoma, and to explore their role in gastric carcinogenesis.

Methods: Tissues from 13 cases of EBV-associated gastric carcinoma (EBVaGC) and 45 cases of matched EBV-negative gastric carcinoma (EBVnGC) were collected, and then subjected to analysis for apoptotic index (AI) using the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick end-labeling (TUNEL) assay. Nuclear cell proliferation-associated antigen ki-67 index (KI), bcl-2, and p53 expression were examined by immunohistochemistry. p53 mutation in exons 5-8 of 13 EBVaGC cases was determined by single-strand conformation polymorphism (SSCP) and DNA sequencing. RT-PCR and Southern hybridization were used to detect the expression of nuclear antigens (EBNAs) 1 and 2, latent membrane protein (LMP) 1, immediately early gene BZLF1 and early genes BARF1 and BHRF1 in 13 EBVaGC cases.

Results: The percentage of AI, KI and p53 overexpression was significantly lower in the EBVaGC group than in the EBVnGC group. However, bcl-2 expression did not show significant difference between the two groups. p53 gene mutations were not found in 13 EBVaGCs. Transcripts of EBNA1 were detected in all 13 EBVaGCs, while both EBNA2 and LMP1 mRNA were not detected. Six of the thirteen cases exhibited BZLF1 transcripts and two exhibited BHRF1 transcripts. BARF1 mRNA was detected in six cases.

Conclusion: Lower AI and KI may reflect a low biological activity in EBVaGC. EBV infection is associated with p53 abnormal expression but not bcl-2 protein in EBVaGC. BZLF1, BARF1, and BHRF1 may play important roles in inhibiting cell apoptosis and tumorigenesis of EBVaGC through different pathways.

Figure 1

Detection of EBV-associated gene expression by RT-PCR and Southern hybridization in EBVaGC. M: DIG-labeled DNA molecular weight marker VIII (Roche); lane 1: EBV-positive LCL (positive control); lane 2: EBV-negative Ramos cells (negative control); lanes 3-13: EBV-positive gastric carcinoma samples.

Figure 2

TUNEL for Cell apoptosis and immunohistochemistry of p53, bcl-2, and ki-67. A: Apoptotic cells; B: Expression of p53; C: Expression of bcl-2; D: Expression of ki-67. (Original magnification ×400).