NCT01759849

Effect of in Vitro Blocking the Common Beta Chain on Cell Viability in Asthma

Effect of in Vitro Blocking the Common Beta Chain on the Function of Blood, Bone Marrow and Sputum Cells Collected From Asthmatic Donors.

• Status: Completed
• Phase: —
• Study type: Observational
• Enrollment: 10 (actual)
• Sponsor: Hamilton Health Sciences Corporation · Academic / Other
• Sex: All
• Age: 18 Years – 65 Years
• Healthy volunteers: Accepted

Summary

This study examines in vitro blockade of signaling through the β-chain, on viability, activation and differentiation of eosinophils and their progenitors collected in sputum, blood and bone marrow samples pre and post-allergen challenge from mild atopic asthmatic subjects.

Detailed description

The experiments will use sputum samples induced from subjects with mild asthma, undergoing allergen inhalation challenges. In general, each sample will be composed of \>10% neutrophils and \>10% macrophages. Samples collected pre-allergen will have a low frequency of eosinophils and lymphocytes (\<1%), however the percentage of eosinophils will increase to approximately 12% following allergen challenges. The sputum samples will be processed in DPBS (without dithiothreitol), and the cell suspension will be adjusted to 1 million cells/ml in DMEM with penicillin and streptomycin. A cytospin will be made for differential cell counts. The mixed cell population at 5 million cells/ml will be incubated for 48 hours at 37 degrees Celcius ± β-chain MAb at a concentration of 100 mcg/ml. After 48 hours the cell culture medium will be removed for assay of cytokines and chemokines by ELISA. Cells will be resuspended in PBS and Binding Buffer (BD Pharmingen, Cat no. 556454),stained for assessments by flow cytometry, and analyzed in duplicate. Experiments will use blood (80 ml) and bone marrow aspirates (5ml) from atopic asthmatics taken pre and 24hr post allergen challenge. Methylcult micro-culture colonogenic assays will be performed to enumerate outgrowth of Eo/Baso-CFU and GM-CFU from CD34+ cells populations collected from the blood and bone marrow samples. Methylcult assays will be performed with CD34+ enriched cell populations in the presence of IL-5,IL-3 and GM-CSF +/- CSL311. Following 14 days culture, colonies will be enumerated.

Conditions

• Asthma

Interventions

Type	Name	Description
OTHER	Allergen	Allergen extract is administered by inhalation.

Timeline

• Start date: 2012-11-01
• Primary completion: 2014-09-01
• Completion: 2014-09-01
• First posted: 2013-01-03
• Last updated: 2017-07-13

Locations

1 site across 1 country: Canada