DNA Replication ; physiology ; Epstein-Barr Virus Infections ; virology ; Herpesvirus 4, Human ; genetics ; metabolism ; physiology ; Humans

OBJECTIVETo study the genetic stability of an immortalized cell line transformed by Epstein-Barr virus (EBV) after long subculture process.

METHODSIn the present study, the genetic stability including chromosome diploidy, karyotypes and microsatellite DNA were evaluated with chromosome banding techniques and microsatellite DNA detection. The telomerase activity of the immortalized cell line was detected by using the telomerase assay kit.

RESULTSFrom passage 1 to 30, there were no change of the diploidy, karyotypes of chromosome and microsatellite DNA, and the telomerase activity is negative.

CONCLUSIONThis study indicates that the immortalized cell line remains stable genetically within limited passages.

Cell Transformation, Viral ; genetics ; Herpesvirus 4, Human ; genetics ; Humans ; Lymphocytes ; cytology ; metabolism ; virology ; Microsatellite Repeats ; genetics ; Polymerase Chain Reaction

The Epstein-Barr virus (EBV) is a gamma herpes virus associated with several types of malignancies. The EBV encodes viral microRNAs (miRNAs) that can target genes within cells. The EBV participates in signal transduction as well as the proliferation and differentiation of cells. How the target genes and functions of EBV-encoded miRNAs contribute to the pathogenesis of EBV is an important research topic. Some target genes have been validated since EBV-encoded miRNAs were discovered and, in this article, we summarize them and their functions.
Animals ; Epstein-Barr Virus Infections ; genetics ; metabolism ; virology ; Herpesvirus 4, Human ; genetics ; physiology ; Humans ; MicroRNAs ; genetics ; metabolism ; RNA, Viral ; genetics ; metabolism

PURPOSE: The aim of the present study was to evaluate the clinical characteristics of the primary Epstein-Barr virus (EBV) hepatitis with elevation of both serum alkaline phosphatase (ALP) and gamma-glutamyltransferase (gamma-GT) levels in children. MATERIALS AND METHODS: A retrospective study was performed by reviewing of the medical records of 36 patients who were diagnosed with primary EBV hepatitis. The patients were divided into 2 groups: patients with elevated serum ALP and gamma-GT levels (group 1) and patients without (group 2). RESULTS: The classic features of infectious mononucleosis (fever, pharyngitis and/or tonsillitis, and cervical lymphadenitis) were seen in 20 (57.1%) of group 1 patients and 18 (50.0%) of group 2 patients. Hepatitis with elevated serum ALP and gamma-GT levels were present in 14 (38.9%) of the all patients. Of these patients, Jaundice occurred in only 2 (5.6%). The mean levels of aspartate aminotransferase and alanine aminotransferase (ALT) as well as the number of patients with ALT greater than 400 IU/L were significantly different between the groups (177 IU/L vs. 94 IU/L, 418 IU/L vs. 115 IU/L, and 50.0% vs. 13.6%; p=0.001, p=0.001, p=0.026, respectively). The mean duration of elevated serum ALT levels was 17.5 days in group 1 and 9.0 days in group 2 (p=0.013). All patients recovered fully without any chronic or serious complications. CONCLUSION: Primary EBV hepatitis with predominant biochemical abnormalities of the elevation of ALP and gamma-GT is frequent and mostly anicteric. This may represent a benign disease, but a delay in recovery of liver function as well.
Alkaline Phosphatase/genetics/*metabolism ; Child ; Child, Preschool ; Female ; Hepatitis/*enzymology/*pathology/virology ; Herpesvirus 4, Human/*pathogenicity ; Humans ; Infant ; Male ; gamma-Glutamyltransferase/genetics/*metabolism

Antigens, Viral ; genetics ; metabolism ; Cell Line ; Cell Phone ; Gene Expression Regulation, Viral ; radiation effects ; Herpesvirus 4, Human ; physiology ; radiation effects ; Humans ; Immunohistochemistry ; Viral Proteins ; genetics ; metabolism

OBJECTIVETo evaluate the application of RNA interference in the study of latent membrane protein-1 (LMP-1) in EB virus-positive nasopharyngeal carcinoma (NPC) cell line C611. Observe the effects of LMP-1 silencing on NPC cell growth.

METHODSFour synthesized small interference RNA (siRNA) were transfected into NPC cell using Oligofectamine reagent. LMP-1 mRNA level was determined by semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR). Flow cytometry and MTT assay were employed to analyze the effects on cell cycle and proliferation.

RESULTSThe most effective sequence found out among the 4 candidates. Single dose of this siRNA caused nearly 90% loss of LMP-1 mRNA in C611 cell. The specific inhibition could last for 96 hours if re-transfection was preformed. LMP-1 siRNA treatment resulted in cell cycle arrest at G0-G1 stage in C611, accompany with a reduction of cell proliferation by 32.9%. While EBV-negative NPC cells appeared unaffected.

CONCLUSIONSThese results provided solid testimony that EBV-encoded latent membrane protein-1 was vulnerable to RNA interference and, selective inhibition of LMP-1 had anti-proliferation effect on NPC cell. RNA interference could be a powerful tool in further investigations of LMP-1 and a novel therapeutic strategy for EBV-related NPC patients.

Cell Line, Tumor ; Cell Proliferation ; Herpesvirus 4, Human ; metabolism ; Humans ; Nasopharyngeal Neoplasms ; genetics ; pathology ; RNA Interference ; RNA, Messenger ; genetics ; RNA, Small Interfering ; Transfection ; Viral Matrix Proteins ; genetics

The oncogenic Epstein-Barr virus (EBV) -encoded latent membrane protein 1 (LMP1) enables this virus's long-term survival within the cells of immune system. Mean while, LMP1 also plays a critical role for the transformation of resting B cells by EBV. It initiates the activation of signalling pathways, such as NF-kappaB, mitogen-activated protein kinase (MAPK), and JAK/STAT cascade by adaptor proteins including the tumor necrosis factor (TNF) receptor associated factors (TRAFs) and the TNF receptor associated death domain protein (TRADD). It increases the expression of adhesion molecules LFA-1, ICAM-1, and costimulatory molecule B7-1 of B cells, and regulates the antibody and cytokine secreted by B cells. LMP1 and CD40 have many common properties in signal transduction. Both of them co-localize in lipid rafts for signal transduction. Considering its close relationship with CD40, the research on LMP1 has become a hot spot in the immunology field.
Animals ; B-Lymphocytes ; immunology ; CD40 Antigens ; genetics ; physiology ; Gene Expression Regulation, Viral ; Herpesvirus 4, Human ; genetics ; metabolism ; physiology ; Humans ; Signal Transduction ; Viral Matrix Proteins ; genetics ; physiology

Recent studies have suggested a probable association between Epstein-Barr virus (EBV) and nasal/nasopharyngeal T cell lymphomas but the role of oncogenes or tumor suppressor genes is poorly understood. We have studied the frequency of p53 expression and its relation to the EBV infection in 33 Korean patients with head and neck (H&N) lymphomas. All cases (23 B cell & 10 T cell) were immunostained for p53 protein using the mAb D07 (Novocastra) and the avidin biotin peroxidase method. EBER in situ hybridization was performed using a fluorescein conjugated EBV oligonucleotide probe (Dako). Among 33 lymphomas, 16 cases stained positively for p53 protein. P53 expression was frequent both in higher grade lymphomas and in advanced stage. Nine cases were EBER positive, EBER was more commonly found in T cell lymphomas than in B cell lymphomas (70% vs 8.7%). EBER positive lymphomas showed a higher frequency of p53 positivity than EBER negative lymphomas (78% vs 38%), although the difference was not statistically significant (p = 0.095). These findings indicate altered expression of p53 protein occurs in H&N lymphomas, especially in late event lymphoma progression and appears to play a role in the development of EBER positive T cell lymphomas.
Follow-Up Studies ; Gene Expression ; Genes, p53 ; Head and Neck Neoplasms/genetics/*metabolism/*virology ; Herpesviridae Infections/genetics/*metabolism ; *Herpesvirus 4, Human ; Human ; Lymphoma, T-Cell/genetics/*metabolism/*virology ; Prognosis ; Protein p53/*biosynthesis ; Support, Non-U.S. Gov't ; Tumor Virus Infections/genetics/*metabolism

Epstein-Barr virus (EBV) is a human herpesvirus associated with important human diseases, including infectious mononucleosis syndrome, malignant lymphoma, and nasopharyngeal carcinoma. The mechanism of EBV entry into host cells remains a subject of intensive research. After decades of study, researchers have identified several key proteins and different patterns of EBV intrusion into host cells. The viral surface glycoproteins, gp350/220, gp42, gB, gH, and gL, are involved in interactions with the CR2 receptor on the surface of B lymphocytes during viral entry. However, the majority of epithelial cells lack CR2 receptor expression, which makes viral invasion much more complex than in B lymphocytes. Three different models have been proposed to explain how EBV enters epithelial cells: (1) "transfer of infection", mediated by B lymphocytes or Langerhans cells; (2) EBV utilizes its own proteins during the process of fusion with the cell membrane; and (3) progeny virions arising from EBV-infected epithelial cells cross lateral membranes into adjacent epithelial cells. This review will discuss the relevant mechanism of viral entry into B lymphocytes and epithelial cells during EBV infection.
Animals ; B-Lymphocytes ; virology ; Epithelial Cells ; virology ; Epstein-Barr Virus Infections ; virology ; Herpesvirus 4, Human ; genetics ; physiology ; Humans ; Viral Proteins ; genetics ; metabolism ; Virus Internalization

BACKGROUND AND OBJECTIVEEBV BamHI fragment H rightward open reading frame 1 (BHRF1), the Epstein-Barr virus (EBV) early gene, is structurally and functionally homologous to the oncogene bcl-2 and may play an important role in the development of EBV-associated tumors. To characterize the polymorphisms of BHRF1 in EBV-associated tumors, we analyzed the sequences of BHRF1 in isolates from nasopharyngeal carcinoma (NPC) and EBV-associated gastric carcinoma (EBVaGC) biopsies as well as throat washing (TW) samples from healthy donors.

METHODSBHRF1 DNA sequences were analyzed by polymerase chain reaction (PCR) and sequencing for 39 NPC samples, 40 EBVaGC samples, and 53 EBV-positive TW samples from healthy donors. The variants of BHRF1 gene were classified according to the signature changes. The EBV types 1 and 2 at nuclear antigen (EBNA) 3C locus were determined by PCR.

RESULTSCompared with EBV standard cell line B95-8, all isolates carried a silent mutation at amino acid (AA) 80 (nucleotide 54616 T→C), the AA88 L→V mutation was found in most isolates, and the AA79 V→L mutation in a few isolates. Other mutations were sporadically distributed. Based on the mutations at AA88 and AA79, 3 distinct variants of BHRF1 genes, designated as 79V88V, 79L88L, and 79V88L, were identified. The 79V88V was the most common variant. The distribution of the BHRF1 variants among the NPC, EBVaGC, and TW samples was not significant. The corresponding regions of bcl-2 homologues were conserved in all isolates except for 3 samples. The distribution of BHRF1 variants in type 1 and type 2 strains was significant different (P < 0.001, contingency coefficient was 0.554).

CONCLUSIONSThe 79V88V is the dominant variant in NPC, EBVaGC, and TW samples from healthy donors and preferential linkages between BHRF1 and EBNA3C variants exist. Conserved BHRF1 in Bcl-2 homologous domains is helpful to remain the important role of BHRF1.

Carcinoma ; Herpesvirus 4, Human ; genetics ; Humans ; Nasopharyngeal Neoplasms ; genetics ; metabolism ; virology ; Polymorphism, Genetic ; Proto-Oncogene Proteins c-bcl-2 ; genetics ; metabolism ; Sequence Analysis, DNA ; Stomach Neoplasms ; genetics ; metabolism ; virology ; Viral Proteins ; genetics ; metabolism