Toll-like receptor-mediated eosinophil-basophil differentiation: autocrine signalling by granulocyte-macrophage colony-stimulating factor in cord blood haematopoietic progenitors.

Eosinophils are multi-functional leucocytes that play a role in inflammatory processes including allergy and infection. Although bone marrow (BM) inflammatory cells are the main source of eosinophil-basophil (Eo/B) differentiation-inducing cytokines, a recent role has been demonstrated for cytokine induction through Toll-like receptor (TLR)-mediated signalling in BM progenitors. Having previously demonstrated that cord blood (CB) progenitors induce Eo/B colony-forming units (CFU) after lipopolysaccharide (LPS) stimulation, we sought to investigate the intracellular mechanisms by which LPS induces Eo/B differentiation. Freshly isolated CD34-enriched human CB cells were stimulated with LPS (and/or pharmacological inhibitors) and assessed for alterations in haematopoietic cytokine receptor expression and signalling pathways by flow cytometry, Eo/B CFU in methylcellulose cultures, and cytokine secretion using Luminex assays. The LPS stimulation resulted in a significant increase in granulocyte-macrophage colony-stimulating factor (GM-CSF)-responsive, as opposed to interleukin-5-responsive, Eo/B CFU, which also correlated with significant increases in CD34(+) cell GM-CSFRα expression. Functionally, CB CD34(+) cells secrete abundant amounts of GM-CSF following LPS stimulation, via a p38 mitogen-activated protein kinase (MAPK)-dependent mechanism; this secretion was responsible for Eo/B CFU formation ex vivo, as shown by antibody blockade. We show for the first time that LPS stimulation of CB progenitor cells results in autocrine activation of p38 MAPK-dependent GM-CSF secretion facilitating Eo/B differentiation ex vivo. This work provides evidence that early life exposure to products of bacterial agents can modulate Eo/B differentiation, representing a novel mechanism by which progenitor cells can respond to microbial stimuli and so affect immune and inflammatory responses.

In vitro effects of budesonide on eosinophil-basophil lineage commitment.

UNLABELLED: 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.

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.

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.

Fish oil supplementation in pregnancy modifies neonatal progenitors at birth in infants at risk of atopy.

Dietary n-3 polyunsaturated fatty acids (PUFA) may represent a mode of allergy prevention. Cord blood (CB) CD34+ hemopoietic progenitors are altered in infants at risk of atopy. We therefore studied the effects of dietary n-3 PUFA supplementation during pregnancy on numbers and function of progenitors in neonates at high risk of atopy. In a double-blind study, atopic, pregnant women were randomized to receive fish oil capsules or placebo from 20 wk gestation until delivery. At birth, CB CD34+ cells were isolated and analyzed by flow cytometry for expression of cytokine (IL-5Ralpha, IL-3Ralpha, granulocyte/macrophage colony-stimulating factor Ralpha) or chemokine (CXCR4 and CCR3) receptors. CB cells were also cultured in methylcellulose assays for eosinophil/basophil colony-forming cells. At age 1 y, infants were clinically assessed for atopic symptoms and skin tests. Percentages of CB CD34+ cell numbers were higher after n-3 PUFA than placebo. Co-expression of cytokine or chemokine receptors on CD34 cells was not altered by n-3 PUFA supplementation. However, there were significantly more IL-5-responsive CB eosinophil/basophil colony forming units (Eo/B-CFU) in the fish oil, compared with the control, group. Overall, there was a positive association between CD34+ cells and IL-5-responsive Eo/B-CFU in CB and 1 y clinical outcomes, including atopic dermatitis and wheeze. Dietary n-3 PUFA supplementation during pregnancy in atopic mothers alters infant cord blood hemopoietic progenitor phenotype. This may have an impact on development of atopic disease.

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.

Anti-allergic therapies: effects on eosinophil progenitors.

Marked eosinophilic infiltration is the typical inflammatory response associated with allergic inflammation. Previous research involving animal and human models has established a role for the eosinophil/basophil hematopoietic progenitor in a systemic process of allergic inflammation. In this article, we will review the evidence implicating eosinophil/basophil progenitors in this systemic response and will discuss the rationale for targeting this cell in the treatment of allergic disease. In this context, we discuss corticosteroid treatment of allergic diseases, such as asthma and its effects on hematopoietic mechanisms, the effects of therapies that inhibit the actions of cysteinyl leukotrienes, the effects of in vivo blockade of the eosinophil-active cytokine interleukin-5, and the effects of antihistamines on hematopoiesis. It is suggested that several anti-allergic therapies exert their beneficial effects on allergic inflammation by influencing eosinophil production systemically. Therefore, targeting the systemic hematopoietic response may provide additional, more beneficial, therapeutic effects.