Mechanisms underlying TNF-alpha effects on agonist-mediated calcium homeostasis in human airway smooth muscle cells.
We have previously shown that tumor necrosis factor (TNF)-alpha, a cytokine involved in asthma, enhances Ca2+ responsiveness to bronchoconstrictor agents in cultured human airway smooth muscle (ASM) cells. In the present study, we investigated the potential mechanism(s) by which TNF-alpha modulates ASM cell responsiveness to such agents. In human ASM cells loaded with fura 2, TNF-alpha and interleukin (IL)-1 beta significantly enhanced thrombin- and bradykinin-evoked elevations of intracellular Ca2+. In TNF-alpha-treated cells. Ca2+ responses to thrombin and bradykinin were 350 +/- 14 and 573 +/- 93 nM vs. 130 +/- 17 and 247 +/- 48 nM in nontreated cells, respectively (P < 0.0001). In IL-1 beta-treated cells, the Ca2+ response to bradykinin was 350 +/- 21 vs. 127 +/- 12 nM in nontreated cells (P < 0.0001). The time course for TNF-alpha potentiation of agonist-induced Ca2+ responses requires a minimum of 6 h and was maximum after 12 h of incubation. In addition, cycloheximide, a protein synthesis inhibitor, completely blocked the potentiating effect of TNF-alpha on Ca2+ signals. We also found that TNF-alpha significantly enhanced increases in phosphoinositide (PI) accumulation induced by bradykinin. The percentage of change in PI accumulation over control was 115 +/- 8 to 210 +/- 15% in control cells vs. 128 +/- 10 to 437 +/- 92% in TNF-alpha-treated cells for 3 x 10(-9) to 3 x 10(-6) M bradykinin. The PI turnover to 10 mM NaF, a direct activator of G proteins, was also found to be enhanced by TNF-alpha. The percentage of change in PI accumulation over control increased from 280 +/- 35% in control cells to 437 +/- 92% in TNF-alpha-treated cells. Taken together, these results show that TNF-alpha can potently regulate G protein-mediated signal transduction in ASM cells by activating pathways dependent on protein synthesis. Our study demonstrates one potential mechanism underlying the enhanced Ca2+ response to bronchoconstrictor agents induced by cytokines in human ASM cells.
Amrani
Y
Y
Department of Medicine, University of Pennsylvania Medical Center, Philadelphia 19104, USA.
Krymskaya
V
V
Maki
C
C
Panettieri
R A
RA
D001920:Bradykinin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D003513:Cycloheximide / Q000494:pharmacology; D003600:Cytosol / Q000378:metabolism; D006706:Homeostasis / Q000187:drug effects; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology; D007700:Kinetics; D009130:Muscle, Smooth / Q000187:drug effects / Q000502:physiology*; D011189:Potassium Chloride / Q000494:pharmacology; D013917:Thrombin / Q000494:pharmacology; D013997:Time Factors; D014132:Trachea / Q000187:drug effects / Q000502:physiology*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D007375:Interleukin-1; D014409:Tumor Necrosis Factor-alpha; D011189:Potassium Chloride; D003513:Cycloheximide; D013917:Thrombin; D001920:Bradykinin; D002118:Calcium