[Inhibiting target gene expression and controlling growth of Epstein-Barr virus transformed cells by antisense RNA transcripts].

BACKGROUND & OBJECTIVES: The latent membrane protein gene (LMP) of Epstein-Barr virus (EBV) was thought to play an important role in the carcinogenesis of nasopharyngeal carcinoma (NPC). In this study, the authors investigated the effects of antisense RNA (AsRNA) on LMP for down regulating at the target gene over expression in EBV transformed lymphoid cells, and set up an antisense system to inhibit LMP expression. METHODS: Constructing the single strand antisense transcription system in vitro, the authors have got large amount of AsRNA. Designing and setting up an antisense tracing system in situ (ATSIS), the authors could observe the living particles of AsRNA/sense RNA duplex dimer. With time lapse phase-contrast microscopy, the agglutination phenotype on living cells was easily detected by MTT test, the inhibition rate on EBV transformed cells was calculated. RESULTS: LMP 1.9 fragment ligated into pGEM vector in reverse orientation have been constructed and produced a plentiful amount of AsLMPmRNA which could incorporated into both B95-8 and C1936 cell lines by endophagocytosis and formed the duplex dimer of As/Sense RNA. This particles have been visualized in situ when labelling 35S isotope by ATSIS. When AsLMPmRNA acted as agents for specific inhibition to LMP over expression, the transform phenotype of cell agglutination have been suppressed and MTT particle formatin was apparently reduced both two EBV tansformed cell lines. CONCLUSION: AsLMPmRNA targets at sense strand have a high effectiveness of down-regulation on EBV-LMP overexpression. This down regulating function of LMP and growth inhibition on transformed cell is demonstrated by the antisenes tracing system in situ (ATSIS). The results provide a clue to overcome the latent EBV infection in human bodies all living long time and to prevent it inducing NPC in high incidence area by antisense strategies.

The 30-bp deletion variant: a polymorphism of latent membrane protein 1 prevalent in endemic and non-endemic areas of nasopharyngeal carcinomas in China.

Development of nasopharyngeal carcinoma (NPC) is closely associated with Epstein-Barr virus (EBV) infection. However, NPC occurs with a marked geographic and racial distribution, whereas EBV infection is ubiquitous in the world. This leads to a question whether certain subtypes of EBV have a greater potential to induce cell transformation. Latent membrane protein 1 (LMP1) is an EBV-encoded oncogenic protein and its 30-bp deleted variant (del-LMP1) has been reported to be predominant in biopsies of NPC. We have assessed the polymorphism of LMP1 in 47 biopsies of NPC, 107 cases of throat washings (TWs) from NPC patients, and 106 cases of TWs from non-NPC patients in Guangzhou, an endemic area of NPC in southern China, as well as 103 cases of TWs from healthy donors in Haerbin, a non-endemic area of NPC in northern China. Our results found a similar extent of the LMP1 polymorphism between NPC patients and non-NPC patients in Guangzhou, with the del-LMP1 being predominant in both Guangzhou and Haerbin. Sequence analyses showed identical substitutions in other coding regions of the del-LMP1 isolated from Guangzhou and Haerbin. These results indicate that del-LMP1 represents a geographic or race-associated polymorphism rather than an NPC disease phenotype-associated polymorphism.