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Biomedical subjects

D C Linch

Publications and source records attributed to D C Linch.

At least 163 records · Page 9Linked to original sources
PubMed1992

Granulocyte-macrophage colony-stimulating factor induces neutrophil adhesion to pulmonary vascular endothelium in vivo: role of beta 2 integrins.

Granulocyte-macrophage colony-stimulating factor (GM-CSF) causes upregulation of neutrophil surface CD11b/CD18 expression, and enhances the adhesion of neutrophils to cultured human endothelial cells in vitro. Systemic administration of GM-CSF results in a rapid, transient decrease in circulating phagocyte numbers. Using a nonhuman primate model (Cynomolgus), we provide histologic evidence that this transient leukopenia is associated with the margination of neutrophils in the pulmonary microcirculation. In four animals receiving 2 to 15 micrograms/kg recombinant human GM-CSF (rhGM-CSF), light microscopic sections of lung contained 36 +/- 8, 17 +/- 7, 21 +/- 6, and 15 +/- 8 (mean +/- SD, n = 20) neutrophils within a graticule grid, as compared with two control animals receiving saline injections whose lung sections contained 2.1 +/- 1.6 and 3.1 +/- 2.1 (mean +/- SD, n = 20) neutrophils within the same grid. Scanning electron microscopy shows activated leukocytes adherent to pulmonary vascular endothelium, but no morphologic evidence of endothelial damage, and no migration of cells into the extravascular space. Margination is associated with an increase in surface expression of CD11b/CD18 on circulating phagocytes, which could contribute to the adhesion to capillary endothelial cells, but CD11b/CD18 levels remain elevated even when demargination is complete. In vitro, monoclonal antibodies (MoAbs) to CD18 and CD11b were able to inhibit neutrophil aggregation and adhesion to endothelium. FMLP-induced neutrophil aggregation was inhibited by 39.8% +/- 11.5% and 44.8% +/- 12.3%, respectively, by MoAbs to CD18 and CD11b (P less than .0005, n = 4 for both); a similar effect was demonstrated on TPA-induced aggregation. MoAb CD18 reduced the adhesion of unstimulated neutrophils to endothelium by 44% (P less than .01, n = 7), and inhibited the amount of GM-CSF-stimulated adhesion by 74% (P less than .001, n = 7), while MoAb to CD11b produced a reduction of unstimulated neutrophil adhesion by 30%, and of GM-CSF-stimulated adhesion by 40% (P less than .01, n = 5, for both). However, when administered in vivo, MoAb CD18 produced only a small, albeit significant, amelioration of GM-CSF-induced margination in vivo, while MoAb CD11b was without effect. These results show that GM-CSF-induced transient leukopenia is associated with enhanced neutrophil adherence to pulmonary vascular endothelium, but suggest that the beta 2 leukocyte integrins CD11/CD18 play only a minor role in this process.

Antibodies, Monoclonal↗
PubMed1992

Down-regulation of human protein kinase C alpha is associated with terminal neutrophil differentiation.

We have established an RNase protection method to quantify the expression of mRNA for the human protein kinase C (PK-C) isoforms alpha, beta 1, beta 2, and gamma. This was used to investigate whether each isoform is differentially expressed during the differentiation of hematopoietic cells. Myeloid and lymphoid cells express PK-C alpha, beta 1, and beta 2 mRNAs in various proportions. PK-C gamma mRNA was detected in human brain, but not in hematopoietic cells. PK-C alpha mRNA decreases as HL-60 cells mature to a neutrophil phenotype in response to retinoic acid, but its abundance does not change during monocytic differentiation in response to vitamin D3. PK-C alpha mRNA and protein were undetectable in peripheral blood neutrophils, but are present in monocytes. The mRNAs for PK-C beta 1 and beta 2 isoforms increase during HL-60 differentiation and are expressed in both neutrophils and monocytes. Therefore, the PK-C alpha isoform is specifically down-regulated during human neutrophil terminal differentiation. These data suggest that mature neutrophil functions do not require the PK-C alpha isoform.

Calcitriol↗
PubMed1992

Interactions of granulocyte-macrophage colony-stimulating factor (CSF), granulocyte CSF, and tumor necrosis factor alpha in the priming of the neutrophil respiratory burst.

Exposure of neutrophils to a range of cytokines augments their response to subsequent agonist-induced activation of the respiratory burst. We have examined the effects of several of these factors, both singly and in combination, on the priming of f-met-leu-phe (FMLP) and complement C5a-stimulated neutrophil H2O2 production, using a whole blood flow cytometric assay designed to minimize artefactual activation. Both granulocyte-macrophage colony-stimulating factor (GM-CSF) and tumor necrosis factor alpha (TNF alpha) produced a similar degree of priming of the FMLP-stimulated burst in vitro (558% +/- 86%, n = 41, and 581% +/- 95%, n = 21, of the response seen with FMLP alone, respectively), but with markedly different kinetics (half-maximal response 20 minutes and 7 minutes, respectively). Preincubation with granulocyte colony-stimulating factor (G-CSF) alone caused only modest priming (202% +/- 39%, n = 14). Priming with cytokine combinations of the FMLP-stimulated burst showed that the combinations of G-CSF and TNF alpha and GM-CSF and TNF alpha are highly synergistic, with recruitment of neutrophils unresponsive to priming by single agents. Priming with the combination of GM-CSF and G-CSF was not significantly different to priming with GM-CSF alone. Similar results were obtained using C5a as the respiratory burst stimulus. Significant priming of the FMLP-stimulated respiratory burst was seen in vivo in patients receiving an infusion of GM-CSF (332% +/- 50% of preinfusion response to FMLP, P less than .005, n = 8). Priming was also seen in patients receiving G-CSF (152% +/- 58%, n = 5), although this did not reach conventional significance levels (.05 less than P less than .1). Although GM-CSF infusion caused priming in vivo, this was 48% less than predicted by preinfusion in vitro responses. This result was not due to inadequate GM-CSF levels as addition of further GM-CSF ex vivo did not correct the response. However, these neutrophils were still able to respond appropriately to ex vivo priming with TNF alpha, with a doubling in H2O2 production.

Complement C5a↗
PubMed1992

Recombinant human granulocyte-macrophage colony-stimulating factor after autologous bone marrow transplantation for malignant lymphoma: a British National Lymphoma Investigation double-blind, placebo-controlled trial.

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is active in enhancing the production of mature myeloid cells in vitro and several phase I/II clinical trials have suggested that its administration may accelerate neutrophil recovery after autologous bone marrow transplantation (ABMT). We have conducted a multicentre randomized double-blind placebo controlled trial in patients with poor prognosis malignant lymphoma receiving an identical high-dose combination chemotherapy regimen with ABMT. 61 patients were entered and 29 in each arm of the trial were evaluated. Treatment with GM-CSF did not affect the period of severe neutropenia (absolute neutrophil count (ANC) of < 0.1 x 10(9)/l) but accelerated recovery to an ANC of 0.5 x 10(9)/l (median 14 d v 20 d in controls, P = 0.001). There was no significant difference in platelet recovery between the groups (GM-CSF group platelet dependent for 25 d v control 19 d, P = NS). The number of positive blood cultures was similar in both groups (GM-CSF 14 v placebo 13) and there were no differences in days of fever > 37.5 degrees C (median 8 v 6) or days on parenteral antibiotics (11 v 10). Patients receiving GM-CSF had a median period of hospitalization following BMT of 24 d (control 25). No significant major toxicity attributable to GM-CSF administration was detected. We have confirmed in a randomized trial that GM-CSF accelerates neutrophil but not platelet recovery following ABMT. We were unable to demonstrate any accompanying changes in clinical outcome and believe that further trials are necessary to assess the clinical value of GM-CSF in BMT.

Adolescent↗
PubMed1992

Mini-BEAM followed by BEAM and ABMT for very poor risk Hodgkin's disease.

High dose chemotherapy and autologous bone marrow transplantation (ABMT) is an effective form of salvage therapy in patients with relapsed or resistant Hodgkin's disease. Patients with large tumour masses at the time of ABMT have a poorer prognosis and we have therefore administered intermediate dose BCNU, etoposide, cytarabine and melphalan (mini-BEAM) prior to high dose therapy with the same agents (BEAM) and ABMT in such patients. In addition we have used the same strategy in patients with bone marrow infiltration at the time of relapse in an attempt to clear the bone marrow for transplant. A total of 23 patients received mini-BEAM and 21 proceeded to BEAM and ABMT. Platelet engraftment was delayed compared to BEAM recipients who had not received mini-BEAM (P = 0.008) but there was only one procedure related death. Responses to BEAM and ABMT were not predicted by the response to mini-BEAM indicating a dose response effect at the upper end of the dose intensity spectrum. At 2 years, the overall survival and progression free survival are 61% and 46% respectively for this group of Hodgkin's patients with extremely poor prognosis.

Adult↗
PubMed1992

Differential effects of granulocyte- and granulocyte-macrophage colony-stimulating factors (G- and GM-CSF) on neutrophil adhesion in vitro and in vivo.

A direct comparison of granulocyte-macrophage colony-stimulating factor (GM-CSF) and granulocyte colony-stimulating factor (G-CSF) effects on neutrophil adhesiveness has been carried out. In vitro, GM-CSF and G-CSF upregulate neutrophil CD11b to a similar degree (to 227 +/- 69%, and 232 +/- 70% of control cells, respectively, p < 0.0005), but GM-CSF is more effective in downregulating neutrophil leucocyte adhesion molecule-1 (LAM-1), reducing levels to 33 +/- 4% (p < 0.0005), while G-CSF causes a fall to only 65 +/- 17% (p < 0.005) of control. The concentration of GM-CSF needed to achieve maximal activity is at least one log less than that of G-CSF. In vivo, both GM-CSF and G-CSF upregulate neutrophil CD11b (to 296 +/- 45% and 370 +/- 150%, respectively of baseline), but surface levels of LAM-1 on circulating cells are unchanged. GM-CSF increased neutrophil adhesion to cultured human endothelium in vitro (from 9.3 +/- 0.7% to 15.4 +/- 1.3%, p < 0.0005, n = 10), while G-CSF was without effect. In vivo, both GM-CSF and G-CSF produce a transient leucopenia, but recovery of peripheral counts occurs much earlier (by 60 minutes) with G-CSF, than with GM-CSF (only 50% of cells have demarginated at 120 min). GM-CSF appears to be greater proadhesive agonist for neutrophils than G-CSF.

Antigens, CD↗
PubMed1992

LOPP alternating with EVAP is superior to LOPP alone in the initial treatment of advanced Hodgkin's disease: results of a British National Lymphoma Investigation trial.

PURPOSE: The purpose of this randomized trial was to compare the efficacy of eight cycles of chlorambucil, vincristine, procarbazine, and prednisone (LOPP) with four cycles of LOPP that alternate with four cycles of etoposide, vinblastine, Adriamycin (doxorubicin; Familitalia Carlo Erba, Ltd, UK), and prednisone (EVAP) in patients with advanced Hodgkin's disease. PATIENTS AND METHODS: Between June 1983 and December 1989, 594 patients were entered onto the study. Of the 594, 295 patients were allocated to receive LOPP, and 299 were allocated to receive LOPP/EVAP. RESULTS: The complete remission (CR) rates were 57% and 64%, respectively, after initial chemotherapy (difference not significant [NS]), and 65% and 75%, respectively, after the subsequent administration of radiotherapy to residual masses (P less than .01). The procedure associated mortality in the LOPP and LOPP/EVAP arms was 1% and 3%, respectively. The actuarial CR relapse-free survival was significantly greater in the LOPP/EVAP arm (P less than .001) as was the overall survival (P less than .05). The CR relapse-free rate, disease-free survival (DFS) rate, and overall survival rate at 5 years were 52%, 32%, and 66%, respectively, in the LOPP arm, compared with 72%, 47%, and 75% in the LOPP/EVAP arm, respectively. CONCLUSION: These results indicate that LOPP and EVAP is superior to LOPP alone as initial treatment for advanced Hodgkin's disease.

Actuarial Analysis↗
PubMed1992

Assessment of X-chromosome inactivation patterns using the hypervariable probe M27 beta in normal hemopoietic cells and acute myeloid leukemic blasts.

The value of the hypervariable X-linked probe M27 beta for use in the analysis of X-chromosome inactivation patterns in normal blood and bone marrow cells and in the assessment of clonality in acute myeloid leukemia (AML) blast cells has been determined. By electrophoresing samples for 30 h, heterozygosity of the M27 beta locus was demonstrable in 324/415 females (78%) and this value could be increased to 93% by electrophoresing for 50 h. Determination of the X-chromosome inactivation patterns in blood and bone marrow samples from hematologically normal females was possible in approximately 90% of heterozygous individuals. The X-chromosome inactivation ratios obtained with M27 beta were comparable with phosphoglycerate kinase or hypoxanthine phosphoribosyl transferase in 46 individuals heterozygous for one of these genes in addition to M27 beta. The results obtained were closely correlated (r = 0.89). In 21 of 35 (60%) AML blasts, however, there was hypermethylation of the M27 beta locus and clonal analysis was not possible. Hypermethylation was not related to FAB type.

Bone Marrow Cells↗
PubMed1992

Fc gamma RII, but not erythropoietin or GM-CSF, mediates calcium mobilization in fetal hemopoietic blast cells.

A proportion of fetal liver hemopoietic blast cells express Fc gamma RII, and addition of the anti-Fc gamma RII monoclonal antibody CIKM5 induces a rise in calcium in these cells in suspension. Although these cells are thus capable of mobilizing intracellular calcium in response to surface receptor mediated events, neither granulocyte-macrophage colony-stimulating factor (GM-CSF) nor erythropoietin produced detectable changes in intracellular calcium ion concentration in these cells.

Antibodies, Monoclonal↗
PubMed1991

Differentiation-linked activation of the respiratory burst in a monocytic cell line (U937) via Fc gamma RII. A study of activation pathways and their regulation.

Activation of the respiratory burst in the monocytic cell line U937 by cross-linking human 40-kDa FcR for IgG (Fc gamma RII) with the IgG1 mAb, CIKM5, is dependent on the maturation state of the cell. Addition of anti-Fc gamma RII to undifferentiated cells does not activate the respiratory burst but differentiation with human rIFN-gamma (200 U/ml) for 13 to 15 days results in maximal stimulation by this agonist, with half-maximal responses in cells incubated for 10 to 12 days. During maturation the development of responsiveness to cross-linking Fc gamma RII occurs later than the development of responsiveness to the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (maximal responses at 7 to 9 days), or the chemotactic peptide FMLP (half-maximal responses at 7 to 9 days). The late development of maximal Fc gamma RII responses is not associated with either increased Fc gamma RII expression, enhanced calcium mobilization induced by anti-Fc gamma RII, changes in protein kinase C activity (PKC) or a switch in PKC isotype expression. Activation of the respiratory burst via Fc gamma RII may not be mediated by activation of PKC as the kinase inhibitors 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride and N-[2-(methylamino)ethyl]-5-isoquinolinesulfonamide dihydrochloride inhibited the Fc gamma RII response by less than 20% at concentrations which inhibit the 12-O-tetradecanoylphorbol-13-acetate-induced respiratory burst by more than 80%. IFN-gamma U937 cells did not metabolize incorporated arachidonate into eicosanoids when stimulated with anti-Fc gamma RII, suggesting that eicosanoids do not mediate activation of the respiratory burst, and this was confirmed by the lack of inhibition by the specific 5'-lipoxygenase and glutathione S-transferase inhibitor, piriprost, and the cyclo-oxygenase inhibitor, indomethacin. In addition there was no significant release of radiolabeled arachidonate in response to anti-Fc gamma RII. The response to anti-Fc gamma RII is inhibited by pertussis toxin, suggesting that signal transduction is via a GTP-binding protein. Agents that elevate intracellular cAMP increased the magnitude of the cAMP transients stimulated by anti-Fc gamma RII and also inhibited the respiratory burst. FMLP responses showed a similar pattern of sensitivity to this range of inhibitors, suggesting that both Fc gamma RII and FMLP receptor share common regulatory mechanisms. However, the termination of the respiratory burst activated via Fc gamma RII and FMLP receptor is independently regulated, in that after FMLP-induced activation there is no subsequent inhibition of the Fc gamma RII-mediated response and vice versa.

Antigens, Differentiation↗
PubMed1991

Lack of effect of granulocyte-macrophage and granulocyte colony-stimulating factors on cultured human endothelial cells.

The hematopoietic growth factors, granulocyte-macrophage colony-stimulating factor (GM-CSF) and granulocyte colony-stimulating factor (G-CSF), enhance the effector functions of mature myeloid cells, including the interaction with vascular endothelium. We examined the direct effect of recombinant human GM-CSF (rhGM-CSF) and recombinant human G-CSF (rhG-CSF) on the growth and function of cultured human umbilical vein endothelial cells (HUVEC). Endothelial cell growth supplement (ECGS) increased the proliferation of passaged and primary cells by 305% +/- 45% (mean +/- SEM, n = 5, P less than .01) over control cells at 4 days; GM-CSF and G-CSF had no effect. Endothelial cell procoagulant activity was increased after 4-hour incubation with recombinant interleukin-1 beta (IL-1 beta) 10 U/mL and recombinant tumor necrosis factor (TNF) 10 U/mL to 1,721% +/- 376% (n = 7, P less than .005) and 247% +/- 71% (n = 4) of control levels, respectively. gamma-Interferon (gamma-IFN) 50 U/mL had no direct effect of its own but was able to prime the response to IL-1 beta. There was no direct or priming effect of GM-CSF (1 ng to 1 microgram/mL) on the expression of procoagulant activity in endothelial cells. GM-CSF and G-CSF (1 ng/mL to 1 microgram/mL) had no effect on the expression of either tissue plasminogen activator (tPA) or plasminogen activator inhibitor-1 (PAI-1) by endothelial cells. The secretion of tPA by endothelial cells was increased, however, after 24-hour incubation with thrombin 4 U/mL (314% +/- 72% of control levels, n = 5, P less than .025). The production of PAI-1 was increased by TNF 200 U/mL (241% +/- 44% of control, n = 3, P less than .005), thrombin 4 U/mL (180% +/- 12% of control, n = 5, P less than .0005) and IL-1 beta 10 U/mL (275% +/- 44% of controls, n = 5, P less than .0005). In four experiments, endothelial cells showed no specific binding of 125I-GM-CSF, whereas peripheral blood (PB) neutrophils demonstrated the presence of 802 +/- 78 high-affinity receptors for GM-CSF. Thus, we found no effect of rhGM-CSF or rhG-CSF on the proliferation activities by these cells. These findings are in accordance with the lack of demonstrable receptors for GM-CSF on cultured HUVEC.

Cell Division↗
PubMed1991

An intracellular 50-kDa Fc gamma-binding protein is induced in human cells by alpha-IFN.

This report describes a novel intracellular Ig Fc-binding protein that is induced by alpha-IFN in a wide range of human cell types. The protein has a m.w. of 45,000 to 50,000 and it will bind human IgG but not IgM, IgA, or IgE. The IgG-binding specificity of this protein (designated Fc gamma BP) is similar to that of the Fc gamma RII receptor in that it binds only to mouse IgG1 and IgG2b but not IgG2a. It is an intracellular protein and is detectable by immunoprecipitation from [35S]methionine-labeled cells but it is undetectable on the cell surface. Fc gamma BP is a phosphoprotein, and the amount of phosphorylated Fc gamma BP increases after treatment with alpha-IFN. Although the function of this protein has yet to be determined, the induction of an Fc-binding protein by alpha-IFN suggests a role in the immune response, particularly during infection by viruses.

Antigens, Differentiation↗
PubMed1991

X-chromosome inactivation patterns using HPRT and PGK polymorphisms in haematologically normal and post-chemotherapy females.

Clonal analysis using X-chromosome inactivation patterns of the hypoxanthine phosphoribosyl transferase (HPRT) and phosphoglycerate kinase (PGK) genes has been used in the assessment of haematological malignancies. In many analyses normal haemopoietic tissue or constitutive DNA from each individual has not been readily available and it has been assumed that when the ratio of the percentage of the two alleles remaining after differential methylation enzyme digestion is greater than 3/1 then an oligoclonal/monoclonal population of cells is present. We report here in a study of 65 haematologically normal females informative for HPRT or PGK that 15 (23%) had a skewed Lyonization pattern with clonal analysis ratios greater than 3/1. No correlation was observed between age and the X-chromosome inactivation pattern to suggest the detection of pre-malignant clonal states. The incidence was similar in 23 females who had received conventional combination chemotherapy, with six women (26%) having a ratio greater than 3/1. In the majority of individuals therefore, conventional chemotherapy does not appear to alter the patient's X-chromosome inactivation pattern. These results indicate that caution must be exercised in interpreting the results of clonal analysis of any given individual.

Adolescent↗
PubMed1991

In vivo effects of macrophage colony-stimulating factor on human monocyte function.

Macrophage colony-stimulating factor (M-CSF) is reported to enhance a variety of functions of mature monocyte/macrophages in vitro. We have examined the effects of a 2 h intravenous infusion of M-CSF obtained from human urine (hM-CSF) on haematological parameters and selected monocyte functions. There was a rapid, small, but consistent reduction in Hb concentration (mean 6.5 +/- 2.3%, P less than 0.0005 by paired t test) by the completion of the hM-CSF infusion and small, transient falls in platelet, monocyte and neutrophil counts were noted in the 2 h following the end of the infusion. No effect on monocyte or neutrophil CD11b cellular adhesion molecule expression was detected. Exposure to hM-CSF in vivo did not directly stimulate the monocyte respiratory burst, but increased the percentage of monocytes responding to f-met-leu-phe from 9.8 +/- 2.5 to 16.6 +/- 4.2 (P less than 0.01). The number of candida ingested and degraded per 100 monocytes increased from 101 +/- 14 pre-infusion to 160 +/- 22 post-infusion (P less than 0.01). There was a rapid increase in the numbers of monocytes entering a skin window membrane from a mean of 226 +/- 71 pre-infusion to 1064 +/- 404 at the end of the infusion, with no effect on neutrophil migration. These data show that the administration of hM-CSF enhances several of the functions of peripheral blood monocytes in vivo, and this may be of benefit in the treatment of selected infections.

Adult↗