There were no significant changes in clinical data in the HS group

There were no significant changes in clinical data in the HS group. Table 2 Clinical characteristics of the study population after bronchial challenge with 0.05 compared to data before allergen challenge. over the AA group. However, iEOS survivability from AA and SNEA groups was higher compared with rEOS under standard conditions, when rEOS survivability increased after their incubation with ASM cells. Bronchial allergen challenge abolished the dominance of blood iEOS in AA patients and prolonged only iEOS survivability. Though the challenge did not affect the adhesion of any eosinophils subtypes, the direct dependence of rEOS and iEOS survivability on their conversation with ASM cells was revealed ( 0.05). These findings provide the premise for eosinophils subtype-oriented asthma treatment. allergen and positive bronchial challenge with methacholine. The SNEA group consisted of the patients with an asthma diagnosis for at least 12 months, and a non-allergic phenotype confirmed by unfavorable allergy history, skin prick tests and no specific allergy symptoms, such as watery runny nose, nasal obstruction, urticaria, rashes, conjunctivitis, without dietary restrictions and any symptoms of digestion. Peripheral blood eosinophil counts were higher than 0.3 109/L during the screening visit or higher than 0.15 109/L if there was a documented eosinophil count higher than 0.3 109/L in the 12 months before the screening. A severe course of the disease was approved with at least a 12-month treatment of high doses of inhaled steroids combined with long-acting beta-agonist long-acting antimuscarinic agent episodic use of oral corticosteroids. The HS was without allergic (had no symptoms of allergy) and other chronic respiratory diseases, with the unfavorable methacholine test. For all groups, the following exclusion criteria were used: clinically significant allergy symptoms, active airway contamination 1 month before the study, exacerbation 1 month before study, use of oral steroids 1 month before study, and smoking. Inclusion and exclusion criteria provided in Physique 1. Open in a separate windows Physique 1 Inclusion and exclusion criteria of the study populace. All recruited individuals were newly selected subjects. The inclusion and exclusion were criteria were verified after the screening visit. AAAllergic asthma; CBCComplete blood count; HSHealthy subjects; SNEASevere non-allergic eosinophilic asthma. 2.3. Study Design and Experimental Plan SNEA patients were asked to visit the clinic once, and AA patients and HS patients were asked to visit twice (at baseline and 24 h after bronchial allergen challenge). All study individuals were invited into the study no earlier than 3 days (well-planned preparation of experiments for each study subject was required), but no later than 2 weeks after their inclusion and exclusion criteria were confirmed. At first visit, for all study subjects, peripheral blood was collected and measured for exhaled fractional exhaled nitric oxide (FeNO). Additionally, AA patients and HS after primary collection of peripheral blood underwent a bronchial challenge with allergen. Initially, the peripheral blood was used for the isolation of granulocytes. Isolated granulocytes were counted and assessed their viability to perform the first quality control of isolation procedures (at least 98% of granulocytes viability and total cells count of 4 107). Compliant samples were used for eosinophils enrichment. The second quality control of isolation procedures was performed to isolated eosinophilseosinophils were counted, evaluated their viability, TAK-715 and assessed purity control by flow cytometer (forward and side light scattering). Approved samples ( 1.5 106/20 TAK-715 mL blood), viability ( 98%), and purity ( 96%) were used for PKN1 eosinophils subtyping. Collected iEOS and rEOS samples were used for third quality control of isolation procedures ( 0.5 106 cells), viability ( 97%). After eosinophils, subtyping combined cell cultures with healthy immortalized ASM cells were prepared immediately, and their adhesive properties were tested after 1 h when adhesion-related viabilityafter 24 h of incubation. The second visit was 24 h after TAK-715 the bronchial allergen challenge for AA and HS subjects, and all procedures were repeated according to baseline. The study design is usually shown in Physique 2. Open in a separate window Physique 2 Experimental study design. AAAllergic asthma; ASMAirway smooth muscle; FeNOFractional exhaled nitric oxide; HSHealthy subjects; iEOSinflammatory eosinophils; rEOSlung-resident eosinophils. 2.4. Lung TAK-715 Function Testing Pulmonary function was tested using an ultrasonic spirometer (Ganshorn Medizin Electronic, Niederlauer, Germany). TAK-715 The results of forced expiratory volume in 1 s (FEV1), forced vital capacity (FVC), and the FEV1/FVC ratio was considered as the highest of three impartial measurements. The data were compared with the.