title: Sodium-calcium exchange in intracellular calcium handling of human airway smooth muscle.
abstract: Enhanced airway contractility following inflammation by cytokines such as tumor necrosis factor alpha (TNFα) or interleukin-13 (IL-13) involves increased intracellular Ca(2+) ([Ca(2+)](i)) levels in airway smooth muscle (ASM). In ASM, plasma membrane Ca(2+) fluxes form a key component of [Ca(2+)](i) regulation. There is now growing evidence that the bidirectional plasma membrane Na(+)/Ca(2+) exchanger (NCX) contributes to ASM [Ca(2+)](i) regulation. In the present study, we examined NCX expression and function in human ASM cells under normal conditions, and following exposure to TNFα or IL-13. Western blot analysis showed significant expression of the NCX1 isoform, with increased NCX1 levels by both cytokines, effects blunted by inhibitors of nuclear factor NF-κB or mitogen-activated protein kinase. Cytokine-mediated increase in NCX1 involved enhanced transcription followed by protein synthesis. NCX2 and NCX3 remained undetectable even in cytokine-stimulated ASM. In fura-2 loaded human ASM cells, NCX-mediated inward Ca(2+) exchange as well as outward exchange (measured as rates of change in [Ca(2+)](i)) was elicited by altering extracellular Na(+) and Ca(2+) levels. Contribution of NCX was verified by measuring [Na(+)](i) using the fluorescent Na(+) indicator SBFI. NCX-mediated inward exchange was verified by demonstrating prevention of rising [Ca(2+)](i) or falling [Na(+)](i) in the presence of the NCX inhibitor KBR7943. Inward exchange-mode NCX was increased by both TNFα and IL-13 to a greater extent than outward exchange. NCX siRNA transfection substantially blunted outward exchange and inward exchange modes. Finally, inhibition of NCX expression or function blunted peak [Ca(2+)](i) and rate of fall of [Ca(2+)](i) following histamine stimulation. These data suggest that NCX-mediated Ca(2+) fluxes normally exist in human ASM (potentially contributing to rapid Ca(2+) fluxes), and contribute to enhanced [Ca(2+)](i) regulation in airway inflammation.
authors.affiliation: Department of Physiology and Biomedical Engineering, Mayo Clinic, Rochester, Minnesota, United States of America.
authors.forename: Venkatachalem
authors.initials: V
authors.lastname: Sathish
authors.forename: Philippe F
authors.initials: PF
authors.lastname: Delmotte
authors.forename: Michael A
authors.initials: MA
authors.lastname: Thompson
authors.forename: Christina M
authors.initials: CM
authors.lastname: Pabelick
authors.forename: Gary C
authors.initials: GC
authors.lastname: Sieck
authors.forename: Y S
authors.initials: YS
authors.lastname: Prakash
mesh_terms: D015153:Blotting, Western; D001980:Bronchi / Q000166:cytology; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D005419:Flavonoids / Q000494:pharmacology; D006801:Humans; D018793:Interleukin-13 / Q000494:pharmacology; D042541:Intracellular Space / Q000187:drug effects / Q000378:metabolism; D017136:Ion Transport / Q000187:drug effects; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D009130:Muscle, Smooth / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000378:metabolism; D010455:Peptides / Q000494:pharmacology; D034622:RNA Interference; D012964:Sodium / Q000378:metabolism*; D019831:Sodium-Calcium Exchanger / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D013890:Thiourea / Q000031:analogs & derivatives / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
chemical_list: C101670:2-(2-(4-(4-nitrobenzyloxy)phenyl)ethyl)isothiourea methanesulfonate; D005419:Flavonoids; D018793:Interleukin-13; D016328:NF-kappa B; D010455:Peptides; C101832:SLC38A3 protein, human; C118258:SN50 peptide; D019831:Sodium-Calcium Exchanger; D014409:Tumor Necrosis Factor-alpha; C110374:sodium-calcium exchanger 1; D012964:Sodium; D020928:Mitogen-Activated Protein Kinases; D013890:Thiourea; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one; D002118:Calcium