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Background: Dendritic cells (DC) are thought to play a key role in the initiation and maintenance of T cell immunity to inhaled antigens. While the density of DC within the bronchial mucosa is increased in stable asthma, there is little information currently available concerning the effects of allergen inhalation on DC in subjects with asthma., Objectives: To enumerate changes in the numbers of circulating CD33(+) myeloid DC in asthmatics, before and after allergen challenge., Methods: Blood DC numbers were enumerated by flow cytometry before and at 3, 6 and 24 h after inhaled allergen and diluent in 10 mild, allergic asthmatic subjects., Results: Blood DC numbers rapidly fell from 3.42 +/- 0.30 x 10(7)/L at baseline, to 2.10 +/- 0.17 x 10(7)/L by 3 h post-challenge (P < 0.01), and remained significantly below baseline values at both 6 and 24 h following allergen challenge. No such changes in DC numbers were noted after diluent challenge. A similar, early fall in circulating lymphocytes was also noted post-allergen challenge, whereas changes in circulating eosinophil and neutrophil numbers occurred more slowly., Conclusions: A significant proportion of myeloid DC rapidly 'disappear' from the circulation following allergen inhalation, suggesting that margination of circulating myeloid DC, and their recruitment into the airway mucosa, is an important feature of the immune response to inhaled allergen.

Background: Airway dendritic cells (DC) play an important role in chronic allergic airway inflammation in experimental animals, but a similar role for DC in human allergic asthma has been difficult to define. This pilot study was undertaken to elucidate the role of DC in allergic asthma by examining their potential to migrate to the lower airways in response to bronchial challenge with specific allergen., Methods: Bronchial biopsy specimens were obtained from seven patients with allergic asthma before and 4-5 hours after allergen challenge. Multicolour immunofluorescence staining was performed on mucosal cryosections to identify changes in the number and phenotypes of DC., Results: A dramatic increase in the number of CD1c+HLA-DR+ DC were observed in the lamina propria after challenge compared with baseline (22.4 v 7.8 cells/mm(2)). The rapid accumulation (within 4-5 hours) of these cells strongly suggests that they were directly recruited from peripheral blood., Conclusion: We have shown for the first time that a specific DC subset rapidly emigrates into the human bronchial mucosa during allergic inflammation. While this study is based on relatively few patients, the consistency of the overall results strongly suggests that the rapid population dynamics of human airway DC closely parallel those in animal models of acute inflammation. These findings support suggestions that DC have an important role in human airway allergy.

Background: Recognition of the importance of dendritic cells (DC) in the initiation of T-cell-dependent immune responses has led to increasing interest in methods for the identification of DC within the circulation. We sought to develop a flow cytometric method that would allow the reliable enumeration of absolute myeloid DC counts in minimally manipulated blood samples., Methods: Myeloid DC were identified by three-color staining of whole blood leukocytes as a discrete population of mononuclear cells expressing high levels of HLA-DR and CD33, yet having little or no expression of CD14 and CD16. This method was analyzed for reproducibility and variation in blood DC number during typical clinical day hours and after exercise. The new method was compared to an established commercial kit method., Results: FACS sorting of the CD33(+) DC showed that they morphologically resembled immature DC, and developed cytoplasmic projections typical of mature DC following overnight culture in granulocyte macrophage-colony stimulating factor (GM-CSF). Within peripheral blood, these DC were found at a mean concentration of 17. 4 +/- 5.4 x 10(6) per liter, corresponding to 0.93 +/- 0.27% of mononuclear cells. Comparison of duplicate samples stained and analyzed in parallel showed that the intrasample variability was very low, with an intraclass correlation coefficient of 0.95. The frequency of CD33(+) myeloid DC and their light scatter characteristics were similar to that of CD11c(+) myeloid cells. Four-color FACS analysis revealed complete identity of CD11c(hi), HLA-DR(+) DC with CD33(+), HLA-DR(+) DC. Only rare CD33(+) DC coexpressed CD123 and HLA-DR. Numbers of blood myeloid DC, identified by CD33 staining, showed no significant variation during standard laboratory hours. However, their numbers rose significantly during vigorous exercise, in parallel to other blood cells., Conclusions: The method described herein is rapid, reproducible, requires only small volumes of blood, can be readily used by a clinical immunology laboratory, and requires fewer antibodies than a currently available commercial method., (Copyright 2000 Wiley-Liss, Inc.)