Biomedical subjects

M Rowe

Publications and source records attributed to M Rowe.

At least 55 records · Page 3Linked to original sources

PubMed1997

The role of inflammatory cytokines and nitric oxide in the pathogenesis of necrotizing enterocolitis.

PURPOSE: The role of inflammatory cytokines in the pathogenesis of necrotizing enterocolitis (NEC) is still undefined. Elevated levels of interleukin (IL)-6 and tumor necrosis factor (TNF)-alpha have been measured in infants with NEC, while elevated levels of nitric oxide (NO) have been reported in newborn infants with clinical sepsis. However, the cellular source of the NO or cytokines is unknown. The authors hypothesized that local intestinal production of NO induced by cytokines may contribute to the pathogenesis of bowel necrosis in NEC by inducing apoptosis (programmed cell death) or necrosis of the enterocytes. We examined the levels of inflammatory cytokines and NO in the intestine of infants undergoing surgical resection for NEC, and the cellular localization of human inducible NO synthase (NOS-2) in the inflamed gut. METHODS: We compared 15 patients undergoing bowel resection for NEC, with six infants (of similar age) undergoing intestinal resection for ileal atresia or stricture, meconium peritonitis, intussusception, or cecal perforation (control). Diagnosis of NEC was confirmed histologically. Representative segments of the surgical specimen were examined for messenger RNA (mRNA) for NOS-2 by Northern blotting and in situ hybridization. Cytokine mRNA was measured by polymerase chain reaction (PCR) because mRNA could not be detected by Northern blotting. The site of NO production was determined by in situ hybridization and immunohistochemistry. Apoptosis was measured using in situ DNA strand break extension (TUNEL). Nitrotyrosine immunoreactivity was assessed to determine if NO mediates cellular injury via peroxynitrite formation. RESULTS: Messenger RNA for NOS-2 was detected in nearly all patients with NEC except for one infant who underwent proximal diverting jejunostomy alone, and who did not have histological evidence of NEC at that site. NOS-2 mRNA was detected less frequently in control patients. In situ hybridization and immunohistochemistry showed that the enterocytes were the predominant source of NOS-2 activity in the intestine of NEC patients. Extensive apoptosis was seen in enterocytes in the apical villi of infants with NEC, and correlated with nitrotyrosine staining. NOS-2 activity was markedly diminished at the time of stoma closure, but remained elevated in infants who died from progressive disease. PCR showed variable cytokine mRNA expression in the intestine. Transforming growth factor (TGF)-beta expression was nearly identical in NEC and control. However, interferon (IFN)-gamma was present in 9 of 10 NEC, but only in one of six control patients. CONCLUSION: The data show that NO is produced in large quantity by enterocytes in the intestinal wall of infants with NEC and leads to apoptosis of enterocytes in apical villi through peroxynitrite formation.

Cytokines↗

PubMed1997

Outcomes of cataract extraction with multifocal intraocular lens implantation: functional status and quality of life.

OBJECTIVE: The authors measured functional status after bilateral implantation of a multifocal versus a monofocal intraocular lens (IOL). METHODS: A retrospective, case-control study was done on 100 subjects implanted bilaterally with a silicone-optic foldable zonal progressive IOL under United States Food and Drug Administration Investigational Device Exemption protocol and 103 control subjects implanted bilaterally with a monofocal IOL of similar design who were matched on age and postoperative corrected distance acuity. The Cataract TyPE specification, a 17-item functional status instrument, was modified, validated, and administered via telephone interview. The measures of vision, functional status, and quality of life were compared between patients and controls. RESULTS: The instrument was valid (Cronbach's alpha = 0.94) and correlated moderately (Pearson's r= 0.34) with Snellen acuity. Multifocal subjects were more likely than monofocal controls to never wear spectacles (41% vs. 11.7%; P < 0.001). Multifocal subjects rated their vision without spectacles significantly better than those with monofocal IOLs (9.0 vs. 7.9; P < 0.001). The difference was most significant in rating of near vision without spectacles (7.8 vs. 5.0; P < 0.001). Multifocal subjects reported less limitation in specific visual tasks without spectacles (0.3 vs. 0.8; P < 0.001, where 1.0 represents "slightly" limited). This difference was consistently observed in subscales related to distance vision activities (0.1 vs. 0.4; P < 0.001), near vision activities (0.6 vs. 1.4; P < 0.001), and social activities (0.1 vs. 0.6; P < 0.001). CONCLUSIONS: In this study, subjects with bilateral multifocal IOLs reported better overall vision, less limitation in visual function, and less spectacle usage than monofocal controls.

Aged↗

PubMed1997

Interoceptive exposure versus breathing retraining within cognitive-behavioural therapy for panic disorder with agoraphobia.

This study compared two components of a cognitive-behavioural treatment for panic disorder. Thirty-eight individuals with panic disorder and agoraphobia were randomly assigned to (a) cognitive restructuring, interoceptive exposure (i.e. repeated exposure to feared bodily sensations) and in vivo exposure to agoraphobic situations; or (b) cognitive restructuring, breathing retraining and in vivo exposure to agoraphobic situations. Assessments were conducted at pre-treatment, post-treatment and six months later. The treatments were equally effective on many measures. However, treatment that included interoceptive exposure was more effective at post-treatment in terms of panic frequency, overall severity and functioning, and more effective at six-month follow-up in terms of panic frequency, phobic fears and general anxiety and functioning. Follow-up results were limited due to attrition.

Adolescent↗

PubMed1997

Epstein-Barr virus EBNA3C represses Cp, the major promoter for EBNA expression, but has no effect on the promoter of the cell gene CD21.

EBNA3C is a potent repressor of transcription when bound to DNA as a fusion with the DNA binding domain (DBD) of GALA. A survey of promoters has revealed that the wild-type, unfused EBNA3C can specifically repress expression from reporter plasmids containing the Epstein-Barr virus Cp latency-associated promoter. Repression of Cp activity required amino acids 207 to 368, which encompasses a region resembling a basic DBD adjacent to a leucine zipper DNA binding motif and a site which binds to the cellular factor CBF1/RBP-Jkappa. However, amino acids 207 to 368 are dispensable when the protein is bound to DNA as a fusion with the GAL4 DBD, thus implicating this region in DNA binding. Mutation of the CBF1/RBP-Jkappa binding site in EBNA3C abrogated repression, strongly suggesting that CBF1/RBP-Jkappa is necessary for targeting the viral protein to Cp. Consistent with this result, mutation of the EBNA2 response element (a CBF1/RBP-Jkappa binding site) in Cp also prevented significant repression. In addition, amino acids 346 to 543, which were previously defined as important for the repressor activity of the GAL4-EBNA3C fusion proteins, also appear to be necessary for the repression of Cp. Since repression by these fusions was not observed in all cell types, it seems likely that EBNA3C either depends on a corepressor which may interact with amino acids 346 to 543 or is modified in a cell-specific manner in order to repress. These data are consistent with EBNA3C contributing to the regulation of EBNA expression in latently infected B cells through CBF1/RBP-Jkappa and another factor, but this need not directly involve EBNA2. Finally, although it has been reported that EBNA3C can upregulate CD21 in some B cells, we were unable to demonstrate any effect of EBNA3C on reporter plasmids which contain the CD21 promoter.

B-Lymphocytes↗

PubMed1996

Lymphoblastoid cells transfected with c-myc: downregulation of EBV-lytic antigens and impaired response of autologous CD4+ T cells in vitro.

Normal EBV-positive lymphoblastoid B-cell lines (LCL) were transfected with vectors containing the c-myc oncogene (pHEBO-E(mu)-myc) or control vectors (pHEBO-E(mu)) and analyzed for the expression of EBV-lytic and latent antigens. While EBV-latent antigens were normal in the c-myc transfectants, there was an almost complete downregulation of EBV-lytic antigens, including BZLF1, EA(D), gp340 and VCA. These observations were consistently repeated on 6 different LCLs transfected with c-myc. Unlike control LCLs, the c-myc transfectants did not release infectious EBV. PCR analysis demonstrated that BZLF1 mRNA was virtually absent in c-myc transfectants, possibly suggesting that the deregulated c-myc imposed a block in the EBV-lytic cycle at this particular level. c-myc transfectants failed to sustain the proliferative response of autologous CD4+ T-cell clones with specificity for EBV-lytic antigens. However, they regained this capacity after incubation with ultraviolet-inactivated EBV or gp340 antigen in vitro, also indicating that their antigen-presenting capacities were not impaired. c-myc transfectants failed to elicit a secondary proliferative response by autologous CD4+ T cells purified from the peripheral blood of EBV-seropositive donors. Exposure of c-myc transfectants to UV-inactivated EBV again resulted in a proliferative CD4+ T-cell response comparable to that elicited by the control LCLs. Collectively, our data provide evidence for the remarkable ability of an oncogene to influence the life cycle of a virus and to modify the antigenicity of the infected cells.

B-Lymphocytes↗

PubMed1996

CD40-induced growth inhibition in epithelial cells is mimicked by Epstein-Barr Virus-encoded LMP1: involvement of TRAF3 as a common mediator.

CD40, a member of the tumour necrosis factor receptor family, is expressed on the surface of B lymphocytes where its ligation provides a potent survival signal. CD40 is also expressed in basal epithelial cells and in a number of different carcinomas where its function remains unknown. We observed that contrary to the studies in normal B cells, CD40 ligation in carcinoma cell lines and in normal primary epithelial cells resulted in growth inhibition and enhanced susceptibility to apoptosis induced by anti-neoplastic drugs, TNF-alpha, Fas and ceramide. This effect was also observed in CD40-transfected Rat-1 fibroblasts. The expression of Bcl-2 did not affect growth inhibition induced by CD40 ligation in epithelial cells but the Epstein - Barr Virus-encoded latent membrane protein 1 (LMP1) blocked the effect. Whilst transient expression of LMP-1 resulted in the inhibition of epithelial cell growth, this effect was not observed with a LMP1 mutant lacking the binding domain for TRAF3, a protein which may mediate signal transduction by interacting with the cytoplasmic domains of both CD40 and LMP1. Transient expression of TRAF3 also inhibited epithelial cell growth, whilst expression of a dominant-negative TRAF3 partially blocked the inhibitory effect of CD40 ligation and of transient LMP1 expression. These results suggest that CD40 regulates epithelial cell growth in a manner mimicked by LMP1 and implicate TRAF3 as a common mediator in the transduction of the growth inhibitory signals generated via the CD40 and LMP1 pathways.

Animals↗

PubMed1996

Expression of the Epstein Barr virus transforming protein LMP1 causes a rapid and transient stimulation of the Bcl-2 homologue Mcl-1 levels in B-cell lines.

The EBV-encoded latent membrane protein 1 (LMP1) suppresses apoptosis in B lymphocytes through up-regulation of Bcl-2. However, the maximum induction of Bcl-2 by LMP1 takes about 48-72 h. We show in this report that up-regulation of the Bcl-2 homologue Mcl-1 by LMP1 preceded the induction of Bcl-2 and that the up-regulation was transient; therefore, Mcl-1 levels decreased when Bcl-2 levels started to increase. This finding supports the hypothesis that Mcl-1 functions as a rapidly inducible, short-term effector of cell viability. LMP1 also blocked the decline in the Mcl-1 levels in response to apoptotic stimulation triggered by elevated cyclic AMP. This effect of LMP1 was associated with a delayed cell death in the EBV-negative Burkitt lymphoma cell line BL41. The maintenance of Mcl-1 expression by LMP1 is likely to be a crucial immediate-early response that enables cells to survive until Bcl-2 can be up-regulated.

Apoptosis↗

PubMed1996

Cytostatic effect of Epstein-Barr virus latent membrane protein-1 analyzed using tetracycline-regulated expression in B cell lines.

Tetracycline-regulated vectors were used to obtain inducible expression in stable transfected B cell lines of two Epstein-Barr virus (EBV) latent genes, LMP1 and EBNA2. The transfected genes were tightly repressed by low, nontoxic concentrations of tetracycline (< or = 1 microgram/ml) and, following removal of tetracycline, were induced to levels comparable to or up to 3x that of EBV-transformed normal lymphoblastoid cell lines. In transfected DG75 cells, induced expression of LMP1, but not of EBNA2, led to the expected upregulation of various cell surface markers, including: CD40, CD54, CD58, and HLA class I.A novel observation was that both LMP1 and EBNA2 independently caused the downregulation of surface IgM, an effect mirrored in EBV-positive Burkitt lymphoma lines undergoing phenotypic drift during the transition from latency I to latency III in which both LMP1 and EBNA2 are upregulated. Most remarkably, induced LMP1 expression almost completely inhibited cell growth for 4 to 5 days, after which the cells recovered a limited proliferative capacity. The cytostatic effect of LMP1 was observed in all three B cell lines studied: DG75, BJAB, and Akata. Further analysis showed that induction of LMP1 coincided with a reduction in the levels of c-myc, and that the cytostatic effect was due to an accumulation of cells at the G2/M phase of the cell cycle. These data suggest a novel function for the LMP1 oncogene in controlling the proliferation of EBV-infected cells by regulating progress through G2/M phase.

Antigens, Surface↗

PubMed1996

A taxonomic study of the genus Listeria using sodium dodecyl sulphate-polyacrylamide gel electrophoresis of intracellular proteins.

Fifty-one strains of Listeria spp. and one strain of Jonesia denitrificans were subjected to sodium dodecyl sulphate-polyacrylamide gel electrophoresis and the resulting histograms compared using the unweighted pair group method with averaging. The majority of L. monocytogenes strains formed a discrete cluster with a minimum correlation of 73% similarity. The position of strain NCTC 11994 within the main L. monocytogenes cluster supports the recommendation by the National Collection of Type Cultures that it, rather than NCTC 10357, be adopted as the type strain of the species. All L. innocua strains formed a discrete cluster with a minimum correlation of 85% similarity. The similarity shown by L. grayi and L. murrayi, both between each other and with the other Listeria strains tested, support their designation as a single species within the genus Listeria. Although the single strain of J. denitrificans showed the least similarity with the other strains it was not sufficient to warrant support for its removal from the genus Listeria. This limited study provides a different perspective both between species in the genus Listeria and between the genera Listeria and Jonesia.

Bacterial Proteins↗

PubMed1996

Child mortality following standard, medium or high titre measles immunization in West Africa.

BACKGROUND: The World Health Organization (WHO) recommended the use of high titre measles vaccine in 1989. Subsequent long term follow-up of several trials yielded results suggesting higher mortality among children inoculated with medium and high titre vaccines compared to standard titre vaccines, although none of the individual trials found significant differences in mortality. METHODS: Long term survival after standard, medium and high titre measles vaccines has been investigated in a combined analysis of all West African trials with mortality data. In trials from Guinea-Bissau, The Gambia and Senegal, children received medium or high titre vaccines from 4 months of age and were compared to control groups recruited at the same time later receiving standard titre vaccine from 9 months of age. All children were followed up to at least 3 years old. RESULTS: Combining trials of high titre vaccines showed higher mortality among the high titre group compared to the standard group: mortality ratio (MR) = 1.33 (95% CI : 1.02-1. 73). Mortality among recipients of medium titre vaccines was not different from that in the standard vaccine group, MR = 1.11 (95% CI: 0.54-2.27). In a combined analysis by sex, the adjusted mortality ratios comparing high titre vaccine with standard vaccine were 1.86 (95% CI : 1.28-2.70) for females and 0.91 (95% CI : 0.61-1.35) for males. The trials were not designed to study long term mortality. Adjustments for several possible sources of bias did not alter the results. CONCLUSIONS: The combined analysis showed a decreased survival related to high titre measles vaccine compared with standard titre vaccines, though solely among females. As a result of these studies from West Africa and a study from Haiti, WHO has recommended that high titre measles vaccine no longer be used.

Age Distribution↗

PubMed1996

Frequency of multiple Epstein-Barr virus infections in T-cell-immunocompromised individuals.

The Epstein-Barr virus (EBV) carrier state is characterized by latent infection of the general B-cell pool and by chronic virus replication at oropharyngeal sites. In Caucasian populations, most healthy carriers seem to harbor one dominant transforming virus strain, usually of type I rather than type 2, which persists over time and is detectable both in the blood and in the throat. This finding implies that once the virus carrier state is established, both viral reservoirs are largely if not completely protected from infection with additional strains. However, it is not known which facets of the immune response offer that protection. Here we address this question by a detailed study of EBV carriage in patients T-cell immunocompromised as a result of chronic human immunodeficiency virus (HIV) infection. Resident EBV strains were rescued from blood and from throat washings by using an in vitro transformation assay which aims to minimize bias toward faster-growing transformants; in this way, a mean of 16 independent isolations were made from each of 35 HIV-positive (predominantly male homosexual) patients. These virus isolates were characterized first at the DNA level by PCR amplification across type-specific polymorphisms in the EBNA2 and EBNA3C genes and across the 30-bp deletion and 33-bp repeat loci in the LMP1 gene and then at the protein level by immunoblotting for the strain-specific "EBNAprint" of EBNA1, -2, and -3C molecular weights. By these criteria, 18 of 35 patients harbored only one detectable EBV strain, usually of type 1, as do healthy carriers. However, the other 17 patients showed clear evidence of multiple infection with different EBV strains. In eight cases these strains were of the same type, again usually type 1, and were more often found coresident in throat washings than in the blood. By contrast, a further nine patients gave evidence of coinfection with type 1 and type 2 strains, and in these cases both virus types were detectable in the blood as well as in the throat. Immunological assays on these HIV-positive patients as a group showed a marked impairment of T-cell responses, reflected in reduced levels of EBV-specific cytotoxic T-cell memory, but an elevation of humoral responses, reflected in raised antibody titers to the EBV envelope glycoprotein gp340 and by the maintenance of virus neutralizing antibodies in serum. We infer that selective impairment of the T-cell system predisposes the host to infection with additional exogenously transmitted EBV strains.

AIDS-Related Opportunistic Infections↗

PubMed1996

Isolation of intertypic recombinants of Epstein-Barr virus from T-cell-immunocompromised individuals.

All wild-type isolates of Epstein-Barr virus (EBV) analyzed to date for allelic polymorphisms of the nuclear antigen EBNA2 gene (in the BamHI YH region of the genome) and of the EBNA3A,-3B, -3C genes (tandemly arranged in the BamHI E region) have proved either uniformly type 1 or uniformly type 2 at all four loci. The absence of detectable intertypic recombination in the wild probably reflects the rarity with which individual carriers, and certainly individual target cells, become coinfected with both virus types. Studying a group of human immunodeficiency virus-positive T-cell-immunocompromised patients known to be at enhanced risk of multiple EBV infections, we have isolated intertypic EBV recombinants from 2 of 40 patients analyzed. These recombinants, whose in vitro transforming capacity appeared at least equal to that of type 1 strains, carried a type 1 EBNA2 allele and type 2 EBNA3A,-3B, and -3C alleles. This was clearly demonstrable at the DNA level by PCR amplification using type-specific primer-probe combinations and was confirmed at the protein level (for EBNA2 and EBNA3C) by immunoblotting with type-specific antibodies. In one patient, the recombinant appeared to be the predominant strain, being the virus most commonly rescued by in vitro transformation both from the blood and from the throat washings on two separate occasions 20 months apart. A regular type 1 virus strain was also present in this individual, but this was not related to the recombinant since the two viruses carried type 1 EBNA2 genes with different patterns of variance from the B95.8 prototype sequence. In the other patient, recombinants were isolated on one occasion from the blood and on a separate occasion, 21 months later, from the throat; these recombinants were almost certainly related, being identical at several genomic polymorphisms and differing only in one facet of the "EBNAprint," the size of the EBNA1 protein. Three different type 1 viruses were also isolated from this patient, two of which carried EBNA2 genes with the same pattern of sequence variation from B95.8 as the recombinant; however, since this is a fairly common pattern of variance, the relationship of these viruses to the recombinant remains an open question. We infer that intertypic recombinants of EBV are not uncommon in HIV-positive T-cell-immunocompromised patients, that they arise in such individuals as a consequence of their increased frequency of mixed-type infections, and that they will prove capable of efficient transmission in the human population.

Amino Acid Sequence↗

PubMed1996

The application of multiple quality-of-life instruments in individuals with mild-to-moderate psoriasis.

The 36-item Short-Form Health Survey Questionnaire (HSQ SF-36), Psoriasis Disability Index (PDI), and Dermatology Life Quality Index (DLQI) were administered to individuals with mild-to-moderate psoriasis to validate the general quality-of-life instrument against the dermatosis-specific, and the dermatology-specific, disability measure. The population consisted of 644 adults with psoriasis involving up to 20% of the body surface area, who were enrolled in 2 US multicentre, evaluator-blinded, parallel-group clinical trials for a new psoriasis medication. Patients averaged 16.5% of maximum possible disability as measured by the PDI, and 23.4% of maximum possible disability as measured by the DLQI. Normalised T-scores showed that the patients approximated US population means on all 8 of the HSQ SF-36 dimensions. The HSQ SF-36 scales did not reflect substantial quality-of-life impairment, although all showed statistically significant correlations with both the PDI and DLQI (correlation coefficients ranging from -0.13 to -0.45). Moreover, while the disability indices were more responsive to psoriasis characteristics than the HSQ SF-36 quality-of-life scales, all 8 HSQ SF-36 dimensions demonstrated sensitivity to at least some objective and/or subjective ratings of severity. The strongest relationships were observed between the PDI, DLQI and the HSQ SF-36 Mental Health and Social Functioning dimensions, suggesting that the HSQ SF-36 is sensitive to psychosocial suffering related to psoriasis, which is not conveyed in objective clinical measures of severity.

Adolescent↗

PubMed1996

Rectal bleeding in Prader-Willi syndrome.

Individuals with Prader-Willi syndrome manifest severe skin picking behavior. We report three patients with this syndrome in whom an extension of this behavior to rectal picking resulted in significant lower gastrointestinal bleeding and anorectal disease. The recognition of this behavior is important to avoid misdiagnosing inflammatory bowel disease in this group of patients.

Adolescent↗

PubMed1995

Heterogeneous expression of Epstein-Barr virus latent proteins in endemic Burkitt's lymphoma.

Epstein-Barr virus (EBV)-infected cells may sustain three distinct forms of virus latency. In lymphoblastoid cell lines, six EBV-encoded nuclear antigens (EBNA1, 2, 3A, 3B, 3C, -LP), three latent membrane proteins (LMP1, 2A, 2B), and two nuclear RNAs (EBERs) are expressed. This form of latency, termed latency III, is also encountered in some posttransplant lymphoproliferative disorders. In EBV-positive cases of Hodgkin's disease, the EBERs, EBNA1, and the LMPs are expressed (latency II), whereas in Burkitt's lymphoma (BL) only the EBERs and EBNA1 have been detected (latency I). We have studied the expression of EBV proteins in 17 cases of EBV-positive endemic BL by immunohistology. Expression of LMP1 was seen in variable proportions of tumor cells in two cases and EBNA2 was detected in some tumor cells in three other cases. Also, the BZLF1 trans-activator protein was expressed in a few tumor cells in 6 cases, indicating entry into the lytic cycle. A phenotypic drift from latency I to latency III has been observed previously in some BL cell lines. Our results suggest that a similar phenomenon may occur in BL in vivo and indicate that the operational definition of EBV latencies is not easily applied to human tumors.

Adolescent↗

PubMed1995

Class I major histocompatibility complex-restricted cytotoxic T lymphocytes specific for Epstein-Barr virus (EBV)-transformed B lymphoblastoid cell lines against which they were raised.

We have raised CD8+ cytotoxic T lymphocytes (CTL) from three Epstein-Barr virus-seropositive donors by incubating peripheral blood lymphocytes with irradiated autologous B95.8-strain EBV-transformed B lymphoblastoid cells (LCL). However, to detect lysis in a standard 51Cr release assay of the LCL against which these CTL were raised, superinfection with recombinant vaccinia expressing the appropriate EBV protein or incubation with the peptide epitope was necessary. The untreated LCL were not lysed, even though Western blotting demonstrated that they expressed the EBV antigens containing the CTL epitopes. We have found CTL of this phenotype that are restricted by human leukocyte antigen-A2, -A3, -B7, or -B39, and which recognize the EBV latent proteins, EBV nuclear antigen (EBNA)-3A, EBNA-3C, or terminal protein. During these experiments, we identified a new human leukocyte antigen-A3-restricted EBNA-3A epitope, residues 603-611, RLRAEAGVK. We raised a spontaneous LCL, transformed by endogenous EBV, from one donor, but this was also not lysed. For at least one of the epitopes, CTL from another donor lysed the LCL without superinfection or addition of peptides. We conclude that the CTL were unable to achieve a high enough avidity of interaction with untreated LCL to trigger effector function, although the LCL were able to stimulate them to grow in vitro for up to 4 mo. To assess whether a small percentage of the LCL might possess a higher antigen density, we used an assay of tumor necrosis factor release from a CTL clone, which was able to detect antigen-bearing cells representing only 1% of a stimulating LCL population. Nevertheless, the untreated autologous LCL line failed to stimulate tumor necrosis factor release.

Amino Acid Sequence↗

PubMed1995

Analysis of VH genes used by neoplastic B cells in endemic Burkitt's lymphoma shows somatic hypermutation and intraclonal heterogeneity.

Tumor cell lines from six typical cases of endemic Epstein-Barr virus (EBV) genome-positive Burkitt's lymphoma (BL) have been investigated for usage and mutational pattern of Ig VH genes. The neoplastic cells all had a t(8;14) (q24;q32) translocation involving the c-myc protooncogene. The VH genes were derived from VH1, VH3 and VH4, and both the IgM-positive (four cases) and IgG-positive (two cases) were extensively mutated from germline sequence. In two cases, early and late passage tumor cells were available, and the VH nucleotide sequences were identical, indicating that mutations had not accumulated in vitro. In a further case, there was evidence of sequence heterogeneity, which appeared to have been generated in vivo, indicating that the tumor cell VH gene was able to undergo posttranslocation somatic hypermutation. Analysis of the relatively nonpolymorphic VH4 genes for the pattern of replacement or silent mutations did not show a role for antigen selection in the expressed sequences.

Amino Acid Sequence↗

PubMed1995

The Epstein-Barr virus latent membrane protein-1 (LMP1) mediates activation of NF-kappa B and cell surface phenotype via two effector regions in its carboxy-terminal cytoplasmic domain.

The Epstein-Barr virus (EBV) encoded latent membrane protein, LMP1, is oncogenic in rodent fibroblasts and is an essential effector protein in EBV-induced growth-transformation of human B lymphocytes. Previous structure-function studies with LMP1 have relied largely on rodent fibroblast transformation as a functional readout, with apparently conflicting results. We have now analysed several LMP1 mutants in various human cell types, including B cells, T cells and epithelial cells, using two independent functional assays; (i) activation of NF-kappa B, and (ii) induction of two cell surface activation markers, CD54 and CD40. The results suggest that the cytosolic N-terminus is not essential for LMP1 function in any cell type studied. The third and fourth transmembrane helices and the intracytosolic loops are dispensable for activation of NF-kappa B, but they do influence the induction of CD54 and CD40. The major effector domain appears to be the cytosolic C-terminus in which were identified two 'C-terminal activating regions', CTAR-1 (residues 194-232) and CTAR-2 (residues 351-386). Whilst the exact results depended upon the host cell line, CTAR-2 was generally more important for activation of NF-kappa B, and both CTAR-1 and CTAR-2 were required for optimal induction of CD54 and CD40. Analysis of NF-kappa B activation by LMP1 in Rat-1 fibroblasts indicated that many mutations that were functional in human cells were poorly tolerated in the rodent cells; a result that is in broad agreement with published Rat-1 transformation data.

Animals↗