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Exposure of human bronchial epithelial cells to fungal aeroallergens induces calcium mobilization and anion secretion by a mechanism involving oxidative stress (869.3)
Author(s) -
O'Grady Kelly,
Maniak Peter,
Kita Hirohito
Publication year - 2014
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.28.1_supplement.869.3
Subject(s) - dids , chemistry , extracellular , ionomycin , intracellular , egta , calcium in biology , alternaria , channel blocker , secretion , calcium , pharmacology , biochemistry , microbiology and biotechnology , biology , membrane , organic chemistry , genetics
Aeroallergens produced by Alternaria alternata have been shown to induce life threatening exacerbations of airway inflammation in sensitized patients with severe asthma. Inflammation produced by Alternaria is associated with ATP release and purinergic receptor activation. In the present study the effects of Alternaria on signaling pathways and ion transport mechanisms associated with mucociliary clearance were investigated in an immortalized human bronchial epithelial cell line (16HBE14o ‐ cells). Cells were grown on Snapwell TM membrane filters until a stable transepithelial resistance (>500 Ω) was established. Cell monolayers were then voltage clamped at zero mV and apically stimulated with 50 μM Alternaria extract to induce a time‐dependent increase in short circuit current (Isc) consistent with anion secretion. Treatment with the CFTR channel blocker, CFTR inh ‐172 significantly reduced the Isc by ~25%. Furthermore, addition of the disulfonic stilbene compound DIDS (250 μM) abolished the Isc response. Measurements of the effect of Alternaria on intracellular Ca 2+ concentration ([Ca 2+ ] i ) showed that the extract caused a persistent increase in [Ca 2+ ] i that was blocked by chelating extracellular Ca 2+ with EGTA. Pretreatment with 250 μM DIDS also abrogated the effects of Alternaria on [Ca 2+ ] i but did not affect the increase in [Ca 2+ ] i produced by 4 μM ionomycin. Similarly, pretreatment of monolayers with glutathione also blocked the Alternaria induced increase in [Ca 2+ ] i . These results indicate that Alternaria aeroallergens are capable of stimulating CFTR and Ca 2+ ‐dependent anion secretion. Moreover, the increase in [Ca 2+ ] i appears to involve a DIDS‐sensitive uptake mechanism that is blocked when oxidant scavenger molecules are present in the extracellular solution. Grant Funding Source : Supported by NIH: HL110539