Effect of priming polymorphonuclear leukocytes with cytokines (granulocyte-macrophage colony-stimulating factor [GM-CSF] and G-CSF) on the host resistance to Streptococcus pneumoniae in chinchillas infected with influenza A virus

Patients infected with influenza A virus (IAV) are at increased risk for bacterial superinfections, and this occurs in association with depressed polymorphonuclear leukocyte (PMNL) function. Recently, we reported that in vitro exposure of human PMNL to granulocyte-macrophage colony-stimulating factor (GM-CSF) reverses IAV-induced cell dysfunction. The present study used an established animal model of IAV infection to examine whether G-CSF and/or GM-CSF can overcome IAV- induced PMNL dysfunction and thereby prevent secondary infections. Preliminary studies determined a dosing schedule of these cytokines that caused significant priming of chinchilla PMNL. In subsequent studies, animals were inoculated intranasally with IAV (day 1) followed 3 days later by Streptococcus pneumoniae, and administered daily intraperitoneal injections with a cytokine or placebo on days 3 through 9. Animals had blood obtained on multiple occasions for PMNL studies, and were followed-up for evidence of pneumococcal disease. Both cytokines caused significant priming of the PMNL chemiluminescence response and this was associated with reversal of the IAV-induced PMNL dysfunction. However, neither cytokine decreased the incidence of pneumococcal disease.     

Studies on levamisole--induced agranulocytosis

Widespread clinical trials of leavo-tetramisole (levamisole) as an immunopotentiating agent in rheumatoid arthritis, metastatic carcinoma, and immunodeficiency states have been complicated by agranulocytosis (AGC) in 2.5%-13% of patients. Other than a relationship with prolonged high dosage, very little is known regarding the pathogenesis of levamisole-induced AGC. Whereas leukoagglutination was negative, fluorochromatic microgranulocytotoxicity (GCY) tests were positive with serum from 10 of 10 acutely neutropenic patients. The antibody was IgM, reacted with 100% of unrelated granulocytes, but not with T or B lymphocytes. Some sera also reacted with monocytes and the myeloid cell line, K-562. Tests for antigen-antibody complexes or cold autoantibodies were negative. Although clinical evidence strongly suggests a haptene (drug) mechanism, in vitro mixing experiments were also negative. An alternative choice parallels the model of aldomet- induced Coombs'-positive hemolytic anemia. Finally, GCY first became positive 2-3 mo prior to the onset of AGC on two patients, suggesting the possibility of identifying those at risk well before the onset of neutropenia.     

Platelet-derived growth factor promotes polymorphonuclear leukocyte activation

The platelet-derived growth factor (PDGF) has several well defined important biologic activities. Platelet-derived growth factor is the major mitogen in human serum for cells of mesenchymal origins; it is a potent chemoattractant protein for human monocytes, neutrophils, fibroblasts, and smooth muscle cells; and has been implicated in transformation by simian sarcoma virus and perhaps in transformation by other agents as well. In this article, PDGF has been shown to stimulate activation of human peripheral blood neutrophils defined by loss of membrane associated calcium as reflected by loss of chlortetracycline fluorescence, release of superoxide anion and specific granule enzymes, and enhanced neutrophil adherence and aggregation. These responses occurred in a dose-dependent fashion at concentrations of PDGF between 10 ng/mL (0.4 nmol/L) and 40 ng/mL (1.5 nmol/L) and were comparable to effects obtained with optimal concentrations of fMLP and C5a. Degranulation induced by PDGF was selective for secondary (specific) granules and not primary (azurophil) granules. Platelet-derived growth factor thus is ideally suited for a pivotal role in attracting inflammatory cells locally and initiating neutrophil activation at sites of blood vessel injury. Platelet-derived growth factor or a closely related protein also may play an important role in attracting and activating neutrophils in association with inflammatory tumors.     

Differentiation of natural killer (NK) cells from human primitive marrow progenitors in a stroma-based long-term culture system: identification of a CD34+7+ NK progenitor

We have recently described a marrow stroma-dependent long-term culture system that supports differentiation of CD34+ human marrow primitive progenitors into natural killer (NK) cells. We postulate that CD7 expression may be an early event in commitment of hematopoietic progenitors to the NK lineage. Here we compare the characteristics of CD34+7- and CD34+7+ marrow cells cultivated in the stroma-based NK culture system. These CD34+ populations were further compared with a marrow derived, more committed, CD34-7+ progenitor to emphasize the continuum of NK development and to highlight differences between progenitors in our assays. No progenitor proliferated when plated in media without stroma, underscoring the importance of stroma in NK differentiation. Plating progenitor populations in interleukin-2 containing media directly on preestablished, allogeneic, irradiated marrow stroma for 5 weeks resulted in CD56+CD3- NK cells; however, characteristics of the cultured populations differed. Fold expansion and cloning efficiency of the CD34+7+ population, determined by a functional limiting dilution assay was significantly higher than of the CD34+7- or CD34+7+ populations. This suggests that the CD34+7+ population is highly enriched for an NK progenitor and a possible intermediate in NK lineage differentiation. Further dividing the CD34+7+ population by the relative fluorescence of CD7 into CD34+7+dim and CD34+7+bright populations showed that the CD34+7+bright population exhibited a significantly higher cloning frequency than parallel experiments with CD34+7+dim cells (11.8% +/- 2.4% v 2.4% +/- 0.7%, n = 6; P = .005). Plating of the more primitive CD34+7- population in a transwell system (which separates progenitors from stroma by a microporous membrane) prevents differentiation into NK cells. In contrast, plating of CD34+7+ progenitors in transwells resulted in generation of NK cells. These data suggest that primitive, but not more mature NK progenitors may require direct contact with stroma for the initial differentiation steps. Finally, differentiation of the NK progenitors in this stroma-dependent model results in expression of CD2 not present on any of the starting populations. This observation suggests that marrow stroma can stimulate CD2 expression on NK progenitors in a previously undescribed fashion that may be analogous to the thymic effect on CD2 expression in immature T lymphocytes. These observations identify early steps in the commitment of primitive marrow CD34+ hematopoietic progenitors to a lymphoid lineage and underscore the importance of coexpression of CD7 with CD34 as an early lymphoid commitment characteristic and direct progenitor-stroma interactions in this process.