Redox-sensitive GPCR signaling drives Gq-dependent Ca2+ mobilization and cytokine production in human bronchial epithelial cells.

Organic dust (OD) from animal production facilities contains a complex mixture of microbial products, metabolites, and particulates that engage airway epithelial signaling pathways. Here we investigated the mechanisms by which OD extract (ODE) activates immune signaling pathways in human bronchial epithelial cells. ODE rapidly stimulate reactive oxygen species (ROS) generation and a biphasic increase in intracellular Ca2+ concentration ([Ca2+]i), consisting of an early transient peak followed by a smaller sustained phase. Antioxidant scavenger pretreatment (glutathione, N-acetyl-cysteine) markedly attenuated both ROS production and Ca2+ mobilization, whereas induction of endogenous antioxidant defenses with bardoxolone abolished the response, indicating redox sensitivity. Pharmacologic inhibition of Gqα with YM-254890 suppressed both phases of the Ca2+ response, implicating Gq-coupled receptor activation. Consistent with an autocrine amplification mechanism, selective antagonists of histamine (H1), cysteinyl leukotriene (CysLT1 and CysLT2), leukotriene B4 (BLT1), and prostaglandin receptors (EP1) each reduced ODE-evoked Ca2+ mobilization. In parallel, inhibitors of histidine decarboxylase, 5-lipoxygenase, and cyclooxygenases (COX-1/COX-2) attenuated Ca2+ signaling, supporting rapid endogenous ligand production and secretion. Downstream of Ca2+ mobilization, ODE activated protein kinase C alpha/beta (PKCα/β) and protein kinase C delta (PKCδ) and induced robust transcription of proinflammatory cytokine and chemokine mRNAs (IL1β, IL6, IL8, IL33, TNFα) within 2 h of exposure. Enzyme-linked immunosorbent assay confirmed increased secretion of IL-1β, IL-6, IL-8, and IL-33, with differential sensitivity to PKC isoforms and NF-κB inhibition. These findings identify a redox-sensitive G-protein coupled receptor (GPCR) network that amplifies Gq-dependent Ca2+ signaling in airway epithelial cells and provides a mechanistic framework for epithelial inflammatory activation following ROS-inducing environmental exposures.NEW & NOTEWORTHY This study investigates how agricultural organic dust promotes airway inflammation. We show that organic dust extract rapidly induces ROS production along with a biphasic Ca2+ response through a redox-sensitive, Gq-coupled G-protein coupled receptor (GPCR) network involving histamine, leukotrienes, and prostaglandins. This signaling cascade activates PKC isoforms that trigger production and secretion of inflammatory cytokines, linking oxidative stress to Ca2+-dependent inflammatory responses by the airway epithelium following organic dust exposure.