Susanna Oddera

Sensitization to Airborne Allergens in Children with Respiratory Symptoms

BACKGROUND Allergy is one of the most common causes of respiratory symptoms in children and youth. OBJECTIVE Evaluate the presence and the type of allergic sensitization in a paediatric population with respiratory symptoms. METHODS We studied 564 consecutive children, 5 months to 17 years of age, with a male to female ratio (M/F) = 1.4, referred to our outpatient clinic in a 12-month period retrospectively. Patients were arbitrarily divided into four groups (grs) according to their age: gr1 = 5 months to 4 years old (181 patients), gr2 = 4 to 7 years (201 patients), gr3 = 7 to 10 years (96 patients), and gr4 = 10 to 17 years (86 patients). Sensitization to house dust mites, pollens, animal dander, and molds was determined by skin prick testing. RESULTS Sensitization to at least one class of allergen occurred in 304 of the 564 patients (53.9%, M/F ratio = 2.0); the percentage of allergic patients increased with age as follows: 29.8% (54 patients) of the patients in gr1, 55.2% (111 patients) in gr2, 68.8% (66 patients) in gr3 and 84.9% (73 patients) in gr4 (chi(2) = 84.1, P < .01). In the entire allergic population and in gr1 to gr3, the most common positive allergic reaction was to house dust mites (P < .01, chi(2) test each comparison). In contrast, gr4 patients showed a nearly equal percentage of sensitization to pollens and to house dust mites (79.5% and 78.1% respectively) (chi(2) = 0.0, P = >.1). Sensitization to only one class of allergen occurred in 51.3% of the allergic patients and the percentage of these monosensitized patients tended to decrease from gr1 to gr4 (chi(2) = 15.2, P < .1). In the monosensitized group, sensitization to house dust mites was the most frequent in gr1 to gr3 (age <10 years) as in the whole sample. In gr4, the frequency of sensitization to house dust mites was similar to that of sensitization to pollens. On the contrary, within the patient group sensitized to two or more allergens (polysensitized patients), sensitization to house dust mites was as frequent as sensitization to pollens already in gr2 as compared with monosensitized patients. CONCLUSIONS In children with respiratory symptoms, the percentage of allergic individuals was high and increased with the age of the patients. This phenomenon was associated with an age-related enhancement in the ratio of polysensitized to monosensitized patients and with an age-related increase in the frequency of sensitization to seasonal allergens (ie, pollens).

Increased expression of the CD80 accessory molecule by alveolar macrophages in asthmatic subjects and its functional involvement in allergen presentation to autologous TH2 lymphocytes.

BACKGROUND Alveolar macrophages (AMs) are more efficient antigen-presenting cells in allergic individuals than in nonatopic subjects. OBJECTIVE We studied whether this difference may be correlated to increased expression of membrane costimulatory molecules, such as the B7 molecules (CD80 and CD86). METHODS Eleven subjects with allergic asthma sensitized to Dermatophagoides pteronyssinus and 5 healthy nonatopic volunteers underwent bronchoalveolar lavage, and the costimulatory molecule expression on AMs was evaluated. Peripheral blood T cells, either freshly isolated or as established D pteronyssinus -specific cell lines, were cultured with autologous monocytes or AMs as antigen-presenting cells. In vitro allergen-induced proliferation and cytokine production were evaluated in the presence of B7-blocking reagents. RESULTS Allergic individuals had a significantly higher proportion of AMs expressing the CD80 molecule than control subjects (28.5% +/- 14.8% vs 1.4% +/- 1.2%; P <.001), whereas no difference was observed in CD86 expression (2.0% +/- 2.3% vs 1.1% +/- 0.6; P >.1). In a large proportion of the asthmatic subjects we studied, AMs were presenting soluble antigens (tetanus toxoid and streptolysin-O) to freshly isolated T cells more efficiently than AMs from nonatopic control subjects. Finally, both T-cell proliferation and cytokine production of D pteronyssinus- specific established T-cell lines were inhibited by a CD80-blocking antibody in a dose-dependent manner. CONCLUSION Costimulation by means of CD80 expressed by AMs is probably involved in the amplification of the allergen-specific T-lymphocyte response in the airways of asthmatic subjects.

CD40 on Adult Human Airway Epithelial Cells: Expression and Proinflammatory Effects

CD40/CD40 ligand interaction is an important pathway for B and T cell cooperation and function; functional CD40 molecules have recently been found on nonhematopoietic cells. We detected CD40 in vivo on normal human respiratory epithelial cells and showed that its expression is increased on inflamed airway epithelium. Subsequently, we analyzed its expression and function on primary cultures of human airway epithelial cells. Our data show that CD40 is up-regulated by IFN-β and IFN-γ, its ligation increases the surface expression of CD54 and CD106 and it may stimulate the release of IL-6 and IL-8. The use of Janus kinase 3 (JAK3) and NF-κB inhibitors suggests that both basal and CD40-induced release of the two cytokines is JAK3-dependent. Using colocalization techniques, we revealed the existence of CD40/JAK3 and CD40/TNFR-associated factor 2 interplay. The extent of these interactions may be partial (2–40% of the cells) or massive (80–90% of the cells) in cultured cells. Stimulation via CD40 causes a significant increase in the number of cells expressing colocalization only in the cultures displaying low frequency of initial colocalization. Thus, airway epithelial cells, activated by CD40, may behave as effector cells of the inflammation process and should be considered priority targets for anti-inflammatory therapy. This work identifies CD40 and the correlated JAK3 signaling molecule as potential molecular targets to block the inflammatory functions of epithelial cells.

Frequency and Specific Sensitization to Inhalant Allergens Within Nuclear Families of Children with Asthma and/or Rhinitis

OBJECTIVE The purpose of this study was to investigate the frequency of allergy in 85 families of pediatric patients with asthma and/or rhinitis. METHODS Families enrolled were drawn according to a table of randomization, from those whose children were referred the outpatient clinic of the Pulmonology Department of Gaslini Institute (Genoa, Italy). In patients and in both their mothers and fathers, allergic sensitization to the three most common classes of inhalant allergens (house dust mites, pollens, and animal danders) was evaluated by skin prick test. RESULTS As compared with their parents, children showed a similar prevalence of positive skin prick test (50.0% and 58.8%, respectively; chi 2 = 1.4; P > .1), but a significantly higher frequency of specific sensitization to house dust mites, pollens, or pets (P < .05, each chi 2). These differences were at least partially related to the higher frequency of polysensitization (i.e., sensitization to more than one class of allergens) in children than in their parents [chi 2 = 4.2, odds ratio (OR) = 2.3, 95% confidence interval (CI95%) = 1.0 to 5.1; P < .05). In addition, both in children (chi 2 = 5.8, OR = 11.3, CI95% = 1.4 to 81.5; P < .05] and in their parents (chi 2 = 7.4, OR = 4.8, CI95% = 1.5 to 16.7; P < .01), allergy to pollens was less frequent in monosensitized than in polysensitized families, while the prevalence of sensitization to house dust mites was similar in monosensitized and in polysensitized families. The analysis of the role of parental sensitization in the development of allergy in the offspring demonstrated that the prevalence of allergic children: (1) seemed to be higher, without reaching statistical significance, in families with sensitized parents than in those with non-sensitized parents (chi 2 = 0.41, P > .1) and (2) was higher in families with one or both polysensitized parents than in those with one or both monosensitized parents (chi 2 = 4.5, OR = 4.0, CI95% = 1.1 to 15.6; P < .05). In addition, within each family, the coincidence of sensitization to house dust mites was more frequent than that to pollens (60.0% and 36.0%, respectively; chi 2 = 7.6, OR = 4.5, CI95% = 1.5 to 14.3; P < .01), and it was not influenced by the number of parents sensitized to same allergen. CONCLUSION These data support the concept that, in addition to genetic predisposition, other factors (i.e., environmental exposure) may influence the development of specific sensitization in children with respiratory symptoms. The different sensitization found in this study between children and their parents occurred in a generation span, so as it is more likely to be related to environmental changes than to genetic factors.

Novel Anti-inflammatory Effects of the Inhaled Corticosteroid Fluticasone Propionate During Lung Myofibroblastic Differentiation

Asthma is characterized by an irreversible subepithelial fibrosis with the appearance of myofibroblasts, which can be now considered important early participants in inflammatory responses as well as potential targets for anti-inflammatory drugs. In this study, we show that fluticasone propionate (FP), a powerful inhaled corticosteroid (ICS), displays novel anti-inflammatory effects on human lung fibroblasts during their myofibroblastic differentiation. Indeed, FP inhibits in lung myofibroblasts, at a very early stage of differentiation, the activation of Janus kinase/STAT pathways induced by IL-13 (tyrosine kinase 2, STAT1, STAT3, STAT6, mitogen-activated protein kinase). Contrarily, in mildly or fully differentiated myofibroblastic cultures, FP still displays a potential anti-inflammatory activity even if it only inhibits tyrosine kinase 2 phosphorylation. Moreover, FP inhibits constitutive and TGF-β-induced expression of α-smooth muscle actin, the main marker of myofibroblastic differentiation, both in very early and in mild differentiated myofibroblasts. Finally, FP displays an additional powerful anti-inflammatory effect, decreasing nuclear translocation of NF-κB independent of the degree of myofibroblastic differentiation. These data 1) suggest that myofibroblasts are priority targets for ICS, which is able to revert them to a normal phenotype even if they appear to be already engaged in their differentiation, and 2) may help to explain why asthma is improved by an early ICS treatment, whereas advanced asthma is more resistant to these drugs.

Effects of Triamcinolone Acetonide on Adult Human Lung Fibroblasts: Decrease in Proliferation, Surface Molecule Expression and Mediator Release

Background: Lung fibroblasts may have a pivotal role in airway inflammation as they are involved in continuous cycles of mediator secretion, proliferation, activation and cross-talk with recruited inflammatory cells. The role of fibroblasts as intermediate participants in the inflammatory network suggests that they could represent an important target for drugs commonly used in asthma; thus, we investigated the effects of triamcinolone acetonide (TAA) on primary human lung fibroblasts. Methods: The in vitro activity of increasing concentrations (10–9 to 10–7 M) of TAA in fibroblast cultures was evaluated as regards the following parameters: proliferation, extracellular matrix (ECM) release, cytokine/chemokine secretion and surface antigen expression. Results: All concentrations of TAA decreased fetal calf serum (FCS)-induced fibroblast proliferation, whereas in the presence of FCS plus basic fibroblast growth factor TAA was only effective at 10–8 and 10–7M. TAA failed to decrease ECM, whereas at 10–8 and 10–7M it decreased IL-6 and IL-8 secretion to different extents. In the presence of IFN-γ the drug was able to reduce VCAM-1 expression at all of the tested concentrations; on the other hand, in TGF-β1-driven cultures a decrease in CD54 expression was detected with TAA at 10–8 and 10–7M. Conclusions: TAA acts on some functional properties of human lung fibroblasts that make these cells active participants in the inflammatory network. The ability of TAA to inhibit lung fibroblast proliferation may prevent or even reverse some of the histological changes that characterize airway remodeling in chronic inflammatory diseases; moreover, IL-6, IL-8 and surface molecule decreases by TAA may suggest a direct anti-inflammatory effect of the drug by suppression of resident lung cell function.

Effects of Mizolastine in vitro on Human Immunocompetent and Airway Cells: Evidence for Safety and Additional Property

Background: Mizolastine is a potent, peripherally acting, selective H1-receptor antagonist with potential anti-inflammatory properties. The aim of the study was to evaluate the in vitro effects of mizolastine on the expression of adhesion molecules by primary human airway epithelial and stromal cultures; moreover, the activity of mizolastine on parameters which reflect the immune response efficacy was investigated. Methods: Airway epithelial and stromal cells were collected from hypereosinophilic subjects by enzymatic digestion of polyps or turbinates. Cells were stimulated with interferon (IFN)-γ (500 IU/ml) in the presence of various mizolastine concentrations (6 × 10–8–6 × 10–6 M) for 24 h and the expression of CD106, CD54, CD58 and HLA class I was evaluated. Peripheral blood mononuclear cells from healthy volunteers were incubated with 1% phytohemagglutinin or anti-CD3 monoclonal antibody (20 ng/ml) in the presence of mizolastine, then T lymphocyte proliferation, HLA-DR expression and T cell subpopulations were evaluated. Results: Both in epithelial and stromal cultures, IFN-γ significantly upregulated all of the tested surface molecules (p < 0.05). The highest dose of mizolastine (6 × 10–6 M), corresponding to 10-fold the peak plasma level after a single oral administration of 10 mg, was able to act on fibroblasts, significantly downregulating the expression of CD54 (p < 0.05). Regarding T lymphocyte proliferation, the addition of mizolastine did not induce any significant change; furthermore, mizolastine was ineffective at all of the tested concentrations on both HLA-DR expression and CD4+/CD8+ ratio. Conclusions: This study demonstrated that mizolastine is able to selectively downregulate CD54 expression on stimulated stromal but not epithelial cells without impairing the immune system effectors. The possible clinical significance of these results are an antiallergic property and CD54 modulation on fibroblasts with a good safety profile as far as the lymphocyte response is concerned.

In childhood asthma the degree of allergen-induced T-lymphocyte proliferation is related to serum IgE levels and to blood eosinophilia.

OBJECTIVE To investigate whether the state of activation of circulating T-cells in childhood asthma could be related to serum IgE levels and/or to blood eosinophilia. METHODS Seventeen atopic asthmatic children, sensitized to Dermatophagoides pteronyssinus (Der p), in stable condition at the time of the study and 15 sex-matched and age-matched controls were studied. The expression of activation surface markers (HLA-DR and CD25) on peripheral blood mononuclear cells (PBMCs) was tested by monoclonal antibodies and FACS analysis, while the PBMC proliferative response to Der p antigens was measured by tritiated thymidine (3HTdR) incorporation. RESULTS As compared to controls, atopic children showed higher eosinophil counts (P < .01), similar lymphocyte counts (P > .1, each comparison) but higher proportion of HLA-DR+ and CD25+ T-lymphocytes (P < .05, each comparison). A significant Der p allergen-induced PBMC proliferation was observed in atopic children (P < .01) but not in controls (P > .1). Both in controls and in atopic children, no correlations were found between lymphocyte counts and eosinophil counts or total or allergen-specific IgE levels (P > .1, each comparison). In contrast, weak correlations were detected between the degree of allergen-induced PBMC proliferation and: a) allergen-specific IgE levels in serum (P < .05) and b) eosinophil counts (P < .05). CONCLUSION These data support the concept that the degree of activation of allergen-specific T-lymphocytes in blood may reflect the intensity of allergic sensitization in childhood asthma.

Cetirizine-induced downregulation of airway fibroblast proliferation and function: a rationale for a different approach to allergy treatment?

Recently, airway fibroblasts captured the attention of both allergists and basic scientists since they are no longer considered as mere bystanders, as far as allergic airway diseases are concerned. The aim of the present study was to assess the effects of different Cetirizine (Cet) concentrations (0.01, 0.05, 0.1 mg/ml) on human airway fibroblast proliferation and on CD54 expression. By means of flow cytometry analysis, we evaluated CD54 expression by airway fibroblasts in basal conditions or after gammaIFN stimulation in the presence of Cetirizine; we also evaluated the effect of the drug on cell proliferation by a [3H]thymidine incorporation assay. All of the tested doses of Cetirizine were able to significantly reduce CD54 upregulation induced by gammaIFN; concerning the fibroblast proliferation, we observed a dose-dependent inhibition of [3H]thymidine incorporation. These results show that Cetirizine exerts a biologic effect directly on human airway fibroblasts, suggesting a new rationale in the use of this compound.

Blood eosinophilia and degree of sensitization to house dust mites in preschool and school children with asthma.

In allergic asthma, there is convincing evidence that changes in eosinophil and lymphocyte state of activation in blood may reflect disease activity. We evaluated whether simple blood eosinophil or lymphocyte counts in atopic children with asthma could reflect the degree of allergic sensitization. Seventy-six asthmatic children, sensitized to house dust mites (HDM), in stable conditions at the time of the study, and 53 sex- and age-matched controls (CTR) were studied. As compared to CTR, allergic patients showed higher eosinophil numbers and percentages (p < 0.001) but similar lymphocyte numbers and proportions (p > 0.1). Both in CTR and in allergic patients, eosinophil counts did not correlate with lymphocyte counts (p > 0.05; each comparison) but positive correlations were observed between eosinophil numbers and percentages and paper radio immunosorbent test (PRIST) levels or radio-allergo sorbent test (RAST) classes (p < 0.001; each comparison). When allergic asthmatic individuals were subdivided according to their age into two subgroups (Gr), no differences were found in eosinophil and lymphocyte counts and in PRIST levels and RAST values between Gr1 (< or =5 years old [preschool children]) and Gr2 (>5 years old [school children]) (p > 0.05; each comparison). Interestingly, although positive correlations between eosinophil counts and PRIST levels were found in both subgroups (p < 0.05; each comparison), only in Gr2 did eosinophil counts correlate positively with RAST classes (p < 0.001). No correlations between lymphocyte counts and PRIST levels or RAST classes were demonstrated (p > 0.05; each comparison). These data suggest that although blood eosinophilia was similar in preschool and in allergic asthmatic school children sensitized to HDM, only in the oldest children did blood eosinophil counts appear to be related to the degree of HDM-specific sensitization.