Innate generation of thrombin and intracellular oxidants in airway epithelium by allergen Der p 1

Allergen-dependent oxidant formation requires purinoceptor activation of ADAM 10 and prothrombin

“…As demonstrated in our previous work, this effect is due to the proteolytic activity of Der p 1 because it can be replicated by the purified allergen and blocked by potent, selective inhibitors specifically designed to target the proteolytic bioactivity of this allergen E5 . Bacterial LPSs, inevitably present in allergen mixtures derived from natural cultures and occurring as contaminants in purified allergens derived therefrom, are potent activators of ROS formation in numerous cell types.…”

“…Der p 1 from D pteronyssinus , commonly used as the general exemplar of these allergens, has recently been identified as the dominant component in the allergenic repertoire of HDM that stimulates the formation of superoxide anion (O 2 − ) in mitochondria of human airway epithelial cells E5 . The production of this ROS is independent of IgE.…”

“…This is because Der p 1, a cysteine peptidase, behaves as a prothrombinase, thereby triggering canonical activation of protease-activated receptor (PAR) 1 and 4 by thrombin 1 . These events are preventable by Allergen Delivery Inhibitors or antagonists of PAR1 and PAR4 G-protein–coupled receptors 1 .…”

Pathways of airway oxidant formation by house dust mite allergens and viral RNA converge through myosin motors, pannexons and Toll‐like receptor 4

“…The oxidative formation of rhodamine may be accomplished by a range of reactive species generated by different mechanisms, so without prejudice we electively refer to these measurements as intracellular ROS to reflect contributions from multiple sources. Advantageously, dihydrorhodamine accumulates preferentially in mitochondria and our previous work using a DNA‐binding triphenylphosphonium analogue of dihydroethidium and a mitochondrial disruptor indicates that a component of this ROS generation is of mitochondrial origin via formation of . Additionally, dihydrorhodamine is an effective reporter of products formed by the facile, spontaneous decomposition of peroxynitrite arising through the reaction of with nitric oxide.…”

“…Our extensive prior work has established that this cell line responds to HDM allergens through a variety of mechanisms also shown to operate in primary cultures of human airway epithelial cells . Notably, intracellular ROS generation which is dependent upon the cleavage of PAR1 and PAR4 by Der p 1 is a feature we have already identified in primary cultures from human lung and in other immortalized epithelial cell lines from human airway . Gene silencing experiments with siRNA duplexes were performed according to the supplier protocol optimized for these experiments.…”