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Biomedical subjects

D McCarthy

Publications and source records attributed to D McCarthy.

At least 19 recordsLinked to original sources
PubMed1992

Expression of functional antigens on neutrophils. Effects of preparation.

We have studied how different conditions of cell labelling and isolation affect the expression of five functional antigens on neutrophils from healthy subjects. Fluorescein isothiocyanate conjugated (FITC) antisera specific for the C3bi receptor CR3 (CD11b), aminopeptidase N (CD13), the LPS:LPS binding protein receptor (CD14) and the receptors for human IgG (Fc gamma RII CDw32 and Fc gamma RIII CD16) were incubated with (i) unfixed whole blood at 4 degrees C and at room temperature (RT, approximately 20 degrees C), and the leukocytes prepared for analysis using the Coulter Q-Prep system, (ii) leukocytes which had obtained following the removal of erythrocytes from whole blood by dextran sedimentation and which had been washed or left unwashed at RT, and (iii) leukocytes which had been prepared from whole blood that had been formaldehyde fixed immediately following venesection. The amount of fluorescence associated with the cells was determined by flow cytometry. The expression of CD14 was low under all conditions. However the expression of CD11b, CD16 and CDw32 was significantly higher (p less than 0.05) on neutrophils obtained by dextran sedimentation (n = 4) than on cells which had been fixed with formaldehyde ex vivo; the increase in expression was even greater if the cells had been washed. In contrast, the expression of CD13 on formaldehyde fixed cells was higher than on cells which had been labelled at 4 degrees C or at room temperature and was similar to or slightly lower than that on cells obtained by dextran sedimentation. Increasing the time between 10 and 60 min for which the whole blood was incubated with antisera at RT or at 4 degrees C, resulted in progressive increases in the expression of CD11b and CD13. When neutrophils which had been obtained by dextran sedimentation were incubated with unlabelled antibodies to CD16 or CDw32 and FITC labelled antibodies to CD11b there was a marked increase in the expression of CD11b. Altogether these findings indicate that the analysis of functional molecules on neutrophils (which may be rapidly up-regulated during activation) should be performed under clearly defined and controlled conditions. Dual fluorescence studies may, in some circumstances, produce misleading results.

Analysis of Variance↗
PubMed1992

A method of preparing blood leucocytes for flow cytometry which prevents upregulation of leucocyte integrins.

LeuCAM (CD11/CD18) cell-surface antigens are easily upregulated on cell manipulation ex vivo. A procedure for preparing leucocytes, in which human blood is immediately treated ex vivo with buffered formaldehyde and then the erythrocytes and platelets are removed by lysis and differential centrifugation, has been successfully applied to the analysis of LeuCAM antigen expression by flow cytometry. We show that the increased expression of monocyte CD11/CD18, which occurs when mononuclear leucocytes are separated by a standard Lymphoprep density gradient separation, can be avoided if cells are fixed immediately. Following this fixation polymorphs are unable to upregulate CD11/CD18 in response to fMLP stimulation in vitro. The technique produces lymphocyte, polymorph and monocyte populations that can be clearly defined on the basis of forward scatter and side scatter, and preserves the expression of various surface antigens; the percentages of gated lymphocytes expressing CD3, CD4, and CD8 were similar to those obtained using a commercial fixing and lysis solution. The processing does not render cells permeable to antibodies, as evidenced by our failure to stain cells with antibodies to intracellular antigens. We believed the method to be useful for measuring CD11/CD18 expression on blood leucocytes from normal or pathological specimens and to have application to the measurement of other cells surface antigens which may also be upregulated by the separation procedures.

Antigens, CD↗
PubMed1992

rec-2-dependent phage recombination in Haemophilus influenzae.

The genetic transformation mutant Rd(DB117)rec- has a pleiotropic phenotype that includes reduced levels of phage recombination. Physical mapping experiments showed that this strain has a 78.5-kbp insertion in the rec-2 gene. The rec-2 dependence of phage recombination was reexamined to determine whether the defective phenotype in Rd(DB117)rec- was due to the simple disruption of the rec-2 gene or whether trans-acting factors from the inserted DNA were responsible. Analysis of strains with transposon insertions in the rec-2 gene showed that they were also defective for phage recombination. Therefore, the phage recombination defect was due solely to the disruption of the rec-2 gene. Strain KB6 is proficient for phage recombination but has a defect in genetic transformation resembling that of Rd(DB117)rec-. The transformation defect of KB6 could be complemented by the wild-type rec-2 gene, showing that the rec-2 contributions to genetic transformation and phage recombination were uncoupled in this strain. The rec-2-dependent phenotype of KB6 suggests that the rec-2 gene participates in genetic transformation and phage recombination in different ways.

Bacteriophages↗
PubMed1992

Leucocyte integrin and CR1 expression on peripheral blood leucocytes of patients with rheumatoid arthritis.

Expression of the leucocyte integrins (CD11a, b, c/CD18) and of CD35 (CR1) on leucocytes from the peripheral blood of patients with rheumatoid arthritis (RA) (n = 14) and control subjects (n = 12) was measured by flow cytometry using a rapid fixation and leucocyte preparation procedure. The mean (SE) percentages of lymphocytes expressing CD11a (RA 93.4 (1.7)%; controls 97.2 (1.8)%) and CD18 (RA 91.3 (2.3)%; controls 97.0 (2.6)%) were lower and the percentage of monocytes expressing CD11b (RA 86.9 (11.4)%; controls 78.4 (11.9)%) and CR1 (RA 62.6 (15.5)%; controls 36.6 (17.6)%) were higher in patients with RA than in controls. In addition, the mean fluorescence intensity of CD18 (RA 22.1 (2.3); controls 30.7 (2.5)) on lymphocytes was decreased and that of CD11b (RA 4.5 (0.8); controls 2.9 (0.9)) and CR1 (RA 2.4 (0.4); controls 1.5 (0.5)) on monocytes was increased in patients with RA compared with controls. The functional importance (if any) of the altered expression of the antigens on lymphocytes is not yet known. Altered expression on monocytes is consistent with activation within the circulation.

Adult↗
PubMed1991

Quantitation of solid-phase immunoassays using a fibre-optic reflectance spectrophotometer.

A reflectance measuring instrument (fibre-optic spectrometer linked to a personal computer) which will quantitate the results of immunoassays that result in the deposition of a coloured chromophore on a membrane or other similar solid phase (SP) is described and the reflectance spectra of some chromophores which are commonly produced or used in SP immunoassays are presented. The instrument has been used in conjunction with silver-enhanced SP-immunogold capture assays to determine the concentrations in serum of IgG and of IgM rheumatoid factor (RF) and the results have been compared with those obtained by rate nephelometry and latex agglutination assays, respectively. It is concluded that fibre-optic reflectance photometry is an accurate and useful means of quantitating SP immunoassays in which coloured chromophores are produced and that the results obtained from such immunoassays are comparable to those given by established procedures.

Agglutination Tests↗
PubMed1991

Biological and clinical effects of intravenous tumor necrosis factor-alpha administered three times weekly.

Tumor necrosis factor (TNF) is a cytokine with pleiotropic biological and antitumor effects in vitro and in mouse models. The immunological effects of the molecule as a single agent, however, have not been well studied clinically. We conducted a Phase I trial of TNF in 53 patients with advanced malignancies in order to determine the biological and clinical effects of TNF when administered as a 30-min i.v. infusion three times/week. Dose levels of TNF ranged from 5 to 275 micrograms/m2; doses of TNF were escalated between patient groups. The most common clinical toxicities of TNF consisted of rigors, anorexia, headache, and fatigue. Dose-limiting toxicity consisted of hypotension, fatigue, and nausea. Four patients treated at the maximally tolerated dose of 225 micrograms/m2 received dexamethasone to determine whether the toxicities of TNF could be ameliorated. No significant differences in hypotension or subjective symptomatology were observed in those patients receiving dexamethasone and those who did not or between injections in which dexamethasone was administered and when it was not. One patient with colorectal carcinoma treated with 50 micrograms/m2 had a partial response lasting about 9 months. Biological responses were evaluated in 8 patients treated at the maximally tolerated dose before therapy and 24 h afterward. TNF significantly (P less than 0.05 for all) enhanced serum beta 2-microglobulin, serum neopterin, and serum interleukin-2 receptor (Tac antigen) levels. Indoleamine 2,3-dioxygenase activity was also increased 24 h following the administration of TNF, although this increase was only of borderline statistical significance (P = 0.07). TNF did not enhance granulocyte bactericidal activity. The expression of cell surface proteins on monocytes, including HLA-DR, HLA-DQ, beta 2-microglobulin, and the Fc receptor, and serum interleukin-1 activity also were not significantly increased by the administration of TNF. Thus, in humans TNF caused biological response modulation with evidence of HLA Class I (beta 2-microglobulin) increase and T-cell (Tac antigen) and monocyte (neopterin) activation.

2',5'-Oligoadenylate Synthetase↗
PubMed1991

Nonsteroidal anti-inflammatory drugs and peptic ulcer disease.

Evidence has accumulated that nonsteroidal anti-inflammatory drugs (NSAIDs) cause clinically important gastroduodenal ulcers. The pathogenesis, which involves the impairment of mucosal resistance to injury in an acid-peptic environment, is multifactorial and controversial. Ulcers caused by NSAIDs can occur either in mucosa inflamed because of infection with Helicobacter pylori or in histologically normal mucosa. The use of these drugs has been linked to an unexpectedly high incidence of ulcer complications, and a history of peptic ulcer disease is common in such cases. Nonsteroidal anti-inflammatory drugs thus appear both to exacerbate an underlying peptic diathesis and to cause de novo ulcers. The association between the use of these drugs and ulcer complications is supported by ulcer prevalence data from cross-sectional studies, and by data from case-controlled and cohort studies, and from randomized, experimental trials. Drug-induced gastric ulcers have been prevented by misoprostol, but not by H2 blocker therapy. Several therapies have been reported to promote ulcer healing despite continued use of NSAIDs, but adequate controlled trials have not been done. Small gastric and duodenal ulcers readily heal, whereas larger gastric ulcers require vigorous and prolonged therapy. The relative efficacies of various therapies in preventing ulcers, healing ulcers, or preventing complications remain to be established.

Anti-Inflammatory Agents, Non-Steroidal↗
PubMed1991

Successful cryopreservation of human bone marrow does not require a controlled-rate freezer.

We studied CFU-GM recovery from bone marrow samples frozen either in a machine for controlled-rate freezing or in a -70 degrees C freezer. We found no statistically significant difference between the two methods. Cooling-curve investigations also demonstrated that it was possible to obtain satisfactory cooling rates simply by manipulation of the shape and volume of the marrow and its container. Five patients received a total of six successful autografts for which their marrow cells were frozen without controlled-rate freezing. Thus it is not necessary to use a sophisticated machine in order to obtain satisfactory cryopreservation of human marrow stem cells.

Bone Marrow↗
PubMed1990

Improved plasmid shuttle vectors for Haemophilus influenzae and Escherichia coli.

In vitro deletion and transposon mutagenesis experiments were performed to localize the region essential for plasmid RSF0885 replication to a 1.7-kb sequence downstream from the beta-lactamase gene. This locus was named rep, for replication. Plasmid RSF0885 can replicate in both Haemophilus influenzae and Escherichia coli. Replication in E. coli depended on transcription run-off from the beta-lactamase promoter into the rep locus. Insufficient transcription into the rep locus could account for the instability of this plasmid in an E. coli background, a property which reduced its usefulness as a shuttle vector. Therefore, four improved shuttle vectors for H. influenzae and E. coli were constructed. They possess the Co1E1 replication origin for maintenance in E. coli and the plasmid RSF0885 rep locus for maintenance in H. influenzae. Together, they provide twelve unique restriction sites for cloning by insertional inactivation of drug-resistance genes.

DNA Replication↗
PubMed1990

Bone marrow transplantation for genetic disorders.

In 1967, a congenital disorder, severe combined immune deficiency disease, (SCID), was the first condition to be successfully corrected by bone marrow transplantation (BMT) from a histocompatible matched sibling donor. Since then the number of inherited disorders in which BMT has been used has been greatly extended. In preface, it should be stressed that BMT represents only one aspect of the management of genetic disorders which includes first and foremost detection and prevention by antenatal screening. Enzyme replacement treatment and the development of genetic engineering techniques to correct the underlying fault are being actively explored. However, reliable screening programmes are only feasible in a minority of disorders, of which thalassaemia is an example. Enzyme replacement treatment has been largely unsuccessful, and despite considerable advances in the understanding of gene regulation, at present BMT represents the only practice capable of correcting genetic disorders and improving the quality of life of affected individuals.

Bone Marrow Transplantation↗
PubMed1990

Interleukin-2 inhibits growth of fibroblasts derived from human bone marrow.

The adherent layer that forms in human bone marrow suspension cultures contains fibroblast colonies which arise from colony-forming cells (CFU-F). We have demonstrated that recombinant interleukin-2 (r-IL2) inhibits growth of CFU-F by preventing their entry into S phase. This inhibition is not mediated by gamma-interferon, nor by T lymphocytes, and can be abrogated by anti-IL2 receptor antibody. r-IL2 may influence haematopoiesis by its effect on bone marrow CFU-F.

Biological Factors↗
PubMed1989

Infusion of vincristine and doxorubicin with oral dexamethasone as first-line therapy for multiple myeloma.

32 previously untreated patients with multiple myeloma received vincristine, doxorubicin ('Adriamycin'), and dexamethasone (VAD) as first-line therapy. The overall response rate was 84%, with 28% of all patients entering complete remission. Response was rapid, with near-maximum response occurring after two courses of treatment and rapid improvement in bone-marrow function. Median response duration was 18 months and this seemed to be unaffected by initial prognostic criteria or by degree of remission achieved. The projected median survival was 44 months, with 75% of all patients and 83% of responders being alive at 2 years. Side-effects due to steroids were common, but there was only 1 treatment-related death. The high response rate and lack of toxicity offer an advantage over other forms of initial treatment, although other strategies will be necessary to prolong the duration of response.

Administration, Oral↗