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Immunology - Molecular Biology - Physiology - Respiratory Medicine

Sodium-Calcium Exchange in Intracellular Calcium Handling of Human Airway Smooth Muscle
Published: Monday, August 15, 2011
Author: Venkatachalem Sathish et al.

by Venkatachalem Sathish, Philippe F. Delmotte, Michael A. Thompson, Christina M. Pabelick, Gary C. Sieck, Y. S. Prakash

Enhanced airway contractility following inflammation by cytokines such as tumor necrosis factor alpha (TNFa) or interleukin-13 (IL-13) involves increased intracellular Ca2+ ([Ca2+]i) levels in airway smooth muscle (ASM). In ASM, plasma membrane Ca2+ fluxes form a key component of [Ca2+]i regulation. There is now growing evidence that the bidirectional plasma membrane Na+/Ca2+ exchanger (NCX) contributes to ASM [Ca2+]i regulation. In the present study, we examined NCX expression and function in human ASM cells under normal conditions, and following exposure to TNFa 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-?B 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 Ca2+ exchange as well as outward exchange (measured as rates of change in [Ca2+]i) was elicited by altering extracellular Na+ and Ca2+ 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 [Ca2+]i or falling [Na+]i in the presence of the NCX inhibitor KBR7943. Inward exchange-mode NCX was increased by both TNFa 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 [Ca2+]i and rate of fall of [Ca2+]i following histamine stimulation. These data suggest that NCX-mediated Ca2+ fluxes normally exist in human ASM (potentially contributing to rapid Ca2+ fluxes), and contribute to enhanced [Ca2+]i regulation in airway inflammation.