Polymorphonuclear leucocytes consume oxygen in sputum from chronic Pseudomonas aeruginosa pneumonia in cystic fibrosis.

BACKGROUND: Chronic lung infection with Pseudomonas aeruginosa is the most severe complication for patients with cystic fibrosis (CF). This infection is characterised by endobronchial mucoid biofilms surrounded by numerous polymorphonuclear leucocytes (PMNs). The mucoid phenotype offers protection against the PMNs, which are in general assumed to mount an active respiratory burst leading to lung tissue deterioration. An ongoing respiratory burst by the PMNs has, however, not been demonstrated previously in endobronchial secretions from chronically infected patients with CF. OBJECTIVE: Based on the accumulating evidence for depletion of molecular oxygen (O(2)) in the mucus in infected CF bronchi, it was hypothesised that the O(2) depletion in the mucus in infected CF bronchi may be accelerated by the respiratory burst of the PMNs due to the reduction of O(2) to the superoxide anion (O(-)(2)) by the phagocyte NADPH oxidase (Phox). METHODS: Methods were established to isolate the O(2) consumption by the respiratory burst from aerobic respiration in freshly expectorated sputum from chronically infected patients with CF. RESULTS: Inhibition of the Phox with diphenylene iodonium (DPI) delayed O(2) depletion, nearly abolished staining of O(-)(2)-producing PMNs with hydroethidine and inhibited the rapid luminol-enhanced chemiluminescence in sputum. Furthermore, the total O(2) consumption was correlated to the concentration of PMNs in the sputum samples. CONCLUSION: The results demonstrate that CF sputum contains PMNs with an active consumption of O(2) for O(-)(2) production and suggest that the respiratory burst is ongoing and causes accelerated O(2) depletion due to formation of O(-)(2) in the lungs of chronically infected patients with CF.

No correlation between CD73 expression and ecto-5'-nucleotidase activity on blood mononuclear cells in vitro. Evidence of CD73 (ecto-5'-nucleotidase) on blood mononuclear cells with distinct antigenic properties.

Monoclonal antibodies against ecto-5'-nucleotidase (ecto-5'-NT) have been clustered to CD73. The aim of the present study was to elucidate whether specific ecto-5'-NT activity on blood mononuclear cells (BMC) was correlated with CD73 expression measured by flow cytometry. During culture of CD73-negative lymphocytes the percentage of cells with ecto-5'-NT activity increased without there being a comparable increase in CD73 expression. After 2 days' culture, 9% (3-16, median and range given) demonstrated ecto-5'-NT activity, whereas CD73-positive cells comprised only 1.6% (0.0-3.0). These results show the existence of lymphocytes with ecto-5'-NT activity but without expression of CD73. The ecto-5'-NT activity increased significantly (p<0.02) during culture of unseparated BMC, whereas the number of anti-CD73 binding sites did not change. As a consequence. the number of anti-CD73 binding sites per U ecto-5'-NT activity decreased during culture. Exposure to interleukin-4 or prostaglandin E2 changed the enzymatic activity but not the number of anti-CD73 binding sites. Treatment of BMC with phosphatidylinositol-specific phospholipase-C released 57% (51%-75%) of the ecto-5'-NT activity into the supernatants, without a detectable decrease in CD73 expression. The fact that ecto-5'-NT was removed from the supernatant after precipitation with anti-CD73 showed that the released ecto-5'-NT activity was due to enzymatic activity of CD73 molecules. The results of this study indicate that CD73 exists on BMC in isoforms with distinct capacities to bind mouse monoclonal anti-CD73.

Treatment of relapse after allogeneic BMT with donor leukocyte infusions in 16 patients.

Sixteen patients with relapse after allogeneic BMT were treated with donor leukocyte infusions (DLI) from the original donor. The diagnoses at relapse were: CML in chronic phase (CP) (two patients), CML in accelerated phase (AP) (four patients), AML (four patients), MDS (one patient), ALL (four patients) and relapse of Hodgkin's disease (one patient). The patients received a mean of 5.2 x 10(8) leukocytes/kg with a range of 1.4-12.3 x 10(8) leukocytes/kg. Six patients obtained complete remission (CR), one with CML in CP, three with CML in AP, one MDS and one ALL. Partial remission (PR) was seen in three patients, one patient with CML in AP, one with AML and one with Hodgkin's disease. Seven patients had no response (NR) to the infusions, including one patient with CML in CP transplanted with a syngeneic donor. Four patients developed marrow hypoplasia after DLI (three CR and one PR) and two patients (ALL with CR and MDS with CR) were hypoplastic at relapse and marrow hypoplasia continued after DLI. GVHD occurred without GVL, but GVL only occurred in one patient with absence of GVHD. Eleven patients died of leukemia, six patients are alive. Three patients with CML are in CR 12, 12 and 32 months after DLI and one patient with ALL is in CR 15 months after DLI.

Elastolytic activity of human blood monocytes characterized by a new monoclonal antibody against human leucocyte elastase. Relationship to rheumatoid arthritis.

The leucocyte elastase of human blood monocytes was investigated by applying a new monoclonal antibody which did not block the enzyme activity against elastin. In a fixed population of mononuclear cells (MNC) and using fluorescence activated cell sorting (FACS), the human leucocyte elastase (HLE) antibody identified a subgroup of CD14+ cells which contained all the elastase activity and which could be blocked by a specific chloromethylketone elastase inhibitor. By anti-CD14 labelling the HLE positive cells were identified as monocytes and amounted to 88% of this cell type (median: range 72-96%). In a parallel study of patients with active rheumatoid arthritis (RA) and control donors the elastolytic capacity of PMA-stimulated live MNC was higher for RA patients than for controls (p less than 0.02). For control donors the number of HLE+ cells correlated well to the elastolysis in the same MNC sample (Spearman's rho:0.83, p less than 0.05); in the samples from RA patients no correlation was found between the normal number of HLE+ cells and the increased elastolysis (rho: 0.54, p greater than 0.20).

[Autologous stem cell transplantation. From bone marrow to selected blood stem cells: 100 consecutive procedures at a single center]. / Autolog stamcelletransplantation. Fra knoglemarv til oprensede blodstamceller: 100 konsekutive procedurer i et enkelt center.

One hundred consecutive autologous stem cell transplants are reported: Non-Hodgkin's lymphoma 51 cases, Hodgkin's disease 27 cases, acute leukaemia 14 cases, multiple myeloma seven cases and chronic myeloid leukaemia one case. Most patients were in their second or later remission. The overall three-year survival for all patients was 60% and the three-year disease-free survival was 50% for lymphoma patients and 30% for acute leukaemia patients. The dominant source of stem cells was bone marrow during 1993, but from 1994 it has been peripheral blood, now totalling 33 cases. There were 12 toxic deaths, all among patients who were heavily treated before bone marrow harvest and transplantation. The patients transplanted with blood stem cells had significantly shorter duration of pancytopenia, and hospital stay, but their disease-free survival was not longer than that of a comparable group of bone marrow transplanted patients. Six patients were transplanted with purified CD34+ cells (selected by avidity column (Ceprate (R)), and had duration of thrombocytopenia and hospital stay similar to the patients transplanted with unmanipulated blood stem cells, but slightly longer duration of neutropenia. We conclude that high-dose therapy with autologous stem cell transplantation in not too heavily pretreated patients is a safe procedure irrespective of the source of stem cells.

Natural killer cells lack ecto-5'-nucleotidase.

Ecto-5'-nucleotidase (ecto-5'-NUC, EC 3.1.3.5., CD73) is a plasma membrane enzyme, which catalyzes the hydrolytic dephosphorylation of purine nucleotides to the corresponding nucleosides so that they can pass through the plasma membrane. The distribution of ecto-5'-NUC is heterogeneous, but all blood mononuclear cell (BMC) subpopulations investigated until now have had ecto-5'-NUC activity. Purified natural killer (NK) cells were prepared by fluorescence-activated cell sorting of CD16-positive cells (purity 94%). In purified NK cells the ecto-5'-NUC activity was less than 1 U/10(6) as estimated by a radioisotope method. Using a cytochemical assay, the proportion of cells with activity was less than 1%. The most likely explanation is that NK cells have no ecto-5'-NUC activity and that the very small ecto-5'-NUC activity observed in this study was caused by contaminating non-NK cells. NK cells thus constitute the only BMC subset known without ecto-5'-NUC activity.

In vitro evaluation of a new filter for leucocyte depletion of platelet concentrate during component preparation.

A newly developed filter for prestorage leucocyte depletion of platelet concentrates (PC) was studied. The filter is designed for leucocyte depletion during the preparation of the pool PC from platelet rich buffy coats. In all the leucocyte depleted PC (LD-PC) leucocyte depletion was satisfactory. 19 of 20 units of LD-PC had a leucocyte content below 3 x 10(5) per PC and 1 contained 8 x 10(5) leucocytes. The standard PC contained 2.53 x 10(8) (0.87 x 10(8)-15.3 x 10(8); n = 20) leucocytes per PC (median and range). The quality of the LD-PC was evaluated by measuring platelet activation, platelet morphology, and pre- and poststorage pH. There were no differences in any of the parameters studied.

Interleukin 2 receptor expression by human blood lymphocytes after vaccination with pneumococcal polysaccharides.

Proliferative responses of unseparated peripheral blood mononuclear cells (PBMC) and blood T cells to recombinant interleukin 2 (rIL-2) were significantly increased 7-21 days after the vaccination with pneumococcal polysaccharides (PPS). In contrast, non-T cells expressed increased responsiveness to rIL-2 only on post-vaccination day 7. Analysis of the proliferative response to rIL-2 among lymphocyte subsets (CD4+Leu8+, CD4+Leu8-, CD8+Leu8+, CD8+Leu8-, CD20+) in cultures of unseparated PBMC revealed that the CD8+Leu8- T cells expressed increased responsiveness 7-14 days after vaccination, whereas neither CD4+ (Leu8+ and Leu8-) nor CD8+Leu8+ T cells showed significantly increased responsiveness after vaccination. The CD20+ B cells, like non-T cells, expressed increased responsiveness to rIL-27 days after the vaccination only. Expression of the 55 kD low-affinity interleukin 2 receptor (IL-2R, CD 25) on freshly isolated PBMC, as judged by direct fluorescence staining with a MoAb anti-55 kD chain, was low (less than 3%) and an increased expression of this receptor was not detected following vaccination. In contrast, binding of 125I-labelled IL-2 to freshly isolated PBMC increased following vaccination (day 7). Scatchard plot analysis revealed a modest increase in the expression of high-affinity IL-2R (Kd = 1-2 pM), whereas the increase in expression of the 75-kD, intermediate-affinity IL-2R (Kd = 300 pM) was more pronounced (from 195 to 295 (means) receptors per PBMC). It is concluded that, following vaccination with PPS increased IL-2R expression is induced on blood lymphocytes. These investigations suggest a role for T cells in the human immune response against PPS.

Monoclonal antibodies with neuroblastoma specificity: a flow cytometric analysis of cross-reactivity with CD34+ hematopoietic stem cells.

The aim of this study was to evaluate the specificity of a number of monoclonal antibodies (MAbs) used for immunological in vitro purging of stem cell grafts from neuroblastoma patients. The extent of cross-reactivity of 10 neuroblastoma-specific MAbs (NB-MAb) with CD34+ stem cells from 14 leukapheresis products was analyzed. The level of cross-reactivity was analyzed on a Coulter (Fullerton, CA) flow cytometer using biotinylated NB-MAbs. There was a marked difference in the reactivity of the ten NB-MAbs with CD34+ stem cells. The antibodies could be divided into three groups with increasing levels of cross reactivity. Four antibodies (126-4, 5.1 H11, UJ127.11, and 14.G2a) all reacted with median levels of less than 2% (range 0.0 to 5.4) of CD34+ stem cells (median of 14 patients). Another three antibodies reacted with a median of 3.1-4.1% of the stem cells (UJ13A, Ab390, and Ab459) but with a wide range (0.2 to 25.6). Finally, M340, HSAN 1.2, and antiThy-1 reacted with a median of 9-16% of the stem cells (range 0.6 to 51.5). We conclude that there is a significant variation in the proportion of CD34+ stem cells reacting with each of the ten neuroblastoma antibodies investigated in this study. Therefore, to avoid a significant loss of CD34+ cells from the stem cell product, we find it important to carefully consider which antibodies to use for immunomagnetic purging.

Effects of immunomodulators on ecto-5'-nucleotidase activity on blood mononuclear cells in vitro.

In this study the effects of immunomodulators on the ecto-5'-nucleotidase (ecto-5'-NT) activity on blood mononuclear cells (BMC) were examined in vitro. Interleukin-4 (IL-4) and interferon-gamma (IFN-gamma) decreased the level of ecto-5'-NT activity on BMC whereas prostaglandin E2 (PGE2) increased the ecto-5'-NT level. All three immunomodulators influenced the ecto-5'-NT activity of isolated monocytes whereas only IL-4 and PGE2 had an effect on the enzyme level on isolated lymphocytes. The effect was dependent upon protein synthesis. The effect was dose dependent: IL-4 was effective at concentrations down to 0.5 U/ml, IFN-gamma down to 40 U/ml and PGE2 at nanomolar concentrations. These data indicate that immunomodulators may also take part in the regulation of ecto-5'-NT activity on BMC in vivo. BMC from 7 patients with different immunodeficiency syndromes showed decreased ecto-5'-NT activity on freshly isolated cells. However, following culture ecto-5'-NT activity was increased above the level found on freshly isolated BMC from healthy persons. On BMC from 3 patients with hypogammaglobulinaemia, the effect of IL-4 on the level of ecto-5'-NT activity was identical to that found on BMC from healthy donors, whereas PGE2 increased ecto-5'-NT activity on BMC from only 1 of the 3 patients investigated. The decreased ecto-5'-NT activity of BMC from patients with immunodeficiency may thus be due to a defective regulation of ecto-5'-NT activity in vivo.

Decreased number of CD73 (ecto-5'-nucleotidase) molecules on lymphocytes from patients with primary immunoglobulin deficiencies. Correlation between number of CD73 molecules and T-lymphocyte function in vitro.

CD73 is a bifunctional glycosyl phosphatidylinositol anchored leucocyte differentiation antigen which has specific ecto-5'-nucleotidase (ecto-5'-NT) activity and is an accessory T-lymphocyte activation molecule. The aim of the present study was to investigate the CD73 expression on blood mononuclear cells (BMC) from a group of patients with primary immunoglobulin deficiency (IGD). This group of patients had both significantly decreased levels of ecto-5'-NT on BMC (P = 0.002) and decreased numbers of CD73 molecules per CD73+ lymphocyte (P = 0.01). Five of the 10 patients had a decreased percentage of CD73+ lymphocytes. Among B-lymphocytes the patients had normal percentages of CD73+ cells but four of the 10 patients had numbers of CD73 molecules per CD73+ B-lymphocyte below the normal range. Among CD4-lymphocytes three out of 10 patients had percentages of CD73+ below the normal range and four out of 10 patients had decreased percentages of CD73+ CD8-lymphocytes. Significant correlations were found between in vitro proliferative responses to mitogens and the number of CD73 molecules per CD73+ lymphocyte (rs = 0.60, P < 0.01) and per CD73+ CD8-lymphocyte (rs = 0.64, P < 0.02). In addition, a positive correlation was found between ability to proliferate and level of ecto-5'-NT on BMC (rs = 0.53, P < 0.05). Furthermore the ability of BMC to synthesize ecto-5'-NT was studied. During 2 days culture ecto-5'-NT activity increased markedly on BMC from both patients and healthy donors. The level of activity on BMC from all patients attained levels higher than on freshly isolated BMC from healthy donors. This shows that the decreased levels of ecto-5'-NT found on freshly isolated BMC from patients with IGD is due to defective regulation of the enzyme activity in vivo.

Interleukin 1-induced down-regulation of antibody binding to CD4 molecules on human lymphocytes.

Interleukin 1 (IL-1) is involved in the early activation of T lymphocytes. The CD4 antigen, described as a phenotypic marker of helper T cells, is also important in early T-cell activation by its ability to bind to MHC class II molecules on antigen-presenting cells, and to transmit positive (and negative) signals to the cells. We observed that purified human monocyte IL-1 as well as recombinant IL-1 alpha and IL-1 beta selectively decreased the binding of monoclonal antibodies to CD4 on the surface of otherwise unstimulated blood T cells, in contrast to prestimulated and continuously grown CD4+ cells. Under optimal growth conditions, the initial reduction in antibody binding to CD4 was followed by an apparent re-expression of the CD4 antigen even in the presence of high concentrations of IL-1. This re-expression did not occur if the cells were cultured at 4 degrees C, or after treatment with actinomycin D or cytochalasin B, indicating that protein synthesis and intact microfilament function were essential for re-expression of CD4 binding. The mechanism by which CD4 molecules are physically and/or functionally modulated by IL-1 is unclear.

Lack of immunomodulating effect of disulfiram on HIV positive patients.

Disulfiram (Antabuse (R)) is metabolized to two molecules of diethyldithiocarbamate, which has been reported to be an immunomodulating agent. In a double blind trial, 15 HIV antibody positive homosexual men were given daily doses of 100 mg or 400 mg of disulfiram or placebo, for 4 weeks. All had a CD4-count below 500 X 10(6)/l and/or a pokeweed mitogen response in a lymphocyte proliferation assay less than 50% of normal controls. None suffered from opportunistic infections. No significant effect of disulfiram on immunological, haematological, biochemical or clinical variables was observed in this short-term trial.

Complement receptor type 1 (CR1, CD35) expression on peripheral T lymphocytes: both CD4- and CD8-positive cells express CR1.

T lymphocytes from 42 healthy blood donors were examined for expression of CR1 on the cell surface using a fluorescence-activated cell sorter. The fraction of CR1-positive T lymphocytes varied in the range from 1 to 8% (median 2.4%). The CR1-positive T lymphocytes constituted a homogeneous group of cells regarding both size and granulation; they seemed to be larger and more granulated than the average T lymphocytes. The CR1-expressing T lymphocytes were found both in the CD4- and in the CD8-positive subpopulations of T lymphocytes, and although the CD4-positive cells expressed CR1 with a slightly higher percentage than did the CD8-positive cells, this difference was not significant. The number of CR1-positive T lymphocytes did not correlate with the level of CR1 on erythrocytes. The presence of CR1 on a small T lymphocyte population suggests that CR1 on T lymphocytes might play a role in the immune regulation.

Recruitment of CD34+ cells during large-volume leukapheresis.

Mobilized peripheral blood stem and progenitor cells (PBPCs) are increasingly used to restore hematopoiesis after myeloablative treatment. To obtain a sufficient number of CD34(+) cells, many studies have focused on the improvement of the collection technique during the leukapheresis procedure (LP), and so-called large-volume leukapheresis (LVL) procedures have been developed. Such procedures can be performed by extending the duration of the LP and/or by increasing the inlet flow rate. However, no previous studies have compared the efficiency of these procedures. In the present study, we compared the kinetics of PBPCs recruitment (including CD34(+) cell subsets), the PBPCs yield, and the collection efficiency as well as the overall feasibility of the procedures during a single LVL performed by standard (group I) (median 85 ml/min; range 50-97 ml/min) and high inlet flow rates (group II) (median 130 ml/min; range 110-150 ml/min). Seven patients with hematological malignancies were enrolled and allocated to each group. The patients' blood volumes (BV) were processed four times. The apheresis product (AP) was collected in four separate bags, which were changed every time one BV had been processed. The CD34(+) cell number and CD34(+) cell subsets were assessed in the four collection bags and in peripheral blood (PB) before every time one BV had been processed and after the leukapheresis. The CD34(+) cell yield exceeded the pre-apheresis CD34(+) cell number per ml BV in 6 out of 7 patients in group I and in 3 out of 7 patients in group II. In group II, the recruitment of CD34(+) cells from the bone marrow (BM) to PB starts in the second collection period--as early as 30-60 min after initiating the procedure. No exhaustion in the recruitment was observed in the two groups for at least 5 h during the leukapheresis, and all CD34(+) cell subsets were recruited at a steady rate. However, the collection efficiency in group II was only half of that in group I. In addition, we experienced many technical problems during the leukapheresis in group II. Thus, in 4 out of 7 patients in this group, it was not possible to perform the maximal inlet flow rate because of catheter problems. In conclusion, due to the technical problems associated with the high inlet flow rate procedure and the fact that the relative number of CD34(+) cells harvested and recruited during the leukapheresis was higher in group I than II and, also reflected an approximately two-fold higher collection efficiency, we recommend that LVL be performed by standard inlet flow rate.

Extracellular accumulation of bioactive substances during preparation and storage of various platelet concentrates.

BACKGROUND: Side effects of platelet transfusion may be associated with infusion of bioactive substances. We therefore studied extracellular accumulation of histamine, plasminogen activator inhibitor (PAI)-1, vascular endothelial growth factor (VEGF), and interleukin (IL)-6 during preparation and storage of various platelet concentrates. METHODS: Twenty buffy-coat-derived platelet pools (BCPC) were prepared and stored in platelet additive solutions (PAS). Twelve apheresis platelet (APC) units were prepared using the COBE Spectra LRS, and 14 were prepared using the Fenwal Amicus Separator. After preparation half of the content was drawn from each APC unit. The normal ranges of the substances were determined in plasma from all donors, and the extracellular concentrations of the substances were determined in supernatants collected on days 0, 1, 3, 5, and 7 of storage from all platelet preparations. RESULTS: The platelet counts were not significantly different in BCPC units and APC units. The BCPC units had a significantly higher white cell count than the APC units (P < 0.0001), but the count was significantly higher in the Amicus APC units than in the COBE APC units (P < 0.0001). The extracellular histamine concentration was significantly (P < 0.001) increased in BCPC units after preparation and without further increase during storage, while there was no accumulation of histamine in APC units. After preparation the PAI-1 concentration was significantly (P < 0.02) higher in BCPC units than in APC units, but during storage PAI-1 increased significantly (P < 0.05) more in APC units than in BCPC units. Similarly, VEGF concentration was significantly (P < 0.05) higher in BCPC units than in APC units after preparation. During storage, however, VEGF increased more in BCPC units compared with COBE Spectra APC units (P < 0.05), but compared with Amicus Separator APC units only for the first 3 days of storage. At days 5 and 7 of storage the VEGF concentration was significantly higher in the Amicus APC units than in the COBE APC units (P < 0.05). IL-6 was not detectable in any of the concentrates after preparation or during storage. CONCLUSION: Platelet concentrates prepared by the apheresis method may contain less white cell derived bioactive substances than platelet concentrates prepared by the buffy-coat method. However, a substantial storage time dependent platelet derived bioactive substance accumulation takes place in all platelet concentrates tested, presumably due to platelet disintegration.

Natural killer cell activity in peripheral blood of highly trained and untrained persons.

Natural killer (NK) cell activity and concentration of CD16+ cells (NK cells) and CD20+ cells (monocytes) in peripheral blood were measured in highly trained racing cyclists and in age- and sex-matched untrained controls. Median NK cell activity was 38.1% (range 20.0%-57.1%) in trained vs 30.3% (range 19.7%-43.1%) in untrained (P = 0.008). Median %CD16+ cells was 17% (range 7%-33%) in trained vs 11% (3%-29%) in untrained (P = 0.007). Indomethacin in vitro enhanced the NK cell activity in both groups. There was, however, no significant difference between the NK cell activity in trained and untrained after exposure to indomethacin in vitro. Indomethacin-enhanced NK cell activity was 45.9% (range 24.4%-67.5%) in trained and 40.0% (range 23.9%-68.5%) in untrained (P = 0.138). Mean %CD14+ cells was 8.3% (range 2%-15%) in trained vs 3.8% (2%-8%) in untrained (P less than 0.0001). The increased NK cell function thus demonstrated in highly trained persons might result in better resistance against infectious disease.

Effect of physical exercise on blood mononuclear cell subpopulations and in vitro proliferative responses.

The present study was designed to examine the effect of physical exercise on subsets and proliferative responses of blood mononuclear cells. Sixteen young, healthy volunteers underwent 60 min of bicycle exercise at 75% of maximal oxygen uptake (VO2max). After an interval of at least 1 week, six of the subjects underwent a 60-min back muscle training period at up to 30% of VO2max. Blood samples were collected before and during the last minutes of exercise, as well as 2 and 24 h later. Blood mononuclear cell (BMNC) subpopulations were determined and the proliferative responses after incubation with phytohaemagglutinin (PHA) or purified derivative of tuberculin (PPD), were quantified by [3H]thymidine incorporation. During bicycle exercise the relative blood concentration of T cells (CD3+ cells) declined, mainly due to a fall in T helper cells (CD4+ cells). The natural killer (NK) cell subset (CD16+ cells) increased during work, but reverted after; the monocytes (CD14+ cells) increased 2 h after work, whereas the B-cell subset (CD20+ cells) did not change. BMNC subsets were not significantly changed by back muscle exercise. The PHA-induced proliferative response decreased during bicycle exercise, whereas the PPD-induced response did not change. No significant changes occurred during back muscle exercise. Investigation of subgroups after incubation with [3H]thymidine showed that the proliferative response per CD4+ cell did not change in relation to exercise, but the contribution of the CD4+ subgroup to proliferation declined during bicycle exercise due to the decreased proportion of CD4+ cells. The suppression of the PHA response during bicycle exercise can be explained in part by a relative fall in CD4+ cells. The pool sizes of BMNC subfraction may be elicited by increased catecholamine and cortisol levels.

Inhibition of human lymphocyte proliferation and cleavage of interleukin-2 by Pseudomonas aeruginosa proteases.

This study was undertaken to determine the effect of Pseudomonas aeruginosa alkaline protease (AP) and elastase (ELA) on human lymphocyte function. AP at 50 micrograms/ml and ELA at 12 micrograms/ml caused a 50% inhibition of phytohemagglutinin-induced proliferation. There was no difference in the effect of proteases on CD4- and CD8-positive cells. To determine the effect of proteases on interleukin-2 (IL-2)-induced cell proliferation, the proteases and IL-2 were added to the IL-2-dependent CTLL-2 cell line. AP and ELA inhibited the proliferation of these cells. When IL-2 was added in excess, the inhibition was partly reversed. ELA at 10 micrograms/ml cleaved IL-2, as judged by size chromatography of a reaction mixture containing 125I-labeled IL-2 and the proteases. The ELA-digested IL-2 exhibited a reduced binding capacity to IL-2 receptors on the lymphocytes. Furthermore, treatment of phytohemagglutinin-stimulated lymphocytes with AP and ELA resulted in inhibition of binding of intact IL-2 to IL-2 receptors on the stimulated lymphocytes. These results indicated that P. aeruginosa-derived enzymes are able to interfere with human lymphocyte function in vitro and that this effect might be due to cleavage of IL-2.