Lynn Crawford

Pathogenesis of Murine Experimental Allergic Rhinitis: A Study of Local and Systemic Consequences of IL-5 Deficiency

Recent studies have demonstrated an important role for IL-5-dependent bone marrow eosinophil progenitors in allergic inflammation. However, studies using anti-IL-5 mAbs in human asthmatics have failed to suppress lower airway hyperresponsiveness despite suppression of eosinophilia; therefore, it is critical to examine the role of IL-5 and bone marrow responses in the pathogenesis of allergic airway disease. To do this, we studied the effects of IL-5 deficiency (IL-5−/−) on bone marrow function as well as clinical and local events, using an established experimental murine model of allergic rhinitis. Age-matched IL-5+/+ and IL-5−/− BALB/c mice were sensitized to OVA followed by 2 wk of daily OVA intranasal challenge. IL-5−/− OVA-sensitized mice had significantly higher nasal mucosal CD4+ cells and basophilic cell counts as well as nasal symptoms and histamine hyperresponsiveness than the nonsensitized group; however, there was no eosinophilia in either nasal mucosa or bone marrow; significantly lower numbers of eosinophil/basophil CFU and maturing CFU eosinophils in the presence of recombinant mouse IL-5 in vitro; and significantly lower expression of IL-5Rα on bone marrow CD34+CD45+ progenitor cells in IL-5−/− mice. These findings suggest that IL-5 is required for normal bone marrow eosinophilopoiesis, in response to specific Ag sensitization, during the development of experimental allergic rhinitis. However, the results also suggest that suppression of the IL-5-eosinophil pathway in this model of allergic rhinitis may not completely suppress clinical symptoms or nasal histamine hyperresponsiveness, because of the existence of other cytokine-progenitor pathways that may induce and maintain the presence of other inflammatory cell populations.

Haemopoietic mechanisms in murine allergic upper and lower airway inflammation

Eosinophil recruitment to the airways, including involvement of haemopoietic eosinophil–basophil progenitors (Eo/B‐CFU), is primarily regulated by interleukin‐5 (IL‐5) and eotaxin. In this study, we investigated the haemopoietic mechanisms in upper and lower airway eosinophilic inflammation. Ovalbumin (OVA) sensitized and challenged BALB/c mice were used to establish isolated upper (UAC), isolated lower (LAC), or combined upper and lower airway (ULAC) inflammation. Airway, blood and bone marrow responses were evaluated in each model. Numbers of airway eosinophils and CD4+ cells were increased significantly in the nasal mucosa in UAC and ULAC mice, and in the lung tissue in LAC and ULAC groups. Levels of IL‐5 and eotaxin were increased significantly in the nasal lavage fluid (NL) in UAC and ULAC mice, and in the bronchoalveolar lavage fluid (BAL) in LAC and ULAC groups. The proportion of IL‐5‐responsive bone marrow Eo/B‐CFU was significantly higher than the control in all treatment groups, but peaked much earlier in the ULAC group. Kinetic studies revealed that IL‐5 and eotaxin in NL, BAL and serum peaked between 2 and 12 hr after OVA challenge in ULAC mice, and at 24 hr in UAC mice, related to the timing of maximal progenitor responses. These data support the concept that the systemic mechanisms linking rhinitis to asthma depend on the location and extent of airway allergen exposure.

Effects of a cysteinyl leukotriene receptor antagonist on eosinophil recruitment in experimental allergic rhinitis

The cysteinyl leukotrienes (cysLTs) are potent lipid mediators in allergic disease, acting through a receptor (cysLT1‐R) which can be targeted in rhinitis and asthma. We investigated the effects of cysLT1‐R antagonism in experimental allergic rhinitis, focusing on bone marrow eosinophil progenitor responses. BALB/c mice were sensitized, then given daily intranasal ovalbumin for 2 weeks, with montelukast sodium (5 mg/kg or 2·5 mg/kg) or placebo by gavage. Bone marrow eosinophil/basophil colonies were enumerated, and colony cells were morphologically assessed as indices of eosinophil differentiation and maturation. Montelukast treatment resulted in a significant decrease of eosinophils in the nasal mucosa, and in either bone marrow interleukin (IL)‐5‐, but not IL‐3‐, or granulocyte‐macrophage colony‐stimulating factor‐responsive eosinophil/basophil colony‐forming units, and IL‐5‐stimulated eosinophil maturation. These results indicate that cysLT1‐R antagonism in vivo limits both IL‐5‐responsive eosinophilopoiesis, acting at several stages of eosinophil differentiation and maturation. The anti‐allergic effects of cysLT1‐R antagonists are consistent with the concept that cysLTs and IL‐5 act together in the recruitment of eosinophils and eosinophil progenitors from the marrow during upper airway allergic inflammation.

The Effect of Desloratadine on Eosinophil/Basophil Progenitors and Other Inflammatory Markers in Seasonal Allergic Rhinitis: A Placebo-Controlled Randomized Study

Background: Eosinophil/basophil (Eo/B) progenitors fluctuate in the peripheral circulation during seasonal allergen exposure in atopic subjects. Several drugs have been shown to modulate Eo/B progenitor levels in the peripheral blood but, to date, the possible effect of antihistamines on Eo/B progenitors has not been explored. Our objective was to evaluate whether the antihistamine desloratadine (DL) can modulate peripheral blood Eo/B progenitors or other markers of allergic inflammation. Methods: We performed a randomized double-blind placebo-controlled study on the effects of DL on peripheral blood Eo/B progenitors in subjects with symptomatic, seasonal allergic rhinitis during a ragweed pollen season. Forty-five subjects were randomized to treatment for 4 weeks with DL 20 mg daily or placebo. Results: The expected fall in the number of Eo/B progenitors from baseline to 2 weeks of treatment was seen in the placebo group [median drop of 1.0 colony-forming unit (CFU)/106 cells], and was greater than in the DL group (median drop of 0.0 CFU/106 cells) (p = 0.013). The change in histamine concentration per colony from baseline to 2 weeks of treatment was lower in the DL group (median decrease of 6.1 pg/colony) compared to placebo (median increase of 1.8 pg/colony) (p = 0.01). An increase in the nasal lavage eotaxin concentration from baseline to 4 weeks of treatment was statistically significant in the placebo group but not in the DL group. Eo/B CFU were not affected by varyingin vitro concentrations of DL. Conclusion: These results suggest that DL can modulate aspects of allergic inflammation in vivo through mechanisms other than simple blockade of H1 histamine receptors.

An evaluation of peripheral blood eosinophil/basophil progenitors following nasal allergen challenge in patients with allergic rhinitis

Objective To evaluate the effect of a single nasal allergen challenge on peripheral blood eosinophil/basophil (Eo/B) progenitor cells and induced sputum eosinophil counts in subjects with allergic rhinitis.

The effect of desloratadine on eosinophil/basophil progenitors and other inflammatory markers in seasonal allergic rhinitis: A placebo-controlled randomized study

BACKGROUND Eosinophil/basophil (Eo/B) progenitors fluctuate in the peripheral circulation during seasonal allergen exposure in atopic subjects. Several drugs have been shown to modulate Eo/B progenitor levels in the peripheral blood but, to date, the possible effect of antihistamines on Eo/B progenitors has not been explored. Our objective was to evaluate whether the antihistamine desloratadine (DL) can modulate peripheral blood Eo/B progenitors or other markers of allergic inflammation. METHODS We performed a randomized double-blind placebo-controlled study on the effects of DL on peripheral blood Eo/B progenitors in subjects with symptomatic, seasonal allergic rhinitis during a ragweed pollen season. Forty-five subjects were randomized to treatment for 4 weeks with DL 20 mg daily or placebo. RESULTS The expected fall in the number of Eo/B progenitors from baseline to 2 weeks of treatment was seen in the placebo group [median drop of 1.0 colony-forming unit (CFU)/10(6) cells], and was greater than in the DL group (median drop of 0.0 CFU/10(6) cells) (p = 0.013). The change in histamine concentration per colony from baseline to 2 weeks of treatment was lower in the DL group (median decrease of 6.1 pg/colony) compared to placebo (median increase of 1.8 pg/colony) (p = 0.01). An increase in the nasal lavage eotaxin concentration from baseline to 4 weeks of treatment was statistically significant in the placebo group but not in the DL group. Eo/B CFU were not affected by varying in vitro concentrations of DL. CONCLUSION These results suggest that DL can modulate aspects of allergic inflammation in vivo through mechanisms other than simple blockade of H1 histamine receptors.

In Vitro Effects of Budesonide on Eosinophil-Basophil Lineage Commitment

IL-5 is the primary cytokine that stimulates the production and survival of eosinophils and basophils from progenitor cells. The inhaled glucocorticoid, budesonide, has been shown to exert a therapeutic effect via suppression of eosinophil/basophil progenitors in vivo. Since various steroids have exhibited the ability to enhance eosinophil/basophil progenitor differentiation, we examined the effects of budesonide in vitro. Bone marrow and cord blood samples were obtained and cultured in the presence of IL-5 alone or IL-5 plus budesonide. Eosinophil/basophil colony-forming units were enumerated from cultured nonadherent mononuclear cells and from purified CD34+ cells. CD34+ cells with and without budesonide were also examined for up-regulation of ERK1/2, MAPK and GATA-1 using real time-PCR. Results: i) up-regulation of eosinophil/basophil colony-forming units is due to the direct effects of budesonide on IL-5-stimulated progenitors; ii) GATA-1 is likely involved in the early amplification of eosinophil/basophil progenitor commitment leading to increased differentiation. A potential transcriptional pathway has been identified which may mediate the effects of budesonide on eosinophil/basophil lineage commitment.

Effects of Budesonide on Eosinophil-Basophil Lineage Commitment

IL-5 is the primary cytokine that stimulates the production and survival of eosinophils and basophils from progenitor cells. The inhaled glucocorticoid, budesonide, has been shown to exert a therapeutic effect via suppression of eosinophil/basophil progenitors in vivo. Since various steroids have exhibited the ability to enhance eosinophil/basophil progenitor differentiation, we examined the effects of budesonide in vitro. Bone marrow and cord blood samples were obtained and cultured in the presence of IL-5 alone or IL-5 plus budesonide. Eosinophil/basophil colony-forming units were enumerated from cultured nonadherent mononuclear cells and from purified CD34+ cells. CD34+ cells with and without budesonide were also examined for up-regulation of ERK1/2, MAPK and GATA-1 using real time-PCR. Results: i) up-regulation of eosinophil/basophil colony-forming units is due to the direct effects of budesonide on IL-5-stimulated progenitors; ii) GATA-1 is likely involved in the early amplification of eosinophil/basophil progenitor commitment leading to increased differentiation. A potential transcriptional pathway has been identified which may mediate the effects of budesonide on eosinophil/basophil lineage commitment.

Effects of the cysteinyl leukotriene receptor antagonist, montelukast, on eosinophil differentiation in an experimental mouse model of allergic rhinitis

The cysteinyl leukotriene (CysLT) LTD 4 is a potent lipid mediator which is synthesized and released from various inflammatory cells, and contributes to the pathophysiology associated with allergic disease. The cloning and characterization of the LTD 4 receptor (CysLTtR), has provided a useful target for therapy of rhinitis and asthma; in particular, montelukast sodium is a selective antagonist that binds with high affinity to CysLT]R on the surface of eosinophils and their progenitors. In the current study, we investigated the effects of montelukast on bone marrow eosinophil progenitor responses, using our recently developed experimental murine model of allergic rhinitis. BALB/c mice were sensitized and challenged with ovalbumin (OVA), and montelukast (5 mg/kg) or placebo was administered by gavage, for 1 or 2 wk, 1 hr before daily intra-nasal OVA challenge. Bone marrow colony assays in methylcellulose were performed, and day 6 eosinophil/basophil colony forming units (Eo/Baso-CFU) were enumerated. Colony cells (day 10) were isolated and histologically stained to allow morphologic assessment of eosinophil differentiation. Bone marrow Eo/Baso-CFU numbers, grown in the presence of interleukin 5 (IL-5) in vitro, were significantly lower than that those from controls in 2 wk (p<0.002), but not 1 wk, montelukast-treated mice. Highly significant suppression of eosinophil maturation compared to control (p<0.001) was observed in colony cells from both 1 wk and 2 wk montelukast-treated groups. These results indicate that montelukast, presumably via CysLT] R antagonism, limits the number of progenitors that differentiate into mature eosinophils; the combination of reduced CFU, as well as lower numbers of mature eosinophils per CFU, suggests that these effects may be occurring at more than one stage of eosinophil differentiation. The anti-allergic effects of montelukast may include targeting the recruitment of eosinophil progenitors from the bone marrow.

Cord blood molecular biomarkers of eosinophilopoiesis: Kinetic analysis of GATA‐1, MBP1 and IL‐5Rα mRNA expression

Ellis AK, Ackerman SJ, Crawford L, Du J, Bedi R, Denburg JA. Cord blood molecular biomarkers of eosinophilopoiesis: Kinetic analysis of GATA‐1, MBP1 and IL‐5Rα mRNA expression.
Pediatr Allergy Immunol 2010: 21: 640–648.
© 2010 John Wiley & Sons A/S