Regulation of endothelin-1-induced interleukin-6 production by Ca2+ influx in human airway smooth muscle cells.
ABSTRACT Endothelin-1 is considered to be an important mediator in the pathophysiology of asthma because it induces contraction, hypertrophy, and proliferation in airway smooth muscle cells as well as inflammatory responses in the airway. Airway smooth muscle cells have been suggested to contribute to airway inflammation in asthma by producing cytokines. Nevertheless, the role of intracellular Ca(2+) signal in cytokine production in human airway smooth muscle cells is still unclear. We investigated the mechanisms by which endothelin-1 induces production of interleukin (IL)-6, a pleiotropic cytokine, in primary cultured human airway smooth muscle cells. Levels of IL-6 protein and mRNA were significantly increased by endothelin-1 in dose- and time-dependent manners. Endothelin-1-induced IL-6 production was markedly attenuated by EGTA and various Ca(2+) channel inhibitors such as 3,5-bis(trifluoromethyl)-1H-pyrazole derivative (BTP-2), 1-[beta-[3-(4-methoxyphenyl)propoxy]-4-methoxyphenethyl]-1H-imidazole hydrochloride (SKF96365), and nifedipine. Endothelin-1-induced increases in intracellular Ca(2+) concentrations were significantly inhibited in Ca(2+)-free solution and by BTP-2, SKF96365, and nifedipine. The IL-6 synthesis was also inhibited by the extracellular signal-regulated kinase (ERK)1/2 inhibitor 1,4-diamino-2,3-dicyano-1,4-bis(o-aminophenylmercapto)-butadiene ethanolate (U0126) and the p38 inhibitor 4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)1H-imidazole (SB203580), but not by the c-Jun NH2-terminal kinase inhibitor anthra[1,9-cd]-pyrazol-6-(2H)-one (SP600125). Endothelin-1 significantly upregulated phosphorylation of ERK1/2 and p38 but blocking Ca(2+) influx pathways did not inhibit either upregulation. These findings demonstrate that endothelin-1-induced IL-6 synthesis in airway smooth muscle cells occurs via two parallel but independent events that include Ca(2+) influx and activation of ERK1/2 and p38.
Article: STIM1 Regulates Platelet-Derived Growth Factor-Induced Migration and Ca(2+) Influx in Human Airway Smooth Muscle Cells.[show abstract] [hide abstract]
ABSTRACT: It is suggested that migration of airway smooth muscle (ASM) cells plays an important role in the pathogenesis of airway remodeling in asthma. Increases in intracellular Ca(2+) concentrations ([Ca(2+)](i)) regulate most ASM cell functions related to asthma, such as contraction and proliferation. Recently, STIM1 was identified as a sarcoplasmic reticulum (SR) Ca(2+) sensor that activates Orai1, the Ca(2+) channel responsible for store-operated Ca(2+) entry (SOCE). We investigated the role of STIM1 in [Ca(2+)](i) and cell migration induced by platelet-derived growth factor (PDGF)-BB in human ASM cells. Cell migration was assessed by a chemotaxis chamber assay. Human ASM cells express STIM1, STIM2, and Orai1 mRNAs. SOCE activated by thapsigargin, an inhibitor of SR Ca(2+)-ATPase, was significantly blocked by STIM1 siRNA and Orai1 siRNA but not by STIM2 siRNA. PDGF-BB induced a transient increase in [Ca(2+)](i) followed by sustained [Ca(2+)](i) elevation. Sustained increases in [Ca(2+)](i) due to PDGF-BB were significantly inhibited by a Ca(2+) chelating agent EGTA or by siRNA for STIM1 or Orai1. The numbers of migrating cells were significantly increased by PDGF-BB treatment for 6 h. Knockdown of STIM1 and Orai1 by siRNA transfection inhibited PDGF-induced cell migration. Similarly, EGTA significantly inhibited PDGF-induced cell migration. In contrast, transfection with siRNA for STIM2 did not inhibit the sustained elevation of [Ca(2+)](i) or cell migration induced by PDGF-BB. These results demonstrate that STIM1 and Orai1 are essential for PDGF-induced cell migration and Ca(2+) influx in human ASM cells. STIM1 could be an important molecule responsible for airway remodeling.PLoS ONE 01/2012; 7(9):e45056. · 4.09 Impact Factor
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ABSTRACT: In addition to hyperresponsiveness in asthma, airway smooth muscle (ASM) also manifests an inflammatory phenotype characterized by augmented expression of mediators that enhance inflammation, contribute to tissue remodelling and augment leucocyte trafficking and activity. Our present review summarizes contemporary understanding of ASM-derived mediators and their paracrine and autocrine actions in airway diseases.British Journal of Pharmacology 12/2010; 163(1):68-80. · 4.41 Impact Factor
Article: Store-operated ca(2+) entry is remodelled and controls in vitro angiogenesis in endothelial progenitor cells isolated from tumoral patients.[show abstract] [hide abstract]
ABSTRACT: Endothelial progenitor cells (EPCs) may be recruited from bone marrow to sustain tumor vascularisation and promote the metastatic switch. Understanding the molecular mechanisms driving EPC proliferation and tubulogenesis could outline novel targets for alternative anti-angiogenic treatments. Store-operated Ca(2+) entry (SOCE), which is activated by a depletion of the intracellular Ca(2+) pool, regulates the growth of human EPCs, where is mediated by the interaction between the endoplasmic reticulum Ca(2+)-sensor, Stim1, and the plasmalemmal Ca(2+) channel, Orai1. As oncogenesis may be associated to the capability of tumor cells to grow independently on Ca(2+) influx, it is important to assess whether SOCE regulates EPC-dependent angiogenesis also in tumor patients. The present study employed Ca(2+) imaging, recombinant sub-membranal and mitochondrial aequorin, real-time polymerase chain reaction, gene silencing techniques and western blot analysis to investigate the expression and the role of SOCE in EPCs isolated from peripheral blood of patients affected by renal cellular carcinoma (RCC; RCC-EPCs) as compared to control EPCs (N-EPCs). SOCE, activated by either pharmacological (i.e. cyclopiazonic acid) or physiological (i.e. ATP) stimulation, was significantly higher in RCC-EPCs and was selectively sensitive to BTP-2, and to the trivalent cations, La(3+) and Gd(3+). Furthermore, 2-APB enhanced thapsigargin-evoked SOCE at low concentrations, whereas higher doses caused SOCE inhibition. Conversely, the anti-angiogenic drug, carboxyamidotriazole (CAI), blocked both SOCE and the intracellular Ca(2+) release. SOCE was associated to the over-expression of Orai1, Stim1, and transient receptor potential channel 1 (TRPC1) at both mRNA and protein level The intracellular Ca(2+) buffer, BAPTA, BTP-2, and CAI inhibited RCC-EPC proliferation and tubulogenesis. The genetic suppression of Stim1, Orai1, and TRPC1 blocked CPA-evoked SOCE in RCC-EPCs. SOCE is remodelled in EPCs from RCC patients and stands out as a novel molecular target to interfere with RCC vascularisation due to its ability to control proliferation and tubulogenesis.PLoS ONE 01/2012; 7(9):e42541. · 4.09 Impact Factor