Adenosine promotion of cellular migration in bronchial epithelial cells is mediated by the activation of cyclic adenosine monophosphate-dependent protein kinase A
ABSTRACT Migration of neighboring cells into the injury is important for rapid repair of damaged airway epithelium. We previously reported that activation of the A(2A )receptors (A(2A)ARs) mediates adenosine-stimulated epithelial wound healing, suggesting a role for adenosine in migration. Because A(2A)AR increases cyclic adenosine monophosphate (cAMP) levels in many cells, we hypothesized that cAMP-dependent protein kinase A (PKA) is involved in adenosine-mediated cellular migration. To test this hypothesis, we stimulated a human bronchial epithelial cell line with adenosine and/or A(2A)AR agonist (5'-(N-cyclopropyl)-carboxamido-adenosine [CPCA]) in the presence or absence of adenosine deaminase inhibitor (erythro-9-(2-hydroxy-3-nonyl) adenine hydrochloride [EHNA]). Cells treated with adenosine or CPCA demonstrated a concentration-dependent increase in migration. Similar results were observed in the presence and absence of EHNA. To confirm A(2A) involvement, we pretreated the cells for 1 hour with the A(2A) receptor antagonist ZM241385 and then stimulated them with either adenosine or CPCA. To elucidate PKA's role, cells were pretreated for 1 hour with either a PKA inhibitor (KT5720) or a cAMP antagonist analogue (Rp-cAMPS) and then stimulated with adenosine and/or CPCA. Pretreatment with KT5720 or Rp-cAMPS resulted in a significant decrease in adenosine-mediated cellular migration. PKA activity confirmed that bronchial epithelial migration requires cAMP and PKA activity. When cells were wounded and stimulated with CPCA, an increase in PKA activity occurred. Pretreatment for 1 hour with either KT5720 or Rp-cAMPS resulted in a significant decrease in adenosine-mediated PKA activation. These data suggest that adenosine activation of A(2A)AR augments epithelial repair by increasing airway cellular migration by PKA-dependent mechanisms.
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ABSTRACT: Extracellular adenosine is a physiologically relevant agonist released by various sources, including endothelial cells (EC) and activated platelets, with complex effects mediated via activation of P1 purinergic receptors. Adenosine-induced EC production of glutathione peroxidase1 and nitric oxide is recognized, and an anti-inflammatory mechanism has been described. Effects of extracellular adenosine on the pulmonary EC barrier function and vascular permeability, however, remain poorly characterized. In this study, we demonstrated the adenosine-induced rapid dose-dependent barrier enhancement in human pulmonary artery EC (HPAEC) as measured by an increase in transendothelial electrical resistance (TER). We have shown that HPAEC express only A2A and A2B adenosine receptors. Pharmacological and siRNA depletion studies indicate that A2A, but not A2B receptor activation is required for the adenosine-induced TER increase. Depletion of Galphas with a specific siRNA significantly attenuated the adenosine-induced TER response in HPAEC. In contrast, depletion of either Galphaq or Galphai2 did not affect the adenosine-induced TER increase. This suggests that the adenosine-induced TER increase is cAMP-dependent. The adenosine-induced barrier enhancement effects were associated with a rearrangement of the EC F-actin component of the cytoskeleton, enhanced cell-surface presentation of cell-cell junctional protein VE-cadherin and an involvement of Myosin-light-chain phosphatase (MLCP). Our results suggest, for the first time, that adenosine regulates the EC barrier function via A2A receptors followed by Galphas engagement and is associated with cytoskeletal activation.Vascular Pharmacology 05/2010; 52(5-6):199-206. DOI:10.1016/j.vph.2009.12.008 · 4.62 Impact Factor
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ABSTRACT: Chronic lung diseases such as asthma, chronic obstructive pulmonary disease and interstitial lung disease are characterized by inflammation and tissue remodeling processes that compromise pulmonary function. Adenosine is produced in the inflamed and damaged lung where it plays numerous roles in the regulation of inflammation and tissue remodeling. Extracellular adenosine serves as an autocrine and paracrine signaling molecule by engaging cell surface adenosine receptors. Preclinical and cellular studies suggest that adenosine plays an anti-inflammatory role in processes associated with acute lung disease, where activation of the A2AR and A2BR has promising implications for the treatment of these disorders. In contrast, there is growing evidence that adenosine signaling through the A1R, A2BR and A3R may serve pro-inflammatory and tissue remodeling functions in chronic lung diseases. This review discusses the current progress of research efforts and clinical trials aimed at understanding the complexities of these signaling pathway as they pertain to the development of treatment strategies for chronic lung diseases.Pharmacology [?] Therapeutics 07/2009; DOI:10.1016/j.pharmthera.2009.04.003 · 7.75 Impact Factor
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ABSTRACT: Lung injury caused by inhalation of dust from swine concentrated animal feeding operations (CAFO) involves the release of inflammatory cytokine interleukin 8 (IL-8), which is mediated by protein kinase C epsilon (PKCε) in airway epithelial cells. Once activated by CAFO dust, PKCε is responsible for slowing cilia beating and reducing cell migration for wound repair. Conversely, the cAMP-dependent protein kinase (PKA) stimulates contrasting effects, such as increased cilia beating and an acceleration of cell migration for wound repair. We hypothesized that a bidirectional mechanism involving PKA and PKC regulates epithelial airway inflammatory responses. To test this hypothesis, primary human bronchial epithelial cells and BEAS-2B cells were treated with hog dust extract (HDE) in the presence or absence of cAMP. PKCε activity was significantly reduced in cells that were pre-treated for one hour with 8-Br-cAMP before exposure to HDE (p<0.05). HDE-induced IL-6 and IL-8 release was significantly lower in cells that were pre-treated with 8-Br-cAMP (p<0.05). To exclude exchange protein activated by cAMP (EPAC) involvement, cells were pre-treated with either 8-Br-cAMP or 8CPT-2Me-cAMP (EPAC agonist). 8CPT-2Me-cAMP did not activate PKA and did not reduce HDE-stimulated IL-6 release. In contrast, 8-Br-cAMP decreased HDE-stimulated TNFα converting enzyme (TACE; ADAM-17) activity and subsequent TNFα release (p <0.001). 8-Br-cAMP also blocked HDE-stimulated IL-6 and keratinocyte-derived chemokine (KC) release in precision-cut mouse lung slices (p<0.05). These data show bidirectional regulation of PKCε via a PKA-mediated inhibition of TACE activity resulting in reduced PKCε-mediated release of IL-6 and IL-8.AJP Lung Cellular and Molecular Physiology 08/2014; 307(8). DOI:10.1152/ajplung.00373.2013 · 4.04 Impact Factor