Desensitization of idiopathic pulmonary fibrosis fibroblasts to Alternaria alternata extract-mediated necrotic cell death.

Alternaria alternata is an allergenic fungus and known to cause an upper respiratory tract infection and asthma in humans with compromised immunity. Although A. alternata's effect on airway epithelial cells has previously been examined, the potential role of A. alternata on lung fibroblast viability is not understood. Since lung fibroblasts derived from patients with idiopathic pulmonary fibrosis (IPF) display a distinct phenotype that is resistant to stress and cell death inducing conditions, the investigation of the role of Alternaria on pathological IPF fibroblasts provides a better understanding of the fibrotic process induced by an allergenic fungus. Therefore, we examined cell viability of control and IPF fibroblasts (n = 8 each) in response to A. alternata extract. Control fibroblast cell death was increased while IPF fibroblasts were resistant when exposed to 50-100 μg/mL of A. alternata extract. However, there was no significant difference in kinetics or magnitude of Ca2+ responses from control lung and IPF fibroblasts. In contrast, unlike control fibroblasts, intracellular reactive oxygen species (ROS) levels remained low when IPF cells were treated with A. alternata extracts as a function of time. Caspase 3/7 and TUNEL assay revealed that enhanced cell death caused by A. alternata extract was likely due to necrosis, and 7-AAD assay and the use of sodium pyruvate for ATP generation further supported our findings that IPF fibroblasts become resistant to A. alternata extract-induced necrotic cell death. Our results suggest that exposure to A. alternata potentially worsens the fibrotic process by promoting normal lung fibroblast cell death in patients with IPF.