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Biomedical subjects

B E Roberts

Publications and source records attributed to B E Roberts.

At least 19 recordsLinked to original sources
PubMed1998

Poly[di(carboxylatophenoxy)phosphazene] (PCPP) is a potent immunoadjuvant for an influenza vaccine.

The adjuvant activity of poly[di(carboxylatophenoxy)phosphazene] (PCPP) on the immunogenicity of formalin-inactivated influenza virions and commercial trivalent influenza vaccine was studied. Regardless of which antigen preparation is used, the addition of 100 micrograms PCPP enhances the HAI antibody response 10-fold over the levels elicited by the vaccine alone. Similarly, PCPP enhanced the IgM, IgG, and IgG1 ELISA antibody titers to influenza antigens at least 10-fold higher than the vaccine alone. In contrast, the IgG2a isotype titers were only enhanced about 2-fold. Immunization of aged mice (22 months old) with trivalent influenza vaccine alone did not sero-convert these mice as measured by HAI or ELISA whereas significant sero-conversion was achieved when mice were immunized with PCPP-formulated trivalent vaccine. The adjuvant activity of PCPP was shown to not be due to a site of injection depot effect. PCPP adjuvanticity was positively correlated to the molecular weight of the polymer.

Adjuvants, Immunologic↗
PubMed1998

Trisomy 12 is seen within a specific subtype of B-cell chronic lymphoproliferative disease affecting the peripheral blood/bone marrow and co-segregates with elevated expression of CD11a.

In order to delineate the specific morphological and immunophenotypic features of B-cell lymphoproliferative disorders associated with trisomy 12, 172 sequential unselected cases of CD19+CD5+ B-cell disorders, primarily affecting the peripheral blood and bone marrow, were studied. Trisomy 12 was found in 24 cases (13.9%), with all cases morphologically classified as either CLL-PL or CLL-mixed by FAB criteria. Trisomy 12 was not found in any cases of typical CLL. Trisomy 12 cases demonstrated a significant higher expression of CD11a (P<0.0001) and CD20 (P<0.0006) when compared to cases with the equivalent morphology and immunophenotype, but without the chromosomal abnormality. Trisomy 12 cases also demonstrated a higher frequency of FMC7, CD38 expression and moderate to strong surface immunoglobulin staining. However, no correlation was detected between the percentages of trisomy 12 cells and cells expressing CD11a, CD38, FMC7 or sIg mean fluorescent intensity. Cells from trisomy 12 positive cases were sorted according to their CD11a expression using fluorescent activated cell sorting. There was a significant increase in the percentage of trisomy 12 cells within the CD11a+ sorted fraction compared to the unsorted population (P < 0.05), implying that trisomy 12 is associated with increased expression of CD11a. With the highly specific morphological and immunophenotypic features demonstrated by trisomy 12 cases in this study, it is highly likely that these cases constitute a specific group of B-cell lymphoproliferative disorders.

Adult↗
PubMed1997

Use of radioactive phosphorus in haematology.

Following the development of the cyclotron in 1932, radio-isotopes became available for use in medicine both as tracer substances and therapeutic agents. The father of nuclear medicine, Dr J. H. Lawrence, pioneered their use in a range of disease states and found that radio-isotopes were of enormous value in the diagnosis and treatment of haemopoetic disease, particularly the myeloproliferative disorders. Radioactive phosphorus 32P emerged as the radio-isotope of choice for the myelosuppressive treatment of myeloproliferative disorders. This article also describes the use of radio-isotopes in the treatment of other disorders: chronic myeloid leukaemia, chronic lymphocytic leukaemia and myeloma, work that is now largely forgotten. All myeloproliferative disorders may evolve without treatment into myelodysplastic syndrome or blast-cell transformation. It is accepted that life is prolonged in myeloproliferative disorders treated with 32P or alkylating agents, yet both are leukaemogenic. The ideal form of treatment for polycythaemia vera is unknown and will remain so, for patients with this disorder often outlive their physician and achieve 90% of normal life expectation. 32P remains the treatment of choice for elderly patients with polycythaemia vera.

Animals↗
PubMed1995

Water-soluble phosphazene polymers for parenteral and mucosal vaccine delivery.

PCPP can be used in two different ways to potentiate an immune response. The soluble form of the polymer has been found to have immunoadjuvant activity. A single subcutaneous injection of polymer/influenza dramatically increases the ELISA, neutralizing, and HI antibodies to influenza virus compared to CFA. The polymer has also succeeded in dramatically increasing the amount of ELISA antibodies to TT. The antibody response elicited was predominantly of the IgG1 isotype. PCPP has also been used to generate micron-sized hydrogel microspheres through a process of divalent ion cross-linking of the polymer strands. These microspheres can induce significantly higher anti-TT serum IgG titers after a single intranasal immunization than TT alone.

Adjuvants, Immunologic↗
PubMed1994

Clonality of CD3 negative large granular lymphocyte proliferations determined by PCR based X-inactivation studies.

AIMS: To examine persistent CD3-large granular lymphocytosis (LGL) cases for clonality, both by lineage specific (T cell receptor) and lineage independent (X-inactivation) molecular methods; and to find out whether X-inactivation studies are more appropriate than gene rearrangement studies for this subset of LGL disorders. METHODS: Patients were selected who had LGL of more than six months' duration and identified as CD3- by immunophenotyping. T cell receptor studies and, where possible, X-inactivation studies of the phosphoglycerate kinase (PGK) gene were carried out. Analysis of subpopulations was carried out on cases heterozygous for PGK by the use of a polymerase chain reaction (PCR) method for X-inactivation. RESULTS: Of 17 CD3- LGL cases studied, all were found to be germline for beta, gamma, and delta T cell receptor studies, and immunoglobulin heavy chain genes. However, six of these were analysed by X-inactivation of the PGK gene and two cases gave clonal band patterns but only within the CD3- subpopulation. CONCLUSIONS: Clonal analysis by the lineage independent method of X-inactivation allows clonal expansion undetected by T and B cell specific markers to be identified. It is therefore a more appropriate method for the analysis of CD3- LGL. This has implications for diagnosis in CD3- LGL disorders.

Adult↗
PubMed1993

Host range selection of vaccinia recombinants containing insertions of foreign genes into non-coding sequences.

A simple yet powerful selection system was developed for the insertion of foreign genes in vaccinia virus. The selection system utilizes the vaccinia virus K1L (29K) host range gene which is located in HindIII M. This gene is necessary for growth in RK-13 cells but not in BSC40 or CV-1 cells. A vaccinia mutant (vAbT33) unable to grow on RK-13 cells was constructed having sequences at the 3' end of the K1L gene and the adjacent M2L gene deleted and replaced with the beta-galactosidase gene regulated by the BamHI F (F7L) promoter. A recombination plasmid containing the hepatitis B surface (HBs) antigen gene regulated by the M2L promoter and the complete sequence of the K1L gene was used to insert the HBs gene into vAbT33. The M2L negative K1L positive recombinant was easily isolated in two rounds of plaque purification by plating the virus on RK-13 cell monolayers. The K1L gene selection system allows the isolation of recombinants arising at frequencies as low as 1/100,000. It was noted that recombinants containing vaccinia sequence duplications (promoters) resulted in intragenomic recombinations that eliminated all sequences between the duplications. A second recombination plasmid was constructed that allowed insertion into the vaccinia genome without the loss of vaccinia coding sequences. This was achieved by insertion of the pseudorabies virus GIII gene regulated by the vaccinia H5R (40K) promoter between the translation and transcription stop signals at the 3' end of the K1L gene. The K1L gene transcription stop signal thus became the stop signal for the inserted GIII gene and an upstream transcription stop signal present in the H5R promoter fragment provided the stop signal for the K1L gene. This manipulation of the vaccinia genome had no effect on the accumulation or 5' end of the M2L gene transcripts. Although the insertion lengthened the 3' end and lowered the accumulation of K1L transcripts it altered neither the virulence nor the immunogenicity of the recombinant.

Base Sequence↗
PubMed1993

Is pure red cell aplasia (PRCA) a clonal disorder?

Pure red cell aplasia (PRCA) is an uncommon disorder, many cases lacking a well defined aetiology. This report describes three cases of PRCA (two idiopathic and one associated with B-CLL) who were investigated to assess the possibility of their PRCA being associated with a clonal proliferation of T-lymphocytes. The results show that one patient had evidence of T-cell receptor (TCR) gamma chain rearrangement, and the other had a TCR delta chain rearrangement. These two cases raise the possibility of PRCA being associated with a clonal proliferation of T-cells and further studies are warranted.

Aged↗
PubMed1992

Molecular attenuation of vaccinia virus: mutant generation and animal characterization.

These studies demonstrated that the inbred BALB/c mouse strain can be optimized for the assessment of vaccinia virus virulence, growth, and spread from the site of inoculation and immune protection from a lethal vaccinia virus challenge. The studies established that manipulation of the vaccinia virus genome generated mutants exhibiting a wide range of attenuated phenotypes. The nine NYCBH vaccinia virus mutants had intracranial 50% lethal doses that ranged from 2 to greater than 7 log10 units. The decreased neurovirulence was due to decreased replication in brain tissue. Three mutants had a decreased ability to disseminate to the lungs, brains, livers, and spleens of mice after intranasal infection. One mutant had a decreased transmission from mice infected by tail scarification to naive cage mates. Although the mutants, with one exception, grew to wild-type titers in cell culture, they showed a growth potential on the scarified skin of mice that was dramatically different from that of the wild-type virus. Consequently, all of the mutants had significantly compromised immunogenicities at low virus immunization doses compared with that of the wild-type virus. Conversely, at high immunization doses most mutants could induce an immune response similar to that of the wild-type virus. Three Wyeth vaccine strain mutants were also studied. Whereas the thymidine kinase, ribonucleotide reductase, and hemagglutinin mutants had a reduced virulence (50% lethal dose), only the thymidine kinase mutant retained its immunogenicity.

Administration, Intranasal↗
PubMed1992

Patterns of membrane CD45 isoform expression by leukaemic blasts and normal mature myeloid cells.

The membrane expression of CD45R isoforms by leukaemic blasts from 92 cases of acute myeloid (AML) and 12 cases of lymphoblastic leukaemia was analysed by single and two-colour flow cytometry. Compared to normal mature granulocytes, which invariably expressed UCHL1 (CD45RO) but not 2H4 (CD45RA), leukaemic myeloid blasts showed 2H4+ UCHL1- and 2H4- ++UCHL1- composite patterns of expression with the first of these phenotypes being associated in particular with the most immature AML subtype (M0). Similar analyses of blast cell fractions from monocytic AML variants, revealed wide variation in CD45R expression in cases of M4, whereas M5 leukaemias were typically 2H4+ ++UCHL1+ with minor 2H4- UCHL1- components. As most normal mature monocytes were also 2H4+ ++UCHL1+, this suggested that monocytic myeloid differentiation was primarily associated with the "acquisition" of UCHL1 and the development of 2H4/UCHL1 co-expressing cells. The expression of membrane CD45RA by leukaemic blasts is an abnormal characteristic and may be related to the maintenance of an undifferentiated state by malignant myeloid precursors.

Adult↗
PubMed1991

An alternative explanation for anomalies in "soluble lipofuscin" fluorescence data from insects, crustaceans, and other aquatic species.

Published attempts to extract lipofuscin from crustaceans and fish to assess age for fisheries research purposes have used essentially the same extraction methodology applied to insects, but have neither shown a conclusive age-dependence of spectrally similar fluorescence nor proved its association with lipofuscin. The reported lipofuscin solvent extraction method for fleshflies, Sarcophaga bullata, was manipulated by varying wash volume. This revealed that almost all age-dependent blue fluorescent material persisting in lipid fractions was actually pteridine-like. This finding was consistent with some previous independent results for Musca domestica. Examination of reported lipofuscin extraction protocols for other insects suggested that this problem was probably widespread. The pteridines are known to occur in unusually high amounts in insects, accumulating with age specifically in some members of this group by storage excretion, probably as a terrestrial water conservation strategy. In addition, there is growing evidence in the gerontological literature for other groups that solvent extracted blue fluorescence is not a true measure of lipofuscin content in tissues. These findings provide considerable insight into anomalies in putative lipofuscin fluorescence data between the insects and various aquatic species and suggest that there may be little basis for expectations of age-dependent fluorescence from aquatic species when the same gross extraction and crude purification methods are used.

Animals↗
PubMed1991

Patterns of CD16 and CD56 expression in persistent expansions of CD3+NKa+ lymphocytes are predictive for clonal T-cell receptor gene rearrangements. The Yorkshire Leukaemia Group.

Phenotypic characteristics, and correlations between the expression of membrane NK-associated (NKa) determinants (CD11b, CD16, CD56 and CD57) and T cell receptor (TCR) genotypic patterns, were examined in 25 patients with persistent (greater than 6 months) expansions of CD3+WT31+NKa+ (CD8+ and CD8dim+) lymphocytes. These studies showed that distinct NKa phenotypic profiles were restricted to cases with rearranged TCR configurations and that clonal CD3+NKa+ components could be predicted in most cases by assessing relationships between membrane CD16 and CD56 expression. For all normal NKa subpopulations, there was a high correlation (P less than 0.0001; n = 31) between the expression of these two membrane determinants. Markedly increased CD16 expression by CD3+NKa+ cells, in relation to CD56 (i.e. a high CD16:CD56 ratio), was found exclusively in cases with rearranged TCR (13/16 cases); 2/3 of the remaining cases showing significantly reduced CD16:CD56 ratios and high (greater than 2.0) CD3+CD56+ absolute numbers. In contrast, 7/9 of the germline TCR cases had a normal CD16:CD56 ratio and 2/9 a decreased ratio with low (less than 1.0) CD3+CD56+ absolute numbers. A high ratio of CD16:CD56 expression by CD3+NKa+ lymphocytes was therefore informative for 82% of TCR rearrangements in this series; and analysis of CD16 and CD56 expression was predictive for germline and rearranged TCR configurations in 24/25 persistent CD3+NKa+ expansions.

Adult↗
PubMed1991

Utilization of DNA recombination for the two-step replacement of growth factor sequences in the vaccinia virus genome.

An efficient procedure for the generation of sequence-specific alterations of the vaccinia virus genome was demonstrated. Homologous DNA recombination within cells infected with vaccinia virus was used for the deletion or replacement of promoter sequences of the viral growth factor gene by a procedure comparable to transplacement in Saccharomyces cerevisiae. This DNA replacement procedure can potentially be used to generate any sequence alteration within the vaccinia virus genome. Deletion of growth factor promoter sequences resulted in a dramatic reduction in growth factor gene transcription and protein synthesis. Replacement of growth factor promoter sequences with promoter sequences of the strong constitutive 40-kDa gene resulted in an increase in gene transcription and protein synthesis and an altered temporal pattern of expression. Virus containing mutations in the growth factor gene demonstrated different plaque morphologies on cell culture monolayers.

Base Sequence↗
PubMed1990

The seroepidemiology of human herpesvirus-6 (HHV-6) from a case-control study of leukaemia and lymphoma.

Sera from an epidemiological case-control study of leukaemias and lymphomas conducted between 1980 and 1986 were examined for reactivity to human herpesvirus-6 (HHV-6) by an indirect immunofluorescence assay. Statistical analyses of the data revealed higher HHV-6 seroprevalence and antibody titres in the cases, particularly evident in the disease subtypes acute myeloid leukaemia, Hodgkin's disease (HD), and low-grade non-Hodgkin's lymphoma. Within the control group alone, HHV-6 seroprevalence was placed at 55% at a serum dilution of 1:40. The controls also displayed higher seropositivity in females as compared with males. Further analyses suggest an association of increased HHV-6 seropositivity and geometric mean titre ratio with HD among young adults lacking social contact in the family group. This finding might indicate late exposure to HHV-6 in such persons and could possibly signify late exposure to a number of viruses, including those hypothesized as playing a role in the aetiology of HD. Previous reports have nominated Epstein-Barr virus as a possible candidate. Our results suggest that HHV-6 should be included in further investigations of the aetiology of HD.

Antibodies, Viral↗
PubMed1990

Effect of warfarin on plasma concentrations of vitamin K dependent coagulation factors in patients with stable control and monitored compliance.

There is a discrepancy in the results of reported studies of levels of vitamin K dependent coagulation factors in patients on warfarin therapy. This may have arisen partly because of the problem of assuring compliance with therapy in outpatients. The plasma concentrations of the vitamin K dependent clotting factors II, VII, IX and X were studied in 23 outpatients whose adherence to prescribed warfarin therapy was determined using a pharmacological indicator of compliance. In these patients, who were shown to have consistently good compliance and stable anticoagulant control over a period of 3-6 months, the activities in plasma of the four coagulation factors were not equally suppressed. Factor IX levels were significantly greater than those of factor VII (P less than 0.0001) which in turn were significantly greater than the levels of factor II (P less than 0.0001) or factor X (P less than 0.0001). There was no significant difference between the levels of factors II and X which were depressed to a similar extent. The proportion of variability of the International Normalized Ratio (INR) explained by linear regression was 51-77% and a model was derived to predict the INR from the mean of the levels of the four clotting factors. The concentrations of the coagulation factors II, VII, IX and X are likely to be highly dependent on the degree of compliance with warfarin therapy which should be taken into account when investigating the behaviour of these factors.

Aged↗
PubMed1990

Relationships between 2H4 (CD45RA) and UCHL1 (CD45RO) expression by normal blood CD4+CD8-, CD4-CD8+, CD4-CD8dim+, CD3+CD4-CD8- and CD3-CD4-CD8- lymphocytes.

We characterized and established relationships between the expression of membrane 2H4 (CD45RA) and UCHL1 (CD45RO) by enriched lymphocyte fractions prepared by selective immunomagnetic depletion of monoclonal antibody-defined populations. Cell fractions analysed in this study could be divided into two broad groups according to the presence (CD3+CD4+CD8-, CD3+CD4-CD8+, CD3+CD4-CD8dim+ and CD3+CD4-CD8-) or absence (CD3-CD4-CD8dim+ and CD3-CD4-CD8-) of the CD3 antigen. Preliminary studies confirmed a reciprocal relationship for CD45RA and CD45RO expression by major lymphoid components and further showed that the level or intensity of membrane 2H4 staining (2H4+, 2H4int and 2H4-) could be directly related to UCHL1 expression. As a reflection of their differential functions, the various CD3+ populations examined showed much greater heterogeneity in 2H4 and UCHL1 expression. CD3+CD4+CD8- cells generally showed significant proportions of 2H4+, 2H4int and 2H4- components, whereas the CD3+CD4-CD8+ population was characterized by a predominance of 2H4+ cells. The results of this current investigation further suggested a higher proportion of dual-positive (2H4+UCHL1+) cells and a much greater degree of inter-individual variation than previously suspected. In contrast to CD3+ lymphocytes, natural killer (NK) associated CD3-CD4-CD8dim+ and CD3-CD4-CD8- populations were mostly 2H4+ with only minor 2H4int components and very low expression of UCHL1. An additional observation of note was that the proportions of 2H4+ and 2H4- cells comprising the CD4+CD8- fraction in any given individual was highly correlated (P = 0.002) with the distributions of 2H4+ and 2H4- components within the CD4-CD8+ fraction. This suggests the possible existence of a common control mechanism for the acquisition of immunological memory by distinct lymphocyte populations and further indicates that individual variations in the distribution of 2H4/UCHL1 lymphocyte subpopulations may be a direct consequence of 'immunological experience' rather than age alone.

Antibodies, Monoclonal↗
PubMed1990

Flow cytometric analysis of membrane CD11b, CD11c and CD14 expression in acute myeloid leukaemia: relationships with monocytic subtypes and the concept of relative antigen expression.

Blast cells from 26 cases of acute myeloid leukaemia (AML) were examined, by single and "two-colour" flow cytometry, for relationships between membrane CD11b (monoclonal antibody OKM1), CD11c (KB90) and CD14 (Leu-M3). Increased expression of all three determinants was associated with myelomonocytic leukaemias, with their relative diagnostic value in discriminating monocytic (M4 and M5) from non-monocytic (M1, M2 and M3) subtypes being CD14 greater than CD11c greater than CD11b. However, the results also indicated, because of the heterogenous expression of CD11c in particular, and to a lesser extent CD11b, that the patterns or histograms of fluorescent staining were potentially more informative than an empirical subdivision of blasts into positive and negative subpopulations. In addition, analysis of phenotypic correlations by simultaneous two-colour fluorescence showed that the expression of CD11b and CD11c determinants by leukaemic myeloid blasts was highly correlated, in contrast to the expression of CD14 and CD11c which were relatively independent. Consequently, CD11c+ myeloid blasts almost always coexpressed CD11b whereas CD14+ cases of AML often comprised CD14+ CD11c+ and CD14+ CD11c- subpopulations. It is concluded from these observations that CD11c immunophenotyping is a useful supplementary investigation, particularly in CD14- cases of myelomonocytic leukaemia. However, it is also apparent that the presence of membrane CD11c per se is not lineage-specific and that the level of expression is perhaps a more discriminatory factor.

Antibodies, Monoclonal↗