Downregulation of KLF5 by EBER1 via the ERK signaling pathway in EBV-positive nasopharyngeal carcinoma cells: implications for latent EBV infection.

Nasopharyngeal carcinoma (NPC) carcinogenesis and malignant transformation are intimately associated with Epstein-Barr virus (EBV) infection. A zinc-fingered transcription factor known as Krüppel-like factor 5 (KLF5) has been shown to be aberrantly expressed in a number of cancer types. However, little is known about the regulatory pathways and roles of KLF5 in EBV-positive NPC. Our study found that KLF5 expression was significantly lower in EBV-positive NPC than in EBV-negative NPC. Further investigation revealed that EBER1, which is encoded by EBV, down-regulates KLF5 via the extracellular signal-regulated kinase (ERK) signalling pathway. This down-regulation of KLF5 by EBER1 contributes to maintaining latent EBV infection in NPC. Furthermore, we uncovered the biological roles of KLF5 in NPC cells. Specifically, KLF5 may influence the cell cycle, prevent apoptosis, and encourage cell migration and proliferation - all of which have a generally pro-cancer impact. In conclusion, these findings offer novel strategies for EBV-positive NPC patients' antitumour treatment.

Epstein-Barr Virus and Helicobacter pylori co-infection in patients with gastric cancer and duodenale ulcer.

We aimed to detect EBV/Hp (Epstein-Barr Virus/Helicobacter pylori) co-infection by determining the number of copies of EBV/EBER-1 in the gastric biopsy samples of the Hp (+) GC, peptic ulcer (PU), and non-ulcer dyspepsia (NUD) cases. The patient group (PG), with 34 patients (34 GC and 30 PU patients) and a control group with 40 NUD cases were included. All patients and controls were Hp positive. EBV/EBNA-1 IgG were measured by the Anti-EBNA-1 ELISA IgG kit. Determination and quantification of EBV/EBER-1 gene region was performed by qPCR. EBV/EBER-1 positivity was 35.29% (12/34), 6.6% (2/30) and 2.5% (1/40) in GC, PU and 40 NUD cases, respectively. A significant difference was found between the GC and NUD cases (p=0.001). A significant difference was found between the groups for mean EBV/EBER-1 copy numbers (p=0.019). No significant difference was found between GC and the NUD cases (p=0.1455) for EBV/EBNA-1 IgG antibody positivity. EBV/EBER-1 positivity (OR=3.319), and age ≥55 years old (OR=2.331) were found to be a significant in multivariate logistic regression. In conclusion, our data suggest that the GC risk by EBVand Hp co-infection increased 3.3 times.

EBV-encoded RNA1-positive cells in the bone marrow specimens of patients with EBV-negative lymphomas and sarcomas.

Epstein-Barr virus (EBV) infection is associated with pathogenesis of various cancers, including extranodal natural killer/T-cell lymphoma, nasal type (ENKL). ENKL tumor cells are positive for EBV-encoded RNA1 (EBER1), which is the most useful marker to identify ENKL tumor cells in histopathology. Currently, EBER1 in situ hybridization (ISH) is recommended to evaluate bone marrow (BM) involvement of ENKL. However, the actual burden of EBER1-positive cells in normal BM specimens remains unclear. In the present study, we performed EBER1 ISH on 111 BM specimens, which were obtained during an initial staging procedure in patients with EBV-negative cancers and were also negative for BM involvement. One or more EBER1-positive cells per whole specimen were observed in 38 specimens (34%). The number of EBER1-positive cells was distributed as follows: single positive cell, n = 17; two positive cells, n = 13; three positive cells, n = 3; and four positive cells, n = 5. These findings suggest that four or fewer EBER1-positive cells can be observed in BM specimens of patients with non-EBV-related cancers. The clinical implications of a small number of EBER1-positive cells in BM specimens of patients with ENKL should be evaluated in further studies.

Sensing of latent EBV infection through exosomal transfer of 5'pppRNA.

Complex interactions between DNA herpesviruses and host factors determine the establishment of a life-long asymptomatic latent infection. The lymphotropic Epstein-Barr virus (EBV) seems to avoid recognition by innate sensors despite massive transcription of immunostimulatory small RNAs (EBV-EBERs). Here we demonstrate that in latently infected B cells, EBER1 transcripts interact with the lupus antigen (La) ribonucleoprotein, avoiding cytoplasmic RNA sensors. However, in coculture experiments we observed that latent-infected cells trigger antiviral immunity in dendritic cells (DCs) through selective release and transfer of RNA via exosomes. In ex vivo tonsillar cultures, we observed that EBER1-loaded exosomes are preferentially captured and internalized by human plasmacytoid DCs (pDCs) that express the TIM1 phosphatidylserine receptor, a known viral- and exosomal target. Using an EBER-deficient EBV strain, enzymatic removal of 5'ppp, in vitro transcripts, and coculture experiments, we established that 5'pppEBER1 transfer via exosomes drives antiviral immunity in nonpermissive DCs. Lupus erythematosus patients suffer from elevated EBV load and activated antiviral immunity, in particular in skin lesions that are infiltrated with pDCs. We detected high levels of EBER1 RNA in such skin lesions, as well as EBV-microRNAs, but no intact EBV-DNA, linking non-cell-autonomous EBER1 presence with skin inflammation in predisposed individuals. Collectively, our studies indicate that virus-modified exosomes have a physiological role in the host-pathogen stand-off and may promote inflammatory disease.

Single-center "Argentine" analysis of post-transplant lymphoproliferative disorders: incidence, histopathological characteristics and EBV status.

INTRODUCTION: Post-transplant lymphoproliferative disorders (PTLDs) are a heterogeneous group of lymphoid proliferations occurring after solid organ or bone marrow transplantation. The primary aims of our study were to characterize cumulative incidence of PTLDs, clinical and pathological features according to the Epstein-Barr virus (EBV) status and survival. METHODS: This was a retrospective cohort study on adult and pediatric patients, from January 2001 to December 2017. The cumulative incidence of PTLD was calculated by analyzing all the patients transplanted at our hospital, based on the database of the Organ Donation and Ablation Authority of Argentina (INCUCAI). The Kaplan-Meier method was used to plot the survival. RESULTS: Fifty-eight cases of biopsy-confirmed PTLD were identified and 12 cases of clinical data were incomplete and these patients were excluded. The median age at the time of the PTLD diagnosis was 17.5 years (interquartile range [IQR] 9 - 57). The median interval between transplant and PTLD diagnosis was 39 months (IQR 9 - 113). The most commonly transplanted organ was the liver (24 cases, 52.2%), followed by kidney (20 cases, 43.5%). The Epstein-Barr encoding region in situ hybridization (EBER ISH) was positive in 29 (69.8%) of the 43 evaluable biopsies. The PTLD cumulative incidence was 1.84% (95%CI 1.77 - 1.91) for solid organ and 0.84% (95%CI 0.48 - 1.2) for bone marrow transplant patients. The overall survival rate at 5 years was 0.77 (95%CI 0.61 - 0.87). Subgroups by the EBV EBER status, transplant type, PTLD subtype and age group (adult vs. pediatric) showed no statistically significant association with the overall survival. CONCLUSION: The PTLD incidence was similar to that of previous series and the EBER did not appear as a relevant factor in our patient survival.

The Impact of Deleting Stem-Loop 1 of Epstein-Barr Virus-Encoded RNA 1 on Cell Proliferation.

Epstein-Barr virus-encoded RNAs (EBERs) are two small, noncoding, structurally conserved transcripts, constitutively expressed at >106 copies per EBV-infected cell. They have been shown to drive cell growth. However, the mechanism(s) involved in EBER-induced proliferation is not clear. In this study, we investigated the molecular mechanisms and structural impact of EBER1. Sequences of EBER1 stem-loops (SL) 1, 3, and 4 were deleted, creating three mutants: ∆SL1, ∆SL3, and ∆SL4. These mutants were cloned into pHebo plasmids and expressed in Jurkat cell lines. Cells transfected with wildtype EBER1 and pHebo were used as controls. Cell proliferation was monitored by microscopy and flow cytometry. Microarray, qPCR, and Western blotting were used to investigate the cell cycle markers. We found significantly higher cell proliferation in wildtype EBER1 cells compared to pHebo, ∆SL1, and ∆SL3, but not ∆SL4 mutants. There was also significant upregulation of S-phase and G2/M phase markers in wildtype EBER1 and ∆SL4 mutant. Furthermore, CDT1, a factor for DNA replication, was upregulated in wildtype EBER1 and ∆SL4 mutant. However, in ∆SL1 mutant, CDT1 was significantly downregulated and translocated to the cytoplasm. These data indicate that the structure of EBER1 is important in cell proliferation.

Epstein-Barr virus noncoding small RNA (EBER1) induces cell proliferation by up-regulating cellular mitochondrial activity and calcium influx.

Epstein-Barr virus encoded RNAs (EBER1 and EBER2) are two non-polyadenylated, non-protein coding small RNAs expressed at high levels in all forms of EBV latent infections. Although not directly involved in cell transformation, a number of studies have reported that these RNAs may be involved in cell proliferation. However, which of the two EBERs play a major role in this process and the mechanisms involved remains unknown. The aim of this study was to investigate the role and mechanism of EBER1-induced cell proliferation. Using stably transfected EBER1 cell lines, and multiple methodologies, we show that EBER1 transfected epithelial, B and T cell lines proliferate at a higher rate, have higher metabolic activity and increased DNA synthesis. The mitochondrial number and activity was also observed to be higher in the EBER1 transfected cells. Moreover, cytochrome c activity and store operated calcium entry (SOCE) were potentiated in the EBER1 expressing cells. Finally, the genes associated with cell proliferation were also observed to be up-regulated in the EBER1 transfected cells. Taken together, our data has unravelled the role of mitochondria and cellular calcium pathway that appear to be involved in EBER1 induced cell proliferation of EBV infected cells.

EBV downregulates the m6A "writer" WTAP in EBV-associated gastric carcinoma.

Epstein-Barr virus-associated gastric carcinoma (EBVaGC) is characterized by the clonal growth of EBV-infected stomach epithelial cells. It has been reported that N6-methyladenosine (m6A) methylation can regulate the splicing, expression, decay and translation of mRNAs. Wilms' tumor 1-associating protein (WTAP) is an m6A "writer" with methyltransferase activity. An m6A RNA methylation quantification kit and immunofluorescence (IF) showed that the m6A total RNA methylation level of the Epstein-Barr virus-negative gastric carcinoma (EBVnGC) cell line (SGC7901) was higher than that in the EBVaGC cell line (GT38). To investigate the underlying mechanism of the downregulated expression of m6A RNA methylation, we analyzed the expression of WTAP. The results showed that the expression of WTAP protein in EBVaGC cell lines was significantly lower than that in EBVnGC cell lines according to western blotting and IF. Through plasmid overexpression and RNA interference technology, we further found that EBV-encoded small RNA1 (EBER1) could downregulate WTAP expression by activating the NF-κB signaling pathway. In addition, WTAP could increase proliferation and inhibit migration in gastric carcinoma cell lines. In summary, EBER1 of EBV potentially regulated WTAP by affecting the NF-κB signaling pathway and WTAP further affected cell proliferation and migration.

Exosomes-carrying Epstein-Barr virus-encoded small RNA-1 induces indoleamine 2, 3-dioxygenase expression in tumor-infiltrating macrophages of oral squamous-cell carcinomas and suppresses T-cell activity by activating RIG-I/IL-6/TNF-α pathway.

OBJECTIVES: Although exosomes carrying Epstein-Barr virus-encoded small RNA-1 (EBER-1) are involved in the immunosuppressive tumor microenvironments of EBV-associated head and neck carcinomas, the effects of EBER-1-associated exosomes on tumor-infiltrating macrophages are poorly understood. MATERIALS AND METHODS: The association between EBV infection and expression of indoleamine 2,3-dioxygenase (IDO) was assessed in 165 paraffin-embedded oral squamous cell carcinoma (OSCC) tissue samples. Using in vitro techniques, we investigated whether stimulation of the RIG-I/IL-6/TNF-α pathway by exosomes carrying EBER-1 is critical for IDO induction in macrophages. We performed a thymidine incorporation and a cell cytolytic assay to test for up-regulated IDO in macrophages that can block the proliferation and function of effector T cells. RESULTS: Some infiltrated macrophages expressed levels of IDO higher than OSCC cells which was significantly associated with presence of EBV. The production of IDO, tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) in human monocyte-derived macrophages (MDMs) was induced by EBV-associated exosomes in vitro. Mechanistically, the retinoic acid-inducible gene I (RIG-I) pathway in MDMs was stimulated by EBV-encoded small RNA-1 (EBER-1) whereas the inhibition of these pathways by BX-795 almost abolished the production of these two cytokines and IDO induction. Also, the EBER-1-activated IDO in MDMs suppressed the proliferation of T lymphocytes and diminished the cytolytic activity of CD8+ T cells. CONCLUSION: Exosomes carrying EBER-1 could induce IDO expression in MDMs, considerably aided by an IL-6 and TNF-α-dependent mechanism via the RIG-I signaling pathway, which might create an immunosuppressive microenvironment affecting T-cell immune responses.

Extranodal NK/T-Cell Lymphoma, Nasal Type: Genetic, Biologic, and Clinical Aspects with a Central Focus on Epstein-Barr Virus Relation.

Extranodal NK/T-Cell Lymphoma, nasal type (ENKTL-NT) has some salient aspects. The lymphoma is commonly seen in Eastern Asia, has progressive necrotic lesions in the nasal cavity, makes midfacial destructive lesions, and shows poor prognosis. The lymphoma cell is originated from either NK- or γδ T-cells, which express CD56. Since the authors first demonstrated the existence of Epstein-Barr virus (EBV) DNA and EBV oncogenic proteins in lymphoma cells, ENKTL-NT has been recognized as an EBV-associated malignancy. Because the angiocentric and polymorphous lymphoma cells are mixed with inflammatory cells on a necrotic background, the diagnosis of ENKTL-NT requires CD56 immunostaining and EBER in situ hybridization. In addition, serum the EBV DNA level is useful for the diagnosis and monitoring of ENKTL-NT. Although ENKTL-NT is refractory lymphoma, the prognosis is improved by the development of therapies such as concomitant chemoradiotherapy. The basic research reveals that a wide variety of intracellular/cell surface molecules, cytokines, chemokines, and micro RNAs are involved in lymphomagenesis, and some of them are related to EBV. Understanding lymphoma behavior introduces new therapeutic strategies, such as the usage of immune checkpoint inhibitors, peptide vaccines, and molecular targeting therapy. This review addresses recent advances in basic and clinical aspects of ENKTL-NT, especially its relation to EBV features.

Clinical features of Epstein-Barr virus in the intestinal mucosa and blood of patients with inflammatory bowel disease.

BACKGROUND: The role of Epstein-Barr virus (EBV) in inflammatory bowel disease (IBD) remains to be elucidated. The aim of this study was to investigate the presence of EBV in the blood and intestinal mucosa of patients with IBD and evaluate the association between EBV positivity and IBD. METHODS: Patients with IBD, hospitalized between January 2015 and April 2018, were enrolled. The EBV-DNA load in blood samples from each subject was analyzed by quantitative real-time polymerase chain reaction. EBV-encoded small-RNA 1 (EBER-1) was detected by in-situ hybridization in intestinal mucosa tissue sections of patients with IBD. RESULT: EBV-DNA was detected in 48 out of 568 patients with IBD (8.4%), and EBER-1 positivity was detected in 27 of these patients (56.3%). Refractory IBD and severe mucosal inflammation were more common in patients with detectable levels of EBER-1 than in those without; the number of EBER-1-positive cells positively correlated with mucosal inflammation (P value < 0.05). Age (≥60 years old) and use of azathioprine were risk factors for EBV infection. There was no significant difference in clinical remission rate and surgical rate between the EBER-1 positive group and EBER-1 negative group, antiviral group and the non-antiviral group, among IBD patients who tested positive for EBV-DNA. CONCLUSION: Elderly patients with IBD, treated with azathioprine, are more susceptible to EBV positivity. Further, EBV mucosal detection correlated with the severity of mucosal damage and refractoriness, but not prognosis.

Association of Plasma Epstein-Barr Virus LMP1 and EBER1 with Circulating Tumor Cells and the Metastasis of Nasopharyngeal Carcinoma.

Epstein-Barr virus (EBV) has been widely recognized to contribute to the development of nasopharyngeal carcinoma (NPC). The present study was to explore the association of plasma Epstein-Barr Virus LMP1 and EBER1 with circulating tumor cells (CTCs) and the metastasis of nasopharyngeal carcinoma. In the present study, we quantified the plasma levels of EBV DNA/RNAs, such as LMP1, LMP2, BART and EBER1 with real-time quantitative PCR, and CTCs with a CellSpotter Analyzer in NPC patients, with or without metastasis. Then the correlation of each biomarker with other biomarkers and tumor metastasis was analyzed. Our data indicated that the plasma levels of EBV LMP1, BART, EBER1, along with CTCs were significantly higher in metastatic NPC patients than in non-metastatic patients. Plasma LMP1 DNA and EBER1 discriminate metastatic NPC patients from non-metastatic patients, correlate with tumor stage and node stage for metastatic NPC patients. In summary, there were significantly higher plasma levels of Epstein-Barr Virus DNAs / RNAs in nasopharyngeal carcinoma patients. LMP1 DNA and EBER1 RNA correlated with the metastasis of nasopharyngeal carcinoma.

Epstein barr virus IgG and EBER-1 in Burkitt's lymphoma children at a referral hospital in western Kenya.

Burkitt's lymphoma (BL) is a frequent childhood B cell non-Hodgkin's lymphoma (NHL) in equatorial Africa associated with infections. Chronic Epstein Barr virus (EBV) infections can lead to host immune stimulation that may trigger genetic translocation(s), neoplastic transformation and proliferation of B cells. We determined EBV immunoglobulin G (IgG) in sera from participants and EBER-1 in tumour sections in confirmed BL cases at Moi Teaching and Referral Hospital (MTRH). A cross sectional study of children with clinical and histology diagnosis of NHL from whom BL status were confirmed by immunohistochemistry (IHC) was carried out. Epstein Barr virus IgG in sera was determine using Enzyme-linked immunosorbant assay, IHC for EBER-1 and MYC protein in tumour sections. Demographic and clinical information were obtained from questionnaires and hospital files respectively. Ninety three percent of sera were EBV IgG positive of which 31.7% were confirmed as BL. All jaw BL tumours and 86.7% of BL tumours carried EBER-1 antigen. Odds ratio of EBER-1 positive was 1.39, 95% CI: 0.16-12.19 in BL tumours regardless of age or gender. EBV infection among the study participants may be associated with BL, however, EBER-1 and MYC negative in BL tumours suggest alternative BL pathogenesis or variant.

Single-center "Argentine" analysis of post-transplant lymphoproliferative disorders: incidence, histopathological characteristics and EBV status

ABSTRACT Introduction: Post-transplant lymphoproliferative disorders (PTLDs) are a heterogeneous group of lymphoid proliferations occurring after solid organ or bone marrow transplantation. The primary aims of our study were to characterize cumulative incidence of PTLDs, clinical and pathological features according to the Epstein-Barr virus (EBV) status and survival. Methods: This was a retrospective cohort study on adult and pediatric patients, from January 2001 to December 2017. The cumulative incidence of PTLD was calculated by analyzing all the patients transplanted at our hospital, based on the database of the Organ Donation and Ablation Authority of Argentina (INCUCAI). The Kaplan-Meier method was used to plot the survival. Results: Fifty-eight cases of biopsy-confirmed PTLD were identified and 12 cases of clinical data were incomplete and these patients were excluded. The median age at the time of the PTLD diagnosis was 17.5 years (interquartile range [IQR] 9 - 57). The median interval between transplant and PTLD diagnosis was 39 months (IQR 9 - 113). The most commonly transplanted organ was the liver (24 cases, 52.2%), followed by kidney (20 cases, 43.5%). The Epstein-Barr encoding region in situ hybridization (EBER ISH) was positive in 29 (69.8%) of the 43 evaluable biopsies. The PTLD cumulative incidence was 1.84% (95%CI 1.77 - 1.91) for solid organ and 0.84% (95%CI 0.48 - 1.2) for bone marrow transplant patients. The overall survival rate at 5 years was 0.77 (95%CI 0.61 - 0.87). Subgroups by the EBV EBER status, transplant type, PTLD subtype and age group (adult vs. pediatric) showed no statistically significant association with the overall survival. Conclusion: The PTLD incidence was similar to that of previous series and the EBER did not appear as a relevant factor in our patient survival.

Down-regulation of increased TRAF6 expression in the peripheral mononuclear cells of patients with primary Sjögren's syndrome by an EBV-EBER1-specific synthetic single-stranded complementary DNA molecule.

AIM: We described earlier a simultaneously increased that the increased expression of miRNA-146a/b was accompanied by an increase in the expression of and TRAF6 and a decrease in the expression of IRAK1 genes in the peripheral mononuclear cells (PBMCs) of patients with primary Sjogren's syndrome (pSS) patients. Recently, the expression of EBV encoded. RNA (EBER) was published in the B cells of salivary glands of in pSS. In the present study, we applied an EBV-EBER1 specific synthetic single stranded complementary DNA molecule (EBV-EBER1-cDNA) to test whether any EBER1 related effect exists also in PBMCs of pSS patients. METHODS: In the PBMCs of pSS patients and healthy controls, we investigated in vitro the effects of a synthetic single stranded EBV-EBER1-cDNA molecule, synthetic double-stranded (ds)RNA polyinosinic-polycytidylic acid [poly (I:C)] and polyadenylic acid potassium salt poly-adenylic acid [poly-(A)] on the expression of TRAF6 gene tested by qRTPCR. The release of interferon -α was detected by ELISA. RESULTS: EBV-EBER1-cDNA resulted in a significant reduction in the expression of TRAF6 in the cells of patients, but in the healthy controls not, whereas the treatments with poly (I:C) and poly-(A) could not reduce the TRAF6 over-expression. No release of EBER1 could be observed in the culture supernatants of patients with pSS. Only the treatment with poly (I:C) resulted in a significant increase of interferon -α release, and only in the heathy controls. No release of EBER1 molecules took place during the culturing of cells. EBV-EBER- cDNA acted functionally on the cells of patients only. CONCLUSION: These findings give a further evidence of the linkage between EBV and pSS, furthermore, they show the possible role of EBV-EBER1 in the induction of increased TRAF6 expression in the peripheral B cells of Sjögren's patients.

Epstein-Barr virus-encoded small RNA 1 (EBER-1) could predict good prognosis in nasopharyngeal carcinoma.

PURPOSE: EBER-1 (a non-coding RNA transcribed by EBV) expression was detected in most of Epstein-Barr virus (EBV)-positive nasopharyngeal carcinoma (NPC) patients. However, the relevance between EBER-1 expression and NPC clinical outcome has not been reported. This study aims to assess the possible correlations of EBER-1 expression and clinical parameters and its potential prognostic predictive ability in NPC patient's outcomes. METHODS: We examined EBER-1 mRNA expression in 301 NPC and 130 non-NPC tissues using in situ hybridization and did statistics. RESULTS: EBER-1 expression was up-regulated in NPC tissues when compared to non-NPC tissues. A receiver operating characteristic analysis revealed that EBER-1 expression could distinguish non-cancerous patients from NPC patients (p < 0.001, sensitivity: 72.5 %, specificity: 83.5 %, AUC = 0.815). A survival analysis revealed that patients with high levels of EBER-1 expression had a significantly good prognosis (Disease-free survival: p = 0.019, overall survival: p = 0.006). CONCLUSION: These results indicated that EBER-1 expression is a potential prognosis factor of NPC and highly negative correlated with the progress of NPC.

Application of circulating plasma/serum EBV DNA in the clinical management of nasopharyngeal carcinoma.

Elevated levels of circulating cell-free Epstein-Barr virus (EBV) DNA have been detected in plasma and serum samples from nasopharyngeal cancer (NPC) patients by quantitative real time PCR (qPCR) test. This qPCR test for circulating EBV DNA was found to be useful in the clinical management of NPC patients. For instance, EBV DNA qPCR test has good sensitivity and specificity in the detection of NPC at disease onset. Increase of the viral DNA load was found in NPC patients at late stages of disease. High EBV DNA load at disease onset or detectable viral load post-treatment was associated with poor survival or frequent relapse in NPC patients. Residual EBV DNA load after primary treatment could be a useful indicator to justify adjuvant chemotherapy. The qPCR test might also be applied to define a poor prognostic group in patients at early stage (I/II) for implementing concurrent chemo-radiotherapy (chemo-RT) to improve patients' outcome. The test is also useful to monitor distant metastases or response to radiotherapy, chemo-RT or surgery. Supplementary tests, however, are needed to pick up EBV negative WHO type I NPC and test improvement is needed to increase sensitivity in detecting stage I disease and local recurrence.

EBV-encoded small RNA1 and nonresolving inflammation in rheumatoid arthritis.

Rheumatoid arthritis (RA) is a chronic autoimmune disease characterized by perpetuated inflammation in multiple joints. To date, there is no cure for RA, and the causal factor for non-resolving inflammation in RA remains unclear. In this study, we initially observed expression of Epstein-Barr virus-encoded small RNA1 (EBER1) in the synovial tissue of all five patients who showed nonresolving RA inflammation. By contrast, EBER1 was detected in the synovial tissue of only one out of seven patients with advanced osteoarthritis (OA; p < 0.01, Fisher's exact test). To confirm this finding, we conducted a second study on synovial tissue samples taken from 23 patients with nonresolving RA inflammation and 13 patients with OA. All synovial samples from patients with nonresolving inflammation of RA showed positive expression of EBER1 (23/23, 100%), whereas none of the synovial samples from patients with OA showed expression of EBER1 (0/13, 0%; p < 0.001, by Fisher's exact test). In vitro, transfection of RA synovial fibroblasts with EBER1 induced the production of interleukin-6. Taken together, these data strongly suggest that nonresolving RA inflammation is strongly related to the presence of EBER1, which might be, at least partially, responsible for synovial fibroblast interleukin-6 production.

Epstein-Barr virus and Hodgkin's lymphoma in Cairo, Egypt.

Fifty-five consecutive cases of Hodgkin's lymphoma (HL), collected between 1996 and 1998 from Cairo, Egypt, were histologically subtyped, phenotyped, and then studied for the presence of Epstein-Barr virus (EBV). We used immunohistochemical stains for EBV latent membrane protein 1 (LMP-1) and in situ hybridization stains for EBV-encoded small RNA (EBER-1) transcripts. Forty-five cases (82%) had classic HL (cHL), and ten cases (18%) had nodular lymphocyte predominant HL (NLPHL), with each group expressing its typical phenotype. LMP-1 stains were positive in 63% and 0% of cHL and NLPHL cases, respectively. EBER-positive Reed-Sternberg cells and variants were also present in 62% and 0% of each group, respectively. The cHL cases showed variable EBER positivity: nodular sclerosis, 58%; mixed cellularity, 100%; lymphocyte depletion, 100%; and unclassifiable, 67%. Our findings are similar to those from other developing countries and point towards a pathogenic role of EBV in cHL.