Different airway smooth muscle cells phenotypes are involved in intrinsic and extrinsic obstruction in guinea pig asthma model

Abstract

Background: Airway obstruction in asthma is driven by airway smooth muscle (ASM) contraction. Airway obstruction can be induced extrinsically by direct stimulation of ASM with contractile agonists or by indirect provocation with antigens, while the airway baseline tone is dependent on intrinsic obstruction. The ASM phenotypes involved in all types of obstruction seem to be related.Methods: To determination the associations of the ASM phenotypes involved in different types of airway obstruction, guinea pigs were sensitized to ovalbumin and repetitively challenged with antigen. At the third challenge, histamine provocation was used to evaluate airway responsiveness (AR), and lung samples were obtained to calculate the airway wall area. ASM cells from the trachea were disaggregated to determine 1) the percentage of cells that expressed transforming growth factor-β1 (TGF-β1), interleukin-13 (IL-13) and sarco-endoplasmic Ca2+ ATPase-2b (SERCA2b) by flow cytometry; 2) SERCA2B gene expression by RT-PCR; 3) the level of reduced glutathione (GSH) by ELISA; and 4) the sarcoplasmic reticulum Ca2+ refilling rate by microfluorometry. The control guinea pig group received only saline instead of ovalbumin. Comparisons were made using t-tests, and the associations were determined using Spearman correlation coefficient analysis.Results: Antigenic challenges induced airway obstruction and progressive incremental changes in airway baseline tone. The AR to histamine and the expression of TGF-β1 in ASM cells was increased in the asthma model. The airway wall mass and expression of IL-13 and SERCA2b in ASM cells were similar between groups. SERCA2B gene expression and GSH levels were reduced in the asthma group. The extent of antigen-induced airway obstruction was directly associated with ASM cell TGF-β1 expression and the degree of AR. The magnitude of AR and antigen-induced airway obstruction showed an inverse correlation with GSH levels. The airway baseline tone showed an inverse association with SERCA2b expression. No relationship was observed between direct or indirect airway obstruction and the airway tone. After caffeine withdrawal, the rate of sarcoplasmic reticulum Ca2+ refilling was similar in both groups.Conclusions: Each type of airway obstruction depends on different ASM phenotypes: 1) direct and indirect airway obstruction seems to be sensitive to the level of ASM oxidative stress; 2) indirect obstruction induced by antigen appears to be influenced by the expression of TGF-β1 in ASM; and 3) the SERCA2b expression level in ASM cells plays a role in the intrinsic airway tone.