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Biomedical subjects

L Brooks

Publications and source records attributed to L Brooks.

At least 19 recordsLinked to original sources
PubMed1992

Distribution and phenotype of Epstein-Barr virus-harboring cells in Hodgkin's disease.

Distribution and phenotype of Epstein-Barr virus (EBV)-harboring cells were determined in Hodgkin's disease (HD) biopsies by in situ hybridization with [35S]-labeled RNA probes specific for the small EBV-encoded nuclear RNAs, EBER1 and EBER2, in some instances preceded by immunohistology for CD20, CD30, CD45RO, and CD68 antigens, the T-cell receptor beta-chain, and latent membrane antigen (LMP) of EBV. Twenty-three of 46 HD cases displayed EBER transcripts in all Hodgkin and Reed-Sternberg (H-RS) cells, and 18 of these cases showed LMP expression exclusively in neoplastic cells. EBER+ small reactive cells were present in 39 cases in low numbers, and in three cases in abundance. Thus, presence of H-RS cells with or without LMP expression was not accompanied by an unrestricted proliferation of reactive EBER+/LMP- lymphoid cells in the majority of HD patients. Simultaneous in situ hybridization with [35S]-labeled immunoglobulin light chain (IgLC) gene probes and nonisotopically labeled EBER probe showed a phenotype of mature B lymphocytes and a polyclonal composition for a large proportion of the EBER+ small cells. However, in contrast to noninfected cells, CD20 expression was not detectable in many of these cells, which may indicate downregulation of certain differentiation antigens in latently EBV-infected small lymphoid cells in vivo.

Antigens, CD↗
PubMed1992

Patterns of Epstein-Barr virus infection in non-neoplastic lymphoid tissue.

Taking advantage of the abundant expression of the small Epstein-Barr virus (EBV)-encoded RNAs (EBERs) in latently infected cells, we have analyzed 72 normal and hyperplastic lymph nodes and three tonsils of acute infectious mononucleosis (IM) for the presence and distribution of EBV+ cells using EBER-specific in situ hybridization, in some cases combined with immunohistologic demonstration of cell type-characteristic antigens. In IM, large numbers of EBV+ lymphoid B blasts were detectable in extrafollicular areas, whereas germinal centers were generally free of EBV+ cells. In reactive lymph nodes, the frequency of EBV+ cells varied with the degree of lymphoid hyperplasia and underlying immune status. The lowest numbers of EBV+ cells were detected in nonactivated lymph nodes and highest in human immunodeficiency virus-associated lymphadenopathy. If present in these lymph nodes, EBV+ cells were almost exclusively localized to extrafollicular areas, as also observed in IM. However, in contrast to IM, these cells were mainly small lymphocytes. Furthermore, in some instances, occasional scattered EBV+ cells were seen within germinal centers, and in two cases diffuse expansions of EBV+ cells occurred within a single germinal center each, indicating that under certain circumstances EBV+ B lymphocytes may participate in physiologic germinal center reactions. These findings reflect the interference of EBV with physiologic lymphoid differentiation pathways and provide a link to EBV-associated malignant lymphomas with a postulated origin from germinal center cells.

B-Lymphocytes↗
PubMed1992

Gold labelling of RNA in virus-induced mitochondrial vesicles in the sheep blowfly Lucilia cuprina.

Membranous vesicles are thought to be the replication site for viral RNA of many plant and animal viruses. A relatively rare site of virus-associated vesiculation is that of the mitochondrial outer membrane. In this study, virus-induced mitochondrial vesicles of the blowfly, Lucilia cuprina, were labelled with ribonuclease/gold and an antibody against double stranded RNA (anti-polyinosinic:polycytidylic acid). Both methods showed the presence of RNA in the vesicles thus indicating they may be a site for viral RNA replication in Lucilia.

Animals↗
PubMed1992

Epstein-Barr virus latent gene transcription in nasopharyngeal carcinoma cells: coexpression of EBNA1, LMP1, and LMP2 transcripts.

Epstein-Barr virus (EBV) genome-positive nasopharyngeal carcinomas (NPCs) regularly express the virus-coded nuclear antigen EBNA1, but not other EBNAs, and a subset of tumors also appear to be latent membrane protein LMP1 positive; the status of NPCs with respect to a second virus-coded latent membrane protein LMP2 is unknown. In the present work the EBV-NPC cell interaction has been analyzed at the RNA level with reverse transcription and polymerase chain reaction-based amplification to detect specific latent viral mRNAs. All four transplantable NPC cell lines studied and 17 of 18 fresh snap-frozen NPC biopsy specimens expressed an EBNA1 mRNA with a BamHI Q/U/K splice structure exactly like that recently identified in group I Burkitt's lymphoma (BL) cell lines and shown to be driven from a novel viral promoter, Fp. The BamHI Y3/U/K-spliced EBNA1 mRNA characteristic of virus-transformed B-lymphoblastoid cell lines (LCLs) was never found in NPCs. These same NPC biopsy specimens were then analyzed for evidence of the various LMP transcripts which are constitutively expressed in LCLs but down-regulated in BL cells. While only 3 of 18 tumors gave a clear LMP1 mRNA-specific signal after first-round amplification with either of two sets of polymerase chain reaction primers, the majority proved to be LMP1 mRNA positive after second-round amplification with nested primers. A rather similar pattern of results was obtained with respect to LMP2B mRNA expression, such transcripts being detectable only in a subset of tumors, and then at apparently low levels. In contrast, clear evidence of LMP2A mRNA expression was obtained in 17 of 17 fresh biopsies. The predominant form of EBV infection in NPCs, with coexpression of EBNA1 and LMP mRNAs, is therefore quite distinct from that seen in BL cells (in which EBNA1 is the only expressed mRNA) and in LCL cells (in which all six EBNA and three LMP transcripts are present). This third form of EBV latency may not be restricted to NPC but may have more general relevance in the context of EBV infection in vivo.

Antigens, Viral↗
PubMed1992

Expression of Epstein-Barr virus genes and of lymphocyte activation molecules in undifferentiated nasopharyngeal carcinomas.

Previous studies investigating the role of Epstein-Barr virus (EBV) in undifferentiated nasopharyngeal carcinoma (NPC) have been performed on extracts from biopsies. The authors analyzed expression and localization of viral gene products in 18 undifferentiated NPCs at the cellular level using immunohistology and in situ hybridization. All cases were EBV-positive. The small nuclear EBV-encoded RNAs, EBERs, were regularly expressed whereas the latent membrane protein, LMP1, and EBV was detectable only in four cases (22%) and the nuclear antigen 2 was not detectable. The BZLF-1 protein of EBV which disrupts viral latency, was not detectable, confirming that the virus is latent in the tumor cells. Although the expression of the CD23 antigen in transplantable NPCs has been reported, our study demonstrates that expression of this antigen in human undifferentiated NPCs is rare. In contrast, almost all cases expressed the CDw70 antigen. Since in normal tissues this antigen is present only in activated lymphoid blasts, this finding may be relevant for the differential diagnosis of undifferentiated NPCs.

Antibodies, Monoclonal↗
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

Effect of deletions in the cauliflower mosaic virus polyadenylation sequence on the choice of the polyadenylation sites in tobacco protoplasts.

Deletions were made in the cauliflower mosaic virus polyadenylation sequence which was cloned downstream of the beta-glucuronidase gene (gus). The populations of mRNAs generated in tobacco mesophyll protoplasts by transient expression with the various constructs were analysed using a polymerase chain reaction procedure. When no deletion was present in the sequence, the mRNA appeared to be polyadenylated at two major polyadenylation sites. A deletion upstream from the AATAAA sequence made the population of polyadenylated mRNAs very heterogenous at their 3' ends. A deletion downstream of the AATAAA sequence had no effect on the choice of the site. Alternative polyadenylation sites were used when the native polyadenylation site was deleted. These results are discussed in relation to data obtained with other polyadenylation sequences from both plants and animals.

Base Sequence↗
PubMed1991

Intrauterine devices in nulliparous women.

The majority of studies on the clinical events following the insertion of an intrauterine device for contraception have observed a higher frequency of adverse effects in nulliparous women. In this review, the significance of nulliparity on the occurrence of medical removal for bleeding and pain, expulsion and pelvic inflammatory disease is estimated. The possible role of the anatomy of the pregravid uterus is discussed, and results obtained through a selective measurement of the endometrial cavity in a population of nulligravidas is presented. It is concluded that IUDs should not be the first choice of contraception in nulliparous women because of an increased risk of long-term adverse effects. The relationship between the length of the intrauterine device and the length of the endometrial cavity does not seem to be of clinical significance for the performance of IUDs in nulliparous women.

Female↗
PubMed1991

Communicating probabilistic information to cancer patients: is there 'noise' on the line?

The objective was to examine the way that cancer patients translate verbal descriptors of probability into numerical estimates. A list of words commonly used on consent forms to describe the likelihood for benefits or risks of therapies was provided to 100 cancer patients. Two formats, paper/pencil or computer, were used to provide the list of words. Two methods, magnitude estimation and linear analogue scaling, were used to obtain probability estimates for each word. In addition, two scenarios were developed to study 'context effects' on numerical interpretations of verbal descriptions of probability. All patients provided numerical values for the words on two occasions, separated by one week, and two interviewers collected the data. Regardless of method or format, each word elicited widely variable numerical interpretations. An ANOVA model, including patient, word, interviewer, time, method and format, indicated that patient and interviewer produced major effects on probability estimates. Agreement between methods and across time was good. Paper/pencil and computer formats yielded similar results. Context effects did not appear to influence the numerical probabilities elicited by the 2 scenarios. It was concluded that, within this group of patients, there was no consensus about numerical meaning of a given word, and that interviewers can systematically influence numerical interpretations. There appears to be a great deal of 'noise' in this particular line of communication between patients and health professionals.

Aged↗
PubMed1991

Magnesium substitution in acute ischaemic heart syndromes.

It has recently been suggested that intravenous infusion of magnesium may reduce mortality and the incidence of serious arrhythmias in patients with ischaemic heart disease and acute myocardial infarction. In the present double-blind, placebo-controlled study, 298 patients with suspected acute myocardial infarction were randomized to receive either intravenous magnesium chloride (80 mmol.24 h-1) or placebo. Infusions were started immediately after admission to the coronary care unit. One hundred and fifty patients received magnesium and 148 the placebo. Ischaemic heart disease was diagnosed in 244 patients. Acute myocardial infarction was observed among 83 patients in the magnesium group and 79 in the placebo group. Both treatment groups were comparable regarding sex, age, clinical status, previous cardiac disease and medication. Serum magnesium was significantly raised during magnesium infusion compared to placebo (P less than 0.01). Fatal events were only observed among patients with myocardial infarction, but neither the in-hospital mortality (magnesium: 12.1%; placebo 10.1%) nor the mortality after a follow-up period of 245 days (median observation time) was affected by magnesium substitution. Magnesium infusion was accompanied by a significantly increased incidence of atrioventricular conduction disturbances. The results suggest that patients suffering from acute ischaemic heart syndromes do not benefit from intravenous magnesium supplementation.

Acute Disease↗
PubMed1991

Epstein-Barr virus infection in oral hairy leukoplakia: virus replication in the absence of a detectable latent phase.

Epstein-Barr virus (EBV) infects both B lymphocytes and oropharyngeal epithelium, and it has been argued that the true reservoir of virus persistence in vivo is the self-renewing basal epithelial compartment. The identification of oral hairy leukoplakia (HL) of AIDS patients as a clinically apparent focus of EBV replication in lingual epithelium therefore provides a means of studying the EBV-epithelial cell interaction in situ. Replicative EBV DNA and productive cycle antigens are restricted to the upper, more differentiated epithelial layers in HL, and here we have applied highly sensitive in situ hybridization and immunohistological methods to examine the lower basal/suprabasal layers for evidence of latent EBV infection. We could not detect EBV DNA in these layers using an in situ DNA hybridization protocol which, on reference B cell lines, detected 1 viral genome/cell. Likewise, using sensitive in situ RNA hybridization for both the small non-polyadenylated EBER RNAs (abundant transcripts seen in all known forms of EBV latency) and the latent membrane protein (LMP) mRNA (the most abundant viral mRNA in B lymphoblastoid cell lines), the basal/suprabasal cells in HL were consistently negative; immunohistological staining with specific monoclonal antibodies also gave no evidence of latently infected LMP-positive cells. When the biopsy extracts were analysed by immunoblotting with selected human antisera, in addition to abundant productive cycle antigens, a band of constant size (66K) was observed which also reacted with immunopurified antibodies monospecific for one of the latency-associated nuclear antigens, EBNA 1; the cellular origin of this EBNA 1 could not be ascertained, but it is possible that in HL the protein is expressed during the productive cycle. The absence of demonstrable EBV latency in the basal/suprabasal cells of HL suggests that this is purely a virus replicative lesion which is sustained by continual re-infection of the maturing epithelium, not by the maturation of latently infected cells from the basal compartment.

Antigens, Viral↗
PubMed1990

Role of arginine 67 in the stabilization of chymotrypsin inhibitor 2: examination of amide proton exchange rates and denaturation thermodynamics of an engineered protein.

We have examined the contribution to protein stability of an interaction involving a charged hydrogen bond from an arginyl side chain (Arg67) in the serine proteinase inhibitor chymotrypsin inhibitor 2 (CI-2), by replacing this side chain with an alanyl residue by protein engineering. Using nuclear magnetic resonance spectroscopy (NMR), we have examined the effect of this mutation on the hydrogen-deuterium exchange rates of several backbone amide protons in the native and engineered proteins at 50 degrees C. These exchange rates provide a localized probe at multiple discrete sites throughout the protein and from comparison of native and mutant exchange rates allow calculation of the difference in free energy of exchange (delta delta Gex) resulting from the mutation. The results show that for the majority of amides observed this mutation results in delta delta Gex of ca. 1.7 kcal mol-1 over the whole CI-2 molecule. However, for two relatively exposed amide protons the exchange rates are found to be far less perturbed, implying that local unfolding mechanisms predominate for these protons. Direct measurement of the stability of both proteins to denaturation by guanidinum hydrochloride shows that the interaction contributes 1.4 kcal mol-1 to the stability of the molecule. This value is comparable to those obtained from the NMR exchange measurements and indicates that the exchange processes reflect the differences in stability between the native and mutant proteins.(ABSTRACT TRUNCATED AT 250 WORDS)

Arginine↗
PubMed1990

Sulfonamide resistance gene for plant transformation.

The sulfonamide resistance gene from plasmid R46 encodes for a mutated dihydropteroate synthase insensitive to inhibition by sulfonamides. Its coding sequence was fused to the pea ribulose bisphosphate carboxylase/oxygenase transit peptide sequence. Incubation of isolated chloroplasts with the fusion protein synthesised in vitro, showed that the bacterial enzyme was transported to the chloroplast stroma and processed into a mature form. Expression of the gene fusion in transgenic plants resulted in a high level of resistance to sulfonamides. Direct selection of transformed shoots on leaf explants was efficient using sulfonamides as sole selective agents. Transformed shoots rooted normally on sulfonamides at concentrations toxic for untransformed ones. Sulfonamide resistance was transmitted to the progeny of transformed plants as a single Mendelian dominant character. These results demonstrate that this chimeric gene can be used as an efficient and versatile selectable marker for plant transformation.

Amino Acid Sequence↗
PubMed1990

Expression of the sulfonamide resistance gene from plasmid R46.

The expression of the sul I gene from plasmid R46, a wide host range plasmid of the IncN incompatibility group, was studied in Escherichia coli. Using a promoter test vector, a promoter was detected upstream of the sul I gene. From a nuclease protection experiment, the transcription was determined to start 360 bp upstream of the coding sequence. Two putative promoter -35 and -10 sequences were found upstream from the predicted transcription start. The presence of this promoter sequence in other R factors was discussed in relation with previous data showing that the sul I genes were transcribed from other promoters. The translation product of the sul I gene was detected in minicells. Its size indicates that the translation starts at the first ATG codon found in the open reading frame.

Base Sequence↗
PubMed1989

Epstein-Barr virus gene expression in malignant lymphomas induced by experimental virus infection of cottontop tamarins.

Inoculation of cottontop tamarins with a large dose of Epstein-Barr virus (EBV) leads to the induction of multiple EBV genome-positive lymphomas. These tumors have been characterized as oligoclonal or monoclonal large-cell malignant lymphomas that closely resemble the EBV genome-positive B-cell lymphomas that arise in human allograft recipients. The expression of latent and lytic EBV-encoded proteins was investigated in these virus-induced tamarin lymphomas and in derived cell lines. The tamarin tumors were found to express EBV nuclear antigen 1 (EBNA 1), EBNA 2, EBNA leader protein, and the latent membrane protein (LMP) as determined both by immunohistochemical staining and by immunoblotting. However, within the limits of the immunoblotting assays, no expression of the EBNA 3a protein family could be detected. Assays for lytic-cycle proteins by using both polyclonal human sera and monoclonal antibodies against viral capsid antigen, early antigen, and membrane antigen (gp340/220) showed minimal, if any, expression of these antigens in the lymphoma biopsies. In contrast, the cell lines derived from these lymphomas, even in early passage, expressed abundant levels of the lytic-cycle antigens and also expressed the EBNA 3a protein as well as EBNA 1, EBNA 2, EBNA leader protein, and LMP. This finding suggests that the virus-lymphoma cell interaction, in particular the switch to lytic cycle, is subject to some form of host control in vivo. The expression of EBNA 2 and LMP in these tamarin lymphomas strengthens their resemblance to posttransplant lymphomas in humans, since these human tumors are also EBNA 2 and LMP positive (L. S. Young, C. Alfieri, K. Hennessy, H. Evans, C. O'Hara, K. Anderson, A. Rickinson, E. Kieff, and J. I. Cohen, submitted for publication). Since both proteins are known to be important effector molecules of virus-induced B-cell growth transformation in vitro, their expression in these lymphomas constitutes the best evidence for a direct oncogenic role for EBV in vivo.

Animals↗
PubMed1987

Retinal vasculitis in Crohn's disease.

Although systemic vasculitis is a prominent part of the extraintestinal complications of Crohn's disease, the association of a retinal vasculitis with this chronic granulomatous disease of the gastrointestinal system is rare. We examined a 26-year-old woman with biopsy-proven Crohn's disease who developed a severe bilateral, obliterative retinal arteritis and phlebitis, leading to a marked loss of vision. Treatment with systemic corticosteroids and cyclophosphamide succeeded in halting progression of the disease.

Adult↗