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M Rowe

Publications and source records attributed to M Rowe.

At least 109 records · Page 6Linked to original sources
PubMed1992

Immunohistochemical demonstration of the Epstein-Barr virus-encoded latent membrane protein in paraffin sections of Hodgkin's disease.

Paraffin sections from 46 cases of Hodgkin's disease were examined for the presence of the Epstein-Barr virus (EBV)-encoded latent membrane protein (LMP) using a sensitive (double layer alkaline phosphatase-anti-alkaline phosphatase) immunohistochemical method. LMP was detected in 22 cases, the majority of positive cases being of nodular sclerosis (12/24), mixed cellularity (6/7), and lymphocyte depletion (3/3) subtypes. Only one of 12 cases of lymphocyte predominant disease was positive. In all cases, reactivity was confined to Hodgkin's and Reed-Sternberg cells. These results provide further evidence for an association between EBV and Hodgkin's disease and indicate that LMP may be readily detected in archival material.

Herpesvirus 4, Human↗
PubMed1992

Three transcriptionally distinct forms of Epstein-Barr virus latency in somatic cell hybrids: cell phenotype dependence of virus promoter usage.

Phenotypically distinct human B cell lines display two transcriptionally distinct forms of Epstein-Barr virus (EBV) latency. Latency I (Lat I) in group I Burkitt's lymphoma (BL) cell lines is characterized by selective expression of the virus-coded nuclear antigen EBNA 1 from a uniquely spliced mRNA driven by the Fp promoter. Latency III (Lat III) in group III BL and EBV-transformed lymphoblastoid cell lines (LCLs) is characterized by expression of EBNAs 1, 2, 3a, 3b, 3c, and -LP from mRNAs driven by the Cp or Wp promoter and of the latent membrane proteins (LMPs 1, 2A, and 2B) from mRNAs driven by the LMP promoters. Here we have altered the group I BL and LCL phenotypes by cell hybridization and screened for attendant changes in EBV latency by PCR analysis of viral mRNAs and immunoblotting of viral proteins. Fusion of group I BL cells with LCLs activated the BL virus genome from a Lat I to Lat III pattern of gene expression. Fusion of LCLs with nonlymphoid lines repressed virus gene expression from Lat III either to Lat I or to another form of latency (Lat II) hitherto not seen in vitro and characterized by selective expression of the Fp-driven EBNA 1 mRNA and of the LMP 1, 2A, and 2B transcripts. There are therefore three forms of EBV latency which can be interconverted by altering cellular phenotype and thereby virus promoter usage.

Antigens, Viral↗
PubMed1992

Three pathways of Epstein-Barr virus gene activation from EBNA1-positive latency in B lymphocytes.

Previous studies on Epstein-Barr virus (EBV)-positive B-cell lines have identified two distinct forms of virus latency. Lymphoblastoid cell lines generated by virus-induced transformation of normal B cells in vitro, express the full spectrum of six EBNAs and three latent membrane proteins (LMP1, LMP2A, and LMP2B); furthermore, these lines often contain a small fraction of cells spontaneously entering the lytic cycle. In contrast, Burkitt's lymphoma-derived cell lines retaining the tumor biopsy cell phenotype express only one of the latent proteins, the nuclear antigen EBNA1; such cells do not enter the lytic cycle spontaneously but may be induced to do so by treatment with such agents as tetradecanoyl phorbol acetate and anti-immunoglobulin. The present study set out to determine whether activation of full virus latent-gene expression was a necessary accompaniment to induction of the lytic cycle in Burkitt's lymphoma lines. Detailed analysis of Burkitt's lymphoma lines responding to anti-immunoglobulin treatment revealed three response pathways of EBV gene activation from EBNA1-positive latency. A first, rapid response pathway involves direct entry of cells into the lytic cycle without broadening of the pattern of latent gene expression; thereafter, the three "latent" LMPs are expressed as early lytic cycle antigens. A second, delayed response pathway in another cell subpopulation involves the activation of full latent gene expression and conversion to a lymphoblastoidlike cell phenotype. A third response pathway in yet another subpopulation involves the selective activation of LMPs, with no induction of the lytic cycle and with EBNA expression still restricted to EBNA1; this type of latent infection in B lymphocytes has hitherto not been described. Interestingly, the EBNA1+ LMP+ cells displayed some but not all of the phenotypic changes normally induced by LMP1 expression in a B-cell environment. These studies highlight the existence of four different types of EBV infection in B cells, including three distinct forms of latency, which we now term latency I, latency II, and latency III.

Antigens, Viral↗
PubMed1992

The Epstein-Barr virus (EBV) nuclear antigen 1 BamHI F promoter is activated on entry of EBV-transformed B cells into the lytic cycle.

In Epstein-Barr virus (EBV)-positive Burkitt's lymphoma cell lines exhibiting the latency I form of infection (i.e., EBV nuclear antigen 1 [EBNA1] positive in the absence of other latent proteins), the EBNA1 mRNA has a unique BamHI Q/U/K splice structure and is expressed from a novel promoter, Fp, located near the BamHI FQ boundary. This contrasts with the situation in EBV-transformed lymphoblastoid cell lines (LCLs) exhibiting the latency III form of infection (i.e., positive for all latent proteins), in which transcription from the upstream Cp or Wp promoters is the principal source of EBNA mRNAs. We carried out cDNA amplifications with oligonucleotide primer-probe combinations to determine whether Fp is ever active in an LCL environment. The results clearly showed that some LCLs express a Q/U/K-spliced EBNA1 mRNA in addition to the expected Cp/Wp-initiated transcripts; this seemed inconsistent with the concept of Cp/Wp and Fp as mutually exclusive promoters. Here we show that Fp is indeed silent in latency III cells but is activated at an early stage following the switch from latency III into the virus lytic cycle. Four pieces of evidence support this conclusion: (i) examples of coincident Cp/Wp and Fp usage in LCLs are restricted to those lines in which a small subpopulation of cells have spontaneously entered the lytic cycle; (ii) transcripts initiating from Fp can readily be demonstrated in spontaneously productive lines by S1 nuclease protection; (iii) the presence of Fp-initiated transcripts is not affected by acyclovir blockade of the late lytic cycle; and (iv) infection of latently infected LCLs with a recombinant vaccinia virus encoding the EBV immediate-early protein BZLF1, a transcriptional transactivator which normally initiates the lytic cycle, results in the appearance of the diagnostic Q/U/K-spliced transcripts.

Antigens, Viral↗
PubMed1992

Characterization of EBV-positive lymphoblastoid cell lines obtained from HIV seropositive patients with or without lymphomas.

Epstein-Barr-virus- (EBV-) positive lymphoblastoid cell lines (LCLs) spontaneously arising in vitro were obtained from the peripheral blood of six HIV-seropositive patients and from the peripheral blood and the bone marrow of one patient (LAM) with AIDS and lymphoma. The LCLs from HIV-seropositive patients had phenotypic, cytogenetic, and biological characteristics indistinguishable from those of normal LCLs obtained by infecting B cells with EBV in vitro. The LCLs from LAM patient comprised composite cell populations. Cloning analysis and cell fractionation procedures showed that, beside normal EBV-infected cells, these lines contained a malignant subset population characterized by c-myc rearrangement, abnormal karyotype, and a surface phenotype similar to that of Burkitt's lymphoma cells. Analyses of Ig heavy chain and c-myc oncogene loci showed that these malignant cells were the progeny of a single precursor. Nevertheless, these cells had heterogeneous EBV-fused termini, a finding which indicates that EBV infection followed c-myc rearrangement.

Blotting, Southern↗
PubMed1992

Epstein-Barr virus, lymphomas and Hodgkin's disease.

Epstein-Barr virus (EBV) is implicated in the pathogenesis of a number of human lymphoid malignancies, including the immunoblastic B lymphomas that arise in immunocompromised individuals, Burkitt's lymphoma, Hodgkin's disease, and certain T cell lymphomas. The immunoblastic lymphomas are most likely a direct consequence of EBV-driven B cell lymphoproliferation that would in normal circumstances be eliminated by virus-specific cell-mediated immune responses. The other EBV-associated malignancies arise in individuals with more or less intact cellular immune responses and appear to have a complex multi-step pathogenesis. Throughout the EBV-positive lymphoid malignancies there is an intimate association between tumour cell phenotype and virus latent gene expression, with each of the three forms of latency seen in in vitro models being exemplified in vivo. Tumours with these different forms of virus latency will differ in their susceptibility to EBV-specific immune T cell control.

Cell Transformation, Neoplastic↗
PubMed1992

Epstein-Barr virus and carcinomas. Expression of the viral genome in an undifferentiated gastric carcinoma.

The Epstein-Barr virus (EBV) is associated with undifferentiated nasopharyngeal carcinomas (NPCs). Recently, EBV DNA has been demonstrated also in some cases of gastric carcinoma with similar morphology as undifferentiated NPC. Here we report on the expression of EBV genes in a gastric undifferentiated carcinoma of nasopharyngeal type (UCNT). The small EBV-encoded nuclear RNAs, EBERs, were expressed in all tumor cells. The BZLF1 transactivator protein, which disrupts viral latency, was detectable in a small subset of tumor cells. The latent membrane protein and the nuclear antigen EBNA2 were not expressed. These results indicate that lytic EBV infection may be possible in undifferentiated epithelial cells. Our findings add to the growing body of evidence suggesting a strong association of gastric UCNT with EBV and point to the possibility of an involvement of the virus in the pathogenesis of this neoplasm.

DNA, Viral↗
PubMed1991

Epstein-Barr virus DNA and latent gene products in Ki-1 (CD30)-positive anaplastic large cell lymphomas.

Epstein-Barr virus (EBV)-specific DNA sequences were detected by polymerase chain reaction analysis in 15 of 47 (32%) DNA extracts prepared from CD30-positive (Ki-1 antigen-positive) anaplastic large cell (ALC) lymphomas. EBV-encoded RNA (EBER) transcripts could be detected by in situ hybridization in the tumor cells of 9 of 11 EBV DNA-positive cases. Twenty-eight cases were examined by immunohistology on cryostat sections for the presence of the EBV-encoded latent membrane protein (LMP), the nuclear antigen 2 (EBNA2), the BZLF1 transactivator protein, and the late viral glycoprotein gp350/250. A distinct LMP-specific membrane and cytoplasmic staining was detected exclusively in lymphoma cells of five cases (18%); two of these cases additionally expressed EBNA2. BZLF1 protein and gp350/250 immunoreactivity was absent in all instances. All LMP-positive cases contained EBV DNA and EBER sequences. The pattern of EBV latent protein expression in ALC lymphomas showed heterogeneity with respect to EBNA2 expression: LMP-positive/EBNA2-negative cases displayed a pattern previously described for undifferentiated nasopharyngeal carcinomas and Hodgkin's disease, whereas LMP-positive and EBNA2-positive cases showed parallels to lymphoblastoid cell lines. Because the LMP gene has transforming potential, our findings support the concept of a pathoetiologic role for EBV in a proportion of CD30-positive ALC lymphomas.

Adult↗
PubMed1991

Activation of Epstein-Barr virus replication in Hodgkin and Reed-Sternberg cells.

Recent evidence has shown that Hodgkin's disease (HD) is associated with Epstein-Barr virus (EBV) in a substantial number of cases and that in these cases EBV DNA is localized exclusively to Hodgkin and Reed-Sternberg (RS) cells. The virus genome is not silent in RS cells because two EBV latent gene products, latent membrane protein (LMP) and EB early region (EBER) transcripts, have recently been reported to be expressed in RS cells. However, little information is available about the possible activation of EBV replicative genes in HD. This prompted us to investigate HD biopsies from 96 patients for expression of replicative gene products. Cryostat sections were immunostained with monoclonal antibodies to protein BZLF1, which controls the switch between EBV latency and replication, and also to LMP. LMP was demonstrated in RS cells in 47 cases (49%). Three of the LMP-positive cases (6%), but none of the LMP-negative cases, expressed the BZLF1 protein. BZLF1 positively was confined to rare RS cells. These three cases showed no detectable early, virus capsid, or membrane antigens. Our findings show that activation of EBV immediate early genes occurs only infrequently in RS cells, indicating that control of viral latency is not severely impaired in HD patients.

Antigens, Viral↗
PubMed1991

Restricted Epstein-Barr virus protein expression in Burkitt lymphoma is due to a different Epstein-Barr nuclear antigen 1 transcriptional initiation site.

Epstein-Barr virus (EBV) expresses six nuclear antigens (EBNAs) and three integral latent membrane proteins (LMPs) in latently infected growth-transformed B lymphoblastoid cell lines (LCLs). In contrast, EBV protein expression in Burkitt lymphoma tissue or in newly established Burkitt lymphoma cell lines is frequently restricted to the EBV genome maintenance protein, EBNA-1. EBNA-1 expression in the absence of other EBNAs and LMP-1 has been an enigma since, in LCLs, all EBNA mRNAs are processed from a single transcript. We now show that the basis for restricted EBV expression in Burkitt lymphoma cells is selective EBNA-1 mRNA transcription from a hitherto unrecognized promoter that is 50 kb closer to the EBNA-1-encoding exon than previously described EBNA-1 promoters. Infected cells with EBNA-1-restricted expression could preferentially persist in vivo in the face of EBV-immune T-cell responses, which are frequently directed against other EBNAs and are also dependent on LMP-1 expression.

Amino Acid Sequence↗
PubMed1991

Induction of bcl-2 expression by Epstein-Barr virus latent membrane protein 1 protects infected B cells from programmed cell death.

Epstein-Barr virus (EBV) not only induces growth transformation in human B lymphocytes, but has more recently been shown to enhance B cell survival under suboptimal conditions where growth is inhibited; both effects are mediated through the coordinate action of eight virus-coded latent proteins. The effect upon cell survival is best recognized in EBV-positive Burkitt's lymphoma cell lines where activation of full virus latent gene expression protects the cells from programmed cell death (apoptosis). Here we show by DNA transfection into human B cells that protection from apoptosis is conferred through expression of a single EBV latent protein, the latent membrane protein LMP 1. Furthermore, we demonstrate that LMP 1 mediates this effect by up-regulating expression of the cellular oncogene bcl-2. The interplay between EBV infection and expression of this cellular oncogene has important implications for virus persistence and for the pathogenesis of virus-associated malignant disease.

Antigens, Viral↗
PubMed1991

Salivary and serum IgA antibodies to the Epstein-Barr virus glycoprotein gp340: incidence and potential for virus neutralization.

Human antibody responses to the Epstein-Barr virus (EBV) glycoprotein gp340 have been measured using purified preparations of the native molecule as the substrate in ELISAs. This glycoprotein is the dominant component of the EBV envelope and a major target for the virus-neutralizing antibody response. Healthy virus carriers (both Caucasian and Chinese) regularly show detectable anti-gp340 IgG in serum and, unexpectedly, 21-30% of these individuals are also serum anti-gp340 IgA positive. Chinese patients with the EBV-genome-positive malignancy nasopharyngeal carcinoma (NPC) show elevated serum IgA antibodies to gp340 but, given the background of responses amongst healthy virus carriers, anti-gp340 IgA titres are a poorer diagnostic indicator of NPC than serum IgA antibodies detectable by immunofluorescence against the multicomponent EBV early antigen (EA). Salivary IgA antibody responses to gp340 are potentially important as a means of neutralizing orally-transmitted virus. We detected salivary IgA (but not IgG) to gp340 in a minority (12-19%) of healthy virus carriers and in a higher proportion (49%) of NPC patients. Even saliva samples chosen for their relatively high anti-gp340 IgA titres showed only weak neutralizing activity against transforming EBV preparations whether from B95.8 cell culture supernatant or from the throat washing of an infectious mononucleosis patient. We conclude that in healthy virus carriers, salivary IgA responses to gp340 are unlikely to provide effective local immunity against re-infection with a second EBV strain.

Adult↗
PubMed1991

Expression of Epstein-Barr virus latent gene products in tumour cells of Hodgkin's disease.

The Epstein-Barr virus (EBV)-encoded latent gene products, latent membrane protein (LMP) and EBV nuclear antigen 2 (EBNA 2), seem to have important roles in EBV-induced cell transformation in vitro, and have been implicated as important effector molecules in EBV-associated lymphomagenesis. Because up to 35% of Hodgkin's disease (HD) samples have been reported to contain EBV genomes, the expression of LMP and EBNA 2 in these tumours was investigated. 84 cases of HD were studied with monoclonal antibodies and immunohistochemical labelling of acetone-fixed cryostat sections. LMP, but not EBNA 2, was demonstrated in Reed-Sternberg (RS) cells of 40 cases (48%); the two proteins were easily detected in transformed lymphocytes of positive control acute infectious mononucleosis tonsils. LMP expression in RS cells varied according to the histological subtype of HD (1/10 cases [10%] of lymphocyte predominance subtype, 16/50 cases [32%] of nodular sclerosis, 23/24 [96%] cases of mixed cellularity type). That the LMP antibodies showed no substantial cross-reactivity with negative control tissues shows that they are useful probes for the diagnosis of latent EBV infection in tissue sections. The findings suggest that EBV is associated with more cases of HD than was previously recognised, that in positive cases RS cells express a latent infection protein phenotype (LMP+, EBNA 2-) which differs from that of other EBV-associated lymphomas, and that LMP expression is related to histologically aggressive subtypes of HD.

Antibodies, Monoclonal↗
PubMed1991

Epstein-Barr virus (EBV)-associated lymphoproliferative disease in the SCID mouse model: implications for the pathogenesis of EBV-positive lymphomas in man.

When human peripheral blood lymphocytes (PBLs) from Epstein-Barr virus (EBV)-seropositive donors are injected intraperitoneally into SCID mice, EBV+ B cell tumors develop within weeks. A preliminary report (Mosier, D. E., R. J. Gulizia, S. M. Baird, D. D. Richman, D. B. Wilson, R. I. Fox, and T. J. Kipps, 1989. Blood. 74(Suppl. 1):52a) has suggested that such tumors resemble the EBV-positive malignancy, Burkitt's lymphoma. The present work shows that generally the human (hu) PBL-SCID tumors are distinct from Burkitt's lymphoma and instead resemble lymphoblastoid cell lines (LCLs) generated by EBV-infection of normal B cells in vitro in terms of: (a) their cell surface phenotype, with expression of B cell activation antigens and adhesion molecules, (b) normal karyotype, and (c) viral phenotype, with expression of all the transformation-associated EBV latent proteins and, in a minority of cells, productive cycle antigens. Indeed, in vitro-transformed LCLs also grow when inoculated into SCID mice, the frequency of tumor outgrowth correlating with the in vitro growth phenotype of the LCL which is itself determined by the identity of the transforming virus (i.e., type 1 or type 2 EBV). Histologically the PBL-derived hu-SCID tumors resemble the EBV+ large cell lymphomas that develop in immuno-suppressed patients and, like the human tumors, often present at multiple sites as individual monoclonal or oligoclonal foci. The remarkable efficiency of tumor development in the hu-SCID model suggests that lymphomagenesis involves direct outgrowth of EBV-transformed B cells without requirement for secondary genetic changes, and that selection on the basis of cell growth rate alone is sufficient to explain the monoclonal/oligoclonal nature of tumor foci. EBV+ large cell lymphoma of the immunosuppressed may arise in a similar way.

Animals↗
PubMed1991

Expression of Epstein-Barr virus replicative proteins in AIDS-related non-Hodgkin's lymphoma cells.

Epstein-Barr virus (EBV) infection in lymphoproliferative lesions has been assumed to be strictly latent. In order to investigate the possible occurrence of EBV replication in AIDS-related lymphoma (ARL) cells, we studied 13 cases by immunohistology using monoclonal antibodies to the EBV-encoded switch-protein BZLF1, early antigens (EAs), late replicative proteins [virus capsid antigens (VCAs) and membrane antigens (MAs)], and to the latent proteins EB nuclear antigen 2 (EBNA 2) and latent membrane protein (LMP). EBV genomes were detected by in situ hybridization. EBV genomes and/or gene products were demonstrated in ten cases, including all immunoblast-rich lymphomas, two Burkitts lymphomas, and a T-cell anaplastic large-cell lymphoma. The BZLF1 protein, which disrupts latency in B cells, was identified in six (60 per cent), and EAs in four (40 per cent) of the EBV-positive ARL. Only one lymphoma (10 per cent) expressed VCAs and MAs. EBNA 2 and LMP were detected in three (30 per cent) and eight (80 per cent) of EBV-positive cases, respectively. EBV DNA was detected in lymphoma cells in 7 of 12 (58 per cent) cases. The most important finding of this study was frequent spontaneous activation of latent EBV in ARL. Production of complete virus, however, was either aborted, or tumour cells expressing late productive cycle proteins (VCA, MA) were rapidly cleared from tissues. It is suggested that host factors that normally inhibit replication of EBV are deficient in AIDS patients.

Adult↗
PubMed1991

O(6)-methylguanine-DNA methyltransferase and uracil DNA glycosylase in human broncho-alveolar lavage cells and peripheral blood mononuclear cells from tobacco smokers and non-smokers.

Because interindividual variations in the activities of DNA repair enzymes may be a risk factor in the pathogenesis of lung diseases, O(6)-methylguanine-DNA methyltransferase (O(6)-MT) and uracil DNA glycosylase (UDG) were measured in broncho-alveolar lavage cell (BALC) and peripheral blood mononuclear cell (PBM) samples from 57 healthy volunteers (25 smokers and 32 non-smokers). According to cotinine determination in 39 cases where serum for this was available, 38% of the self-acclaimed non-smokers had greater than 10 ng/ml of cotinine in their serum. Whether grouped into smokers and non-smokers according to clinical history or by serum cotinine, there were no statistically significant differences between these groups in O(6)-MT or UDG in either of the cell types. However, a tendency towards lower values in smokers was seen. The highest intraindividual variation in O(6)-MT activity was 7-fold, while the highest interindividual variation reached 18-fold. For UDG, the respective values were 24- and 307-fold. Although the distribution of O(6)-MT in BALC was different from that in PBM, the data are consistent with unimodality in both of the cell types. These findings suggest that exposure to cigarette smoke is not entirely responsible for the wide interindividual variation in O(6)-MT and UDG DNA repair activities.

Adult↗
PubMed1991

The Epstein-Barr virus carrier state: dominance of a single growth-transforming isolate in the blood and in the oropharynx of healthy virus carriers.

Epstein-Barr virus (EBV) isolates can be broadly classified as type 1 or type 2 on the basis of allelic polymorphism of the virus-encoded nuclear antigens EBNAs 2, 3a, 3b and 3c, and individually identified based on Mr values of their EBNA proteins (EBNA type). Here we have used this natural heterogeneity amongst isolates to re-examine the question of EBV persistence in vivo, asking in particular whether virus carriage in oropharyngeal epithelium and/or in B lymphoid tissues involves infection with a single or with multiple virus strains. Firstly, 76 healthy virus carriers were classified into serotype groups on the basis of preferential antibody reactivity to type 1 EBNAs (serotype 1) or to type 2 EBNAs (serotype 2); 60 of the 76 donors were serotype 1, four of the 76 donors were serotype 2 and 12 of the 76 donors were anti-EBNA 2, 3a, 3b, 3c antibody-negative and therefore could not be serotyped. Representative donors from each group were then selected for virus isolations from blood (by spontaneous in vitro transformation) and from throat washings (by cord blood cell transformation). All 13 serotype 1 donors tested and six of seven non-serotypeable donors gave a type 1 virus isolate, whereas all four serotype 2 donors and one of the seven non-serotypeable donors gave a type 2 isolate. Multiple transforming virus isolates from any one donor, whether from blood or throat washings, were all of the one strain characteristic of that particular donor; sequential isolations showed retention of the same strain over several years. Finally, throat washing samples from these same donors were examined for amplifiable EBV DNA in the polymerase chain reaction using EBV type-specific oligonucleotide primers and probes derived from the polymorphic EBNA 2 and EBNA 3c loci. The results were consistent with earlier virus isolation studies, each individual donor showing amplification either of type 1 or type 2 sequences. We conclude that multiple EBV infections must occur rarely, if at all, in healthy virus carriers; EBV persistence in vivo is characterized by dominance of a single transforming virus strain.

Adult↗