Transition of asthmatic bronchial fibroblasts to myofibroblasts is inhibited by cell–cell contacts

Abstract

The role of airway wall remodelling in bronchial asthma is well established. Myofibroblasts, the cells displaying features intermediate between fibroblasts and smooth muscle cells, are involved in this process but the mechanism of myofibroblasts activation in the onset of the disease remains obscure. Myofibroblasts can differentiate from various cell types, including resident fibroblasts, and the fibroblasts to myofibroblasts transition (FMT) can be reproduced in vitro. We aimed to investigate the process of FMT in human bronchial fibroblasts (HBF) derived from non-asthmatic (n = 7) and asthmatic (n = 7) subjects. We also tested whether cell-cell contacts affect FMT by using N-cadherin blocking antibody. HBF plated in low or high cell density were treated with TGF-β(1) up to one week to induce FMT. The percentage of myofibroblsts was counted and expression of α-smooth muscle actin was evaluated by cytoimmunofluorescence, flow cytometry and immunobloting. We demonstrated that the intensity of FMT induced by TGF-β(1)in vitro was strongly enhanced in asthmatic as compared to non-asthmatic HBF populations. This process was facilitated by low cell plating density in both groups of cultures. Furthermore, we proved that neither HBF-conditioned medium nor growth arrest in G(0)/G(1) phase of cell cycle could stop the TGF-β(1)-induced FMT in asthmatic cell populations. However, even in sparse asthmatic HBF, the blocking of N-cadherin resulted in the inhibition of FMT. Our findings show for the first time that the initial absence or an induced loss of cell-cell adhesions in asthmatic HBF populations is important for the completion of FMT.

Lithium Attenuates TGF-β1-Induced Fibroblasts to Myofibroblasts Transition in Bronchial Fibroblasts Derived from Asthmatic Patients

“…We have previously demonstrated that HBFs derived from patients with bronchial asthma display inherent features in vitro that facilitate FMT in response to prolonged incubation with TGF- β 1 [14, 34]. Here, we aimed to investigate the interference of lithium with TGF- β 1 -induced FMT in HBF populations derived from asthmatic patients (AS, n = 8) and non-asthmatic subjects (NA, n = 8).…”

“…Primary HBF were isolated from bronchial biopsies as described previously [14, 34]. The first group consisted of patients with moderate to severe bronchial asthma (AS group) with a mean age of 43.1 ± 9.5 years (females 64%), the average duration of the disease was 11.3 ± 9.4 years, and decrease in the first second forced expiratory volume (FEV1: 78.4 ± 22.4% of predicted).…”

“…Myofibroblasts were identified by immunodetection of α -SMA as described previously [34]. In brief, cells growing on glass coverslips were fixed in 3.7% paraformaldehyde, permeabilised in 0.1% Triton X-100, blocked with 1% BSA, and incubated with a mouse monoclonal antibody against human α -SMA (clone 1A4, Sigma-Aldrich) and Alexa Fluor 488 goat anti-mouse IgG (clone A11001, Sigma-Aldrich).…”

“…We have previously shown that HBFs derived from patients with asthma displayed more pronounced FMT in response to TGF- β in comparison with nonasthmatic counterparts. This indicates susceptibility towards a TGF- β -induced phenotypic switch, an event crucial for the asthmatic process [14, 34]. In the current study, we aimed to elucidate the role of lithium compounds in TGF- β 1 -induced FMT in HBF populations and the mechanisms responsible for this process.…”

Protomyofibroblast Pathway in Early Thermal Burn Healing

“…Thus, MF differentiation is regulated by the combination of specific cells and a series of ECM molecular components [20,21,22]. Cell-cell contacts [23] and mechanical force transductions [24,25,26,27] participate in the process.…”