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Human alveolar epithelial cells engulf apoptotic eosinophils by means of integrin- and phosphatidylserine receptor-dependent mechanisms: A process upregulated.

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Presentation on theme: "Human alveolar epithelial cells engulf apoptotic eosinophils by means of integrin- and phosphatidylserine receptor-dependent mechanisms: A process upregulated."— Presentation transcript:

1 Human alveolar epithelial cells engulf apoptotic eosinophils by means of integrin- and phosphatidylserine receptor-dependent mechanisms: A process upregulated by dexamethasone  Darren W. Sexton, MSc, Morgan G. Blaylock, PhD, Garry M. Walsh, PhD  Journal of Allergy and Clinical Immunology  Volume 108, Issue 6, Pages (December 2001) DOI: /mai Copyright © 2001 Mosby, Inc. Terms and Conditions

2 Fig. 1 Representative light (original magnification 400×) and electron (original magnification 5000×) photomicrographs of epithelial cells ingesting apoptotic eosinophils at 60 minutes' interaction. A549 epithelial-cell engulfment of apoptotic eosinophils (A) , which are visualized as dark-staining peroxidase-positive cells, compared with ingestion by SAECs (B) . Electron micrographs of partial apoptotic-eosinophil ingestion (C) and near-total engulfment of an apoptotic eosinophil (D) by A549 cells. The arrow in (C) demonstrates evidence of partial digestion of an apoptotic eosinophil (ie, a phagosome containing a degenerated eosinophil, including an eosinophil granule). Journal of Allergy and Clinical Immunology  , DOI: ( /mai ) Copyright © 2001 Mosby, Inc. Terms and Conditions

3 Fig. 2 A , Effect of increasing concentrations of dexamethasone on apoptotic-eosinophil uptake by A549 cells compared with SAECs (percentage engulfment with each point representing the mean ± SEM of 5 experiments: *P < .05, **P < .005). B , Representative photomicrographs of basal ingestion of apoptotic eosinophils by resting A549 cells (left) and engulfment after treatment of the A549 monolayers with the glucocorticoid dexamethasone (right , 10–6 mol/L). C , Stimulation of A549 epithelial cells for 24 hours with a concentration range of dexamethasone cells increased their ingestion capacity for phagocytosis of apoptotic eosinophils. Each bar represents the mean ± SEM of 5 experiments. Journal of Allergy and Clinical Immunology  , DOI: ( /mai ) Copyright © 2001 Mosby, Inc. Terms and Conditions

4 Fig. 3 Representative expression data from flow cytometric analysis of A549 cell-surface receptors involved in phagocytosis of apoptotic eosinophils (β5, αvβ5, αvβ3, CD36, or PSR). In each histogram the dotted line represents the isotype-matched negative control, the solid line represents immunostaining with the indicated primary antibody, and the heavy solid line denotes immunostaining with the indicated primary antibody after incubation of the A549 monolayer with dexamethasone (10–6 mol/L for 24 hours). CD44 represents the positive control. Journal of Allergy and Clinical Immunology  , DOI: ( /mai ) Copyright © 2001 Mosby, Inc. Terms and Conditions

5 Fig. 4 Inhibition of apoptotic-eosinophil phagocytosis by A549 cells (percentage engulfment) through integrin, CD36, or PSR blockade. The contribution of these receptors to apoptotic-eosinophil engulfment by both resting and dexamethasone (10–6 mol/L)–treated A549 cells was assessed by using mAbs against these receptors and the appropriate isotype controls. Significant, comparable, and incomplete inhibition was observed with mAbs specific for αvβ5, αvβ3, CD36, and PSR, with no significant inhibition observed for the anti-CD11b mAb. Each bar represents the mean ± SEM of 5 experiments (*P < .05, **P < .005 compared with the relevant isotype-matched control in each case). Journal of Allergy and Clinical Immunology  , DOI: ( /mai ) Copyright © 2001 Mosby, Inc. Terms and Conditions

6 Fig. 5 Representative experiments showing immunostaining of A549 monolayers with isotype negative control, CD44, or αvβ5 mAbs. A , Negative staining with an isotype control mAb together with DAPI staining (blue) of epithelial nuclei. Note the presence of a red-immunostained CD9-PE–positive apoptotic eosinophil (arrow) . B , Positive immunostaining of A549 monolayer with an αvβ5 mAb visualized with anti-mouse antibody-FITC, demonstrating a phagosome containing a CD9-PE–labeled apoptotic eosinophil (arrow) . C , Demonstration of αvβ5 integrin receptor clustering around an apoptotic eosinophil inside a newly formed phagocytic vessel within an A549 epithelial cell. Note the diffuse membrane immunostaining for integrin αvβ5 on A549 cells, while the apoptotic eosinophil is surrounded by a αvβ5-positive phagosome, which has resulted in a high level of FITC-positive immunostaining in this area. All representative photomicrographs are at 400× magnification. The area of interest in B has been enhanced with Adobe PhotoShop. Journal of Allergy and Clinical Immunology  , DOI: ( /mai ) Copyright © 2001 Mosby, Inc. Terms and Conditions

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