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Relative enlargement of the pulmonary artery (PA) on chest CT imaging is associated with respiratory exacerbations in patients with COPD or cystic fibrosis. We sought to determine whether similar findings were present in patients with asthma and whether these findings were explained by differences in ventricular size.

This article presents an overview of future uses for biologic therapies in the treatment of immunologic and allergic conditions. Discussion is centered on the use of existing therapies outside of their current indication or on new therapies that are close to approval. This information may help familiarize practicing allergists and immunologists with therapies they may soon encounter in their practice as well as help identify conditions and treatments that will require further study in the near future.

MCs and histamine in survival, proliferation, and gene expression of MDSCs in mouse models of Th2 disease. It has been shown recently that MCs are required for differential regulation of the immune response by granulocytic versus monocytic MDSCs. Granulocytic MDSCs promoted parasite clearance, whereas monocytic MDSCs enhanced tumor progression; both activities were abrogated in MC‐deficient mice. Herein, we demonstrate that the lack of MCs also influences MDSC trafficking. Preferential trafficking to the liver was not seen in MC‐deficient mice. In addition, evidence that the MC mediator histamine was important in MDSC trafficking and activation is also shown. MDSCs express HR1–3. Blockade of these receptors by HR1 or HR2 antagonists reversed the histamine enhancement of MDSC survival and proliferation observed in cell culture. In addition, histamine differentially influenced Arg1 and iNOS gene expression in MDSCs and greatly enhanced IL‐4 and IL‐13 message, especially in granulocytic MDSCs. Evidence that histamine influenced activity seen in vitro translated to in vivo when HR1 and HR2 antagonists blocked the effect of MDSCs on parasite expulsion and tumor metastasis. All of these data support the MDSC‐mediated promotion of Th2 immunity, leading to the suggestion that allergic‐prone individuals would have elevated MDSC levels. This was directly demonstrated by looking at the relative MDSC levels in allergic versus control patients. Monocytic MDSCs trended higher, whereas granulocytic MDSCs were increased significantly in allergic patients. Taken together, our studies indicate that MCs and MC‐released histamine are critical for MDSC‐mediated immune regulation, and this interaction should be taken into consideration for therapeutic interventions that target MDSCs.

Allelic variants at codons 16 and 27 of the β2-adrenergic receptor gene (ADRB2) have shown clinical and pharmacological implications in asthma, hypertension, ischemic heart failure, diabetes, obesity, and cystic fibrosis. We have developed a simultaneous genotyping assay for the c.46AG and c.79CG allelic variants using hybridization probes and melting curve analysis. The assay was optimized on a panel of 30 DNA samples of known ADRB2genotype as determined by sequencing with 100% concordance between the two techniques. Melting temperature (Tm) ranges for the different genotypes were obtained using data from three independent experiments. Single peaks for p.Arg16Arg (Tm = 57.76°C ± 0.10°C) and p.Gly16Gly (Tm = 66.73°C ± 0.18°C) and two melting peaks for p.Arg16Gly were obtained. Similarly, single peaks for p.Gln27Gln (Tm = 53.98°C ± 0.19°C) and p.Glu27Glu (Tm = 64.93°C ± 0.16°C) and two peaks for p.Gln27Glu were detected. Independent operators easily assigned genotypes in a sample set of 385 asthmatic patients. Haplotype and allele frequencies were in concordance with previously published data: Arg allele frequencies in children/adults were 0.34/0.30 in Caucasians and 0.45/0.52 in African Americans, and Gln allele frequencies were 0.58/0.52 in Caucasians and 0.82/0.84 in African Americans. Thus, the ADRB2genotyping assay represents a highly reliable and rapid technique for routine clinical use in the simultaneous detection of ADRB2variants.

Uncontrolled asthma in children can result in significant morbidity, activity limitations, and financial burden. Current National Asthma Education and Prevention Program asthma management guidelines recommend that all children with persistent asthma receive longterm inhaled corticosteroids (ICSs) controller therapy. Other controller medications, such as cromolyn and nedocromil, are less effective alternatives to ICSs in children with persistent asthma. Although comparative studies need to be conducted in children, the results of several adult studies suggest that ICSs are more effective than leukotriene modifiers as longterm controller monotherapy in patients with moderate to severe persistent asthma. Early ICS intervention is important in children with persistent asthma, as the greatest losses in lung function typically occur in the early stage of disease. Once initiated, ICS therapy should be administered on a regular basis, because discontinuing ICS use may result in increased asthma symptoms, asthma exacerbations, and rescue medication use. Importantly, numerous studies have demonstrated that, when used at recommended doses, ICSs are safe for long-term use in children with asthma, with no clinically significant effects on growth, bone mineral density, vision, or adrenal function. Physicians should choose the most appropriate device for ICS delivery based on the patient's individual needs. (Pediatr Asthma Allergy Immunol 2005; 18[4]:247–258.)

To review the current data and treatment options for mild persistent asthma.

Because of potential toxicities of inhaled corticosteroid (ICS) use in pediatric asthma, alternative or steroid-sparing therapy is desirable. There are no previous studies evaluating montelukast's steroid-sparing effects in children with asthma.

The initial 12-week, double-blind phases of three studies demonstrated that budesonide inhalation suspension (BIS) is effective and well tolerated in infants and young children (6 months to 8 years of age) with persistent asthma.

Background:Basophils represent an important source of inflammatory mediators and cytokines after IgE-dependent activation in human beings. Objective:To assess the role of basophils in allergic asthma, we measured the number of basophils in the bronchial mucosa and their capacity to express IL-4 mRNA and protein during allergen-induced late asthmatic responses. Methods:Fiberoptic bronchoscopic bronchial biopsies were obtained at 24 hours from sites of segmental bronchial allergen challenge and control sites in 19 patients with atopic asthma and 6 nonatopic healthy volunteers. Basophil numbers were assessed by immunohistochemistry through use of mAb 2D7. IL-4 mRNA–positive cells were detected through use of in situ hybridization and colocalized to basophils through use of sequential immunohistochemistry/in situ hybridization. IL-4 protein was detected and colocalized to basophils through use of dual immunohistochemistry. Results:After allergen challenge, there was an increase in the median number of 2D7-positive basophils per square millimeter in the bronchial mucosa in patients with asthma (0.9 cells/mm2at baseline to 8.8 cells/mm2after challenge; P= .002), which also was significantly higher than what was seen in nonasthmatic controls (P= .01). Similarly, IL-4 mRNA– positive cells were increased at 24 hours in patients with asthma (1.4 to 14) in comparison with controls (0 to 0; P= .02). Colocalization studies revealed that 15% and 41% of the basophil population in patients with asthma after allergen-challenge expressed, respectively, IL-4 mRNA and protein. Conversely, 19% of IL-4 mRNA–positive cells and 72% of IL-4 protein–positive cells were accounted for by basophils. Conclusion:After allergen provocation in sensitive patients with atopic asthma, basophils are recruited to the bronchial mucosa and express IL-4 mRNA and protein, which might contribute to local IgE synthesis and/or tissue eosinophilia or other aspects of allergic inflammation during late responses and ongoing asthma. (J Allergy Clin Immunol 2001;108:205-11.)

Background:Disialoganglioside GD3 is expressed on the surface of selected cell types. Anti-GD3 mAb administered to human subjects with malignant melanoma produces signs and symptoms of immediate hypersensitivity reactions. Objective:The expression of GD3 by human mast cells was assessed during mast cell development in vitro and in samples of lung and skin. Methods:GD3 on tissue- and in vitro–derived mast cells was analyzed after double labeling of cells for tryptase (G3 mAb) or Kit (YB5.B8 mAb) and GD3 (R24 mAb). Glycolipids in extracts of fetal liver–derived mast cells were examined by using high-performance thin-layer chromatography. Results:Flow cytometry showed that the percentage of GD3+cells increased in parallel to Kit+cells during the recombinant human stem cell factor–dependent development of fetal liver–derived mast cells. Double-labeling experiments showed that GD3+cells were also surface Kit+and granule tryptase positive, identifying them as mast cells in preparations of lung-, skin-, fetal liver–, and cord blood–derived cells. The major acidic glycolipid detected was NeuAcα2-8NeuAcα2-3Galβ1-4Glcβ1-1′Cer (GD3). Among peripheral blood leukocytes, only basophils and about 10% of the T cells were labeled with anti-GD3 mAb. Anti-GD3 mAb–conjugated magnetic beads were used to purify mast cells to greater than 90% purity from dispersed skin cells enriched to approximately 12% purity by means of density-dependent sedimentation but were less proficient for dispersed human lung mast cells, most likely because of other cell types that express GD3. Conclusion:GD3 is expressed on the surface of developing human mast cells in parallel to tryptase in secretory granules and, like Kit, can serve as a target for their enrichment by immunoaffinity techniques. (J Allergy Clin Immunol 2001;107:322-30.)

Background:Allergic eye disease is common, but little is known about the underlying disease mechanisms. Conjunctival allergen challenge causes symptoms similar to those of seasonal allergic conjunctivitis and is a useful model to study. Objective:We have used allergen challenge to investigate the course of the ocular response, tear inflammatory mediators, tissue adhesion protein expression, and cellular infiltration. Methods:Eighteen atopic patients and 4 nonatopic control subjects were challenged with extracted mixed grass or Dermatophagoides pteronyssinusin one eye and control vehicle in the other. The clinical response was recorded, and tears were collected over a 6-hour period. Conjunctival biopsy specimens were taken from the challenged eye at 6 or 24 hours. Results:An early-phase response (maximal at 20 minutes) showed a significant increase in tear histamine and tryptase levels, reducing to control levels again by 40 minutes. At 6 hours, a late-phase response occurred with increased symptoms, a second peak of tear histamine and eosinophil cationic protein but not tryptase, upregulation of the adhesion molecules E-selectin and intercellular adhesion molecule, and a cellular infiltrate of mast cells, neutrophils, eosinophils, macrophages, and basophils, with T cells increased only in bulbar biopsy specimens. Conclusions:The early peaks of tear histamine plus tryptase indicate that the mast cell is responsible for the early-phase response, but basophils may be involved in the late-phase response. Both tear and biopsy findings underline the significance of the late-phase response as the transition between a type I response and clinical disease. (J Allergy Clin Immunol 2000;106:948-54.)

Background:Because biopsy criteria for diagnosing systemic mastocytosis are not precise, the value of serum α-protryptase levels in the work-up of suspected systemic mastocytosis should be considered. Objective:A retrospective analysis was performed on subjects with total tryptase serum levels that were high (≥20 ng/mL), while β-tryptase serum levels were normal (<1 ng/mL) or modestly elevated (1 to 5 ng/mL). Methods:Over a 3.5-year period, 52 qualifying specimens were identified from 1369 consecutive samples. The corresponding subjects were divided into those with suspected mastocytosis and those with suspected anaphylaxis. Subjects with suspected mastocytosis were subdivided into 3 subgroups on the basis of biopsy results (positive, negative, or not available). Subjects with suspected anaphylaxis were subdivided into living and deceased subgroups. Results:Among the 15 subjects who underwent biopsy, α-protryptase serum levels (the difference between directly-measured levels of serum total tryptase and β-tryptase), when greater than 75 ng/mL (n = 9), were always associated with a positive biopsy result for systemic mastocytosis; levels from 20 to 75 ng/mL (n = 6) were associated with a positive biopsy result in 50% of subjects. α-Protryptase serum levels may be a more sensitive screening test than a bone marrow biopsy for this disorder. Also, elevated α-protryptase serum levels in some adult patients return to normal over time, suggesting that mast cell hyperplasia resolved in these patients. Finally, a high α-protryptase level may reveal anaphylaxis to be a presenting manifestation of systemic mastocytosis or mast cell hyperplasia. Conclusion:Levels of serum α-protryptase, relative to those of β-tryptase, appear to be useful in the diagnostic work-up and follow-up of subjects with suspected systemic mastocytosis. (J Allergy Clin Immunol 1999;103:1092-9.)

We have previously shown the development in vitro of tryptase+human mast cells from fetal liver cells cocultured with murine 3T3 fibroblasts. In this study, recombinant human stem cell factor (rhuSCF), the ligand for the c-kitproto-oncogene product called Kit, stimulated the growth and differentiation primarily of mast cells from dispersed fetal liver cells, whereas recombinant human interleukin-3 (rhulL-3) stimulated the differentiation of basophils along with other cell types. Cultures of fetal liver cells were initiated and maintained in the presence of rhuSCF or rhulL-3 for up to 6 weeks. Metachromatic cells in cytospins were identified as mast cells primarily on the basis of tryptase expression, and as MCT or MCTC by immunohistochemistry using monoclonal antibodies against tryptase and chymase, whereas basophils were metachromatic, polymorphonuclear, and lacked these proteases. Levels of tryptase and histamine were measured by radioimmunoassay, tryptase and chymase activities by peptide hydrolysis, and cell surface Kit by flow cytometry with the monoclonal antibody YB5.B8. The predominant presence of mast cells occurred only in the cultures supplemented with rhuSCF. The percentage and total number of mast cells increased over time with increasing concentrations of rhuSCF and reached a plateau at 55 ng/mL. At this concentration of rhuSCF, mast cells first appeared by day 7; by day 42, 106% of the starting number of cells were present and 85% of these were tryptase+, 31 % being weakly chymase+. These mast cells appeared immature by ultrastructural criteria; most cells were mononuclear, but some had nuclei with deeply divided lobes. DNA synthesis in tryptase+mast cells at days 21 and 28 of culture with rhuSCF was demonstrated by incorporation of bromodeoxyuridine. Calculated levels of histamine (1.2 pg/mast cell) and tryptase (0.9 pg/mast cell) were similar to those determined previously in coculture experiments with murine 3T3 fibroblasts. Chymase activity was undetectable in most cell extracts. On day 0, 4% to 20% of fetal liver cells expressed cell surface Kit. In the presence of rhuSCF, the percentages and total numbers of Kit+cells and the apparent concentration of Kit per cell increased along with the number of tryptase+cells. In the presence of rhulL-3, toluidine blue+, tryptase– cells first and maximally appeared at day 14 (11% ± 2.5%). The percentage of these toluidine blue+cells then declined to about 6% by days 21 and 35, while the total number of positive cells declined over 10-fold. Kit+cells in the presence of rhulL-3 declined from 9% on day 3 to 2% on day 35. Thus, rhuSCF induces the differentiation of Kit+, tryptase+, toluidine blue+mast cells, whereas rhulL-3 induces the differentiation of Kit-, tryptase-, toluidine blue+basophils. In conclusion, human SCF appears to be a major growth factor for the differentiation of human mast cells.

Human fetal livers contain progenitor cells that become mast cells after 4 weeks of culture with recombinant human stem cell factor. Expression of cell surface CD29 (β1), CD18 (β2), CD61 (β3), and β5 integrins was investigated on such cells by flow cytometry and adhesion measurements. High surface expression of CD49e, CD51, and CD61 along with kit was apparent by 4 weeks of culture, whereas expression of each at day 0 was low to undetectable. CD29 and CD49d were detected on cells from day 0 to 4 weeks of culture; CD49b, CD49c, CD49f, CD18, and CD54 expression was negligible. The fetal liver-derived mast cells spontaneously adhered to vitronectin. No evidence for degranulation was found during vitronectin-dependent adhesion. Adhesion occurred in part through the CD61/CD51 receptor. No evidence for adhesion to vitronectin through CO29 and β5 integrins was obtained. Almost all of the vitronectin-adherent cells expressed CD51, CD61, kit, and tryptase, and exhibited meta-chromasia with toluidine blue. Thus, among the fetal liver-derived cells, developing mast cells were selectively adherent to vitronectin. These mast cells and the other cell types present also adhere spontaneously to fibronectin and to laminin, this adhesion being partially inhibited by antibodies against CD61 and CD29 integrins. In conclusion, human mast cells acquire functional vitronectin receptors as they develop from fetal liver progenitors under the influence of rhSCF. This may be important for the recruitment, localization, and retention of developing mast cells.

Tryptase, a neutral protease, is selectively concentrated in the secretory granules of human mast cells, and its release into the circulation serves as a clinical marker of mast cell activation. The current study describes a new, more sensitive ELISA utilizing a newly developed, mouse monoclonal IgG1 antibody for capture called B12 and capable of detecting tryptase in normal plasma and serum. The greater sensitivity of the new immunoassay results in part from a greater portion of tryptase being detected. Mean levels of tryptase in serum from normal subjects from Richmond, Virginia (4.9 ng/ml;n=56), Munich, Germany (3.8 ng/ml;n=19), and Amersfoort, The Netherlands (1.9 ng/ml;n=8) were as indicated. In 62 subjects with ongoing allergic rhinitis, tryptase levels were no different in serum than for 19 normal controls, indicating that local mast cell activation is not necessarily reflected in the circulation. In 61 subjects sensitive to honey bee or yellow jacket venom by history, the 17 destined to have a severe, hypotensive response to a sting challenge had higher levels of tryptase at baseline than mild reactors, nonreactors, and controls, suggesting that baseline levels of tryptase may predict the severity of the clinical response to allergen in sensitive subjects.

Although interleukin-4 (IL-4) in mice is known to augment the proliferation of mast cells and to modulate the expression of certain mast cell protease transcripts, its effect on human mast cells is less well understood. The current study examined the effects of recombinant human IL-4 (rhulL-4) on stem cell factor (SCF)-dependent fetal liver-derived human mast cells in liquid culture. In no case did rhulL-4 augment proliferation of mast cells. rhulL-4 selectively inhibited certain aspects of the development of mast cells in cultures of fetal liver cells with rhuSCF. These include lower numbers and percentages of cells expressing tryptase and surface Kit, smaller cells, and lower contents of cells for tryptase, histamine, and Kit. Development of metachromasia was not attenuated. The downregulation of Kit, the surface receptor for SCF, is probably a critical factor, because cells lacking this molecule would not be able to respond to SCF. In contrast to mast cell progenitors, mast cells already developed in vitro from fetal liver cells are relatively resistant to rhulL-4, but are still dependent for survival on the presence of rhuSCF.

The effect of age on asthma severity is poorly understood.