Anti-Fas mAb-induced apoptosis and cytolysis of airway tissue eosinophils aggravates rather than resolves established inflammation - PubMed (original) (raw)

Anti-Fas mAb-induced apoptosis and cytolysis of airway tissue eosinophils aggravates rather than resolves established inflammation

Lena Uller et al. Respir Res. 2005.

Abstract

Background: Fas receptor-mediated eosinophil apoptosis is currently forwarded as a mechanism resolving asthma-like inflammation. This view is based on observations in vitro and in airway lumen with unknown translatability to airway tissues in vivo. In fact, apoptotic eosinophils have not been detected in human diseased airway tissues whereas cytolytic eosinophils abound and constitute a major mode of degranulation of these cells. Also, Fas receptor stimulation may bypass the apoptotic pathway and directly evoke cytolysis of non-apoptotic cells. We thus hypothesized that effects of anti-Fas mAb in vivo may include both apoptosis and cytolysis of eosinophils and, hence, that established eosinophilic inflammation may not resolve by this treatment.

Methods: Weeklong daily allergen challenges of sensitized mice were followed by airway administration of anti-Fas mAb. BAL was performed and airway-pulmonary tissues were examined using light and electron microscopy. Lung tissue analysis for CC-chemokines, apoptosis, mucus production and plasma exudation (fibrinogen) were performed.

Results: Anti-Fas mAb evoked apoptosis of 28% and cytolysis of 4% of eosinophils present in allergen-challenged airway tissues. Furthermore, a majority of the apoptotic eosinophils remained unengulfed and eventually exhibited secondary necrosis. A striking histopathology far beyond the allergic inflammation developed and included degranulated eosinophils, neutrophilia, epithelial derangement, plasma exudation, mucus-plasma plugs, and inducement of 6 CC-chemokines. In animals without eosinophilia anti-Fas evoked no inflammatory response.

Conclusion: An efficient inducer of eosinophil apoptosis in airway tissues in vivo, anti-Fas mAb evoked unprecedented asthma-like inflammation in mouse allergic airways. This outcome may partly reflect the ability of anti-Fas to evoke direct cytolysis of non-apoptotic eosinophils in airway tissues. Additionally, since most apoptotic tissue eosinophils progressed into the pro-inflammatory cellular fate of secondary necrosis this may also explain the aggravated inflammation. Our data indicate that Fas receptor mediated eosinophil apoptosis in airway tissues in vivo may cause severe disease exacerbation due to direct cytolysis and secondary necrosis of eosinophils.

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Figures

Figure 1

Figure 1

Study design. All animals were immunized with OVA and 14 days later exposed to aerosol challenge with OVA for 7 days to establish tissue and lumen eosinophilia. Treatment with anti-Fas mAb or isotype control IgG was administered intra-nasally at day 22 and outcome measurements including BAL and tissue sampling were made at 8 and 24 h after treatment.

Figure 2

Figure 2

After eosinophilia had been established in the immunized mice anti-Fas mAb or isotype control Ab was administered locally to the lungs followed by BAL and tissue sampling at 8 and 24 hours. The number of eosinophils in airway lumen (A) and airway tissue (B) was quantified as described in detail in the methods section. White bars represent mice given control Ab and black bars mice given anti-Fas mAb. Error bars indicate the standard error of the mean for each group of mice (n = 8, ** = p < 0.01). Peribronchial eosinophilia induced by the OVA challenges is shown in (C).

Figure 3

Figure 3

Representative light micrographs of mouse lung tissue using Htx-staining in control (A) and anti-Fas mAb treated animals (B) at 24 h. Htx-staining shows dark condensed (pycnotic) nuclei of eosinophils and disturbed epithelial lining. Very few TUNEL-positive apoptotic cells were present in control treated animals (C) whereas a large number of TUNEL-stained cells was detected in anti-Fas mAb treated animals (D), almost all of which were shown to be apoptotic eosinophils by double chromotrope 2R and TUNEL staining (see also Figure 4).

Figure 4

Figure 4

Transmission electron micrographs of lung tissue from mice with OVA-induced eosinophilia. Control mice treated with the isotype control Ab showed no sign of eosinophil apoptosis at 8 h (A) and 24 h (not shown). In mice treated with anti-Fas mAb there were numerous apoptotic eosinophils in the lung tissues at both 8 and 24 hours after treatment (B and C, respectively). The apoptotic eosinophils were rarely engulfed although macrophages (labeled M) commonly occurred in the tissue (B). By 24 h a majority of the apoptotic eosinophils exhibited signs of secondary necrosis and severe inflammation was recorded including neutrophil infiltration (arrow) and derangement of the epithelial lining (labeled E).

Figure 5

Figure 5

These micrographs illustrate characteristic eosinophil phenotypes present in mouse airways in this study: (A) viable non-degranulating eosinophil, the only phenotype found in lung tissues of allergen challenged animals; (B) apoptotic eosinophil exhibiting nuclear condensation, cell shrinkage, and an intact cell membrane; (C) an apoptotic eosinophil exhibiting secondary necrosis involving cell membrane rupture and piecemeal degranulation; (D) a cytolytic eosinophil exhibiting chromatolysis, cell membrane rupture, and spilling of electron-dense specific granules into the tissue (arrow).

Figure 6

Figure 6

CC-chemokine mRNA expression 8 h (A) and 24 h (B) after treatment with anti-Fas mAb or isotype control (IgG). Two days post allergen challenge expression of 5 different CC-chemokines in the lung was up-regulated compared to immunized control animals receiving saline challenges. Treatment with anti-Fas mAb post allergen challenge further increased the expression of eotaxin MIP-1α, and MIP -1β, and additionally induced the expression of IP-10 and MCP-1. Data are mean ± SEM. **p < 0.01 indicates differences between OVA and saline treatments. §§ p < 0.01 indicates difference between anti-Fas mAb treated and control-treated OVA-challenged animals.

Figure 7

Figure 7

Observations in allergen challenged animals demonstrating anti-Fas-induced mucus-exudate plugs (A,C,D,E) compared to control isotype antibody (IgG) treatment (A,B). The occurrence of mucus-exudate plugs is expressed as percentage affected airways in each tissue section in control mice (white bars) and in mice treated with anti-Fas mAb (black bars) at 8 and 24 hours (A). Bars indicate the standard error of the mean for each group of animals (n = 8, ** = p < 0.01). Histochemical staining with periodic acid-Schiff reagent (PAS) illustrated mucus-containing cells (B,C) and so did the transmission electron micrograph (D). Tethered secretions and lumen plugs occurred foremost in anti-Fas treated airways (C,D;E). Fibrinogen immuno-reactivity was distributed in the mucus plugs exclusively in anti-Fas treated airways (E).

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References

    1. Fadok VA, Bratton DL, Henson PM. Phagocyte receptors for apoptotic cells: recognition, uptake, and consequences. J Clin Invest. 2001;108(7):957–962. doi: 10.1172/JCI200114122. - DOI - PMC - PubMed
    1. Henson PM, Bratton DL, Fadok VA. The phosphatidylserine receptor: a crucial molecular switch? Nat Rev Mol Cell Biol. 2001;2(8):627–633. doi: 10.1038/35085094. - DOI - PubMed
    1. Davies DE, Wicks J, Powell RM, Puddicombe SM, Holgate ST. Airway remodeling in asthma: new insights. J Allergy Clin Immunol. 2003;111(2):215–225. doi: 10.1067/mai.2003.128. quiz 226. - DOI - PubMed
    1. Persson CG. Centennial notions of asthma as an eosinophilic, desquamative, exudative, and steroid-sensitive disease. Lancet. 1997;350(9083):1021–1024. doi: 10.1016/S0140-6736(96)02335-5. - DOI - PubMed
    1. Alam R, Busse WW. The eosinophil – quo vadis? J Allergy Clin Immunol. 2004;113(1):38–42. doi: 10.1016/j.jaci.2003.10.054. - DOI - PubMed

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