Extracellular adenosine signaling via the adenosine receptor 2B (AR 2B) is a potent profibrotic and proinflammatory molecule that has been shown to contribute to tissue remodeling and excessive matrix protein production under chronic conditions in kidney disease. Investigation of the adenosine pathway on interstitial renal fibroblasts is lacking even though fibroblasts are a major source of ECM production and contribute to the pathogenesis of renal failure in late stage kidney disease. Analysis of mRNA from the interstitial renal fibroblast cell line NRK-49F revealed these cells express AR1 and AR2B. Utilizing highly selective small molecule agonists and antagonists specific to each individual adenosine receptor, it was determined by mobilization of intracellular cAMP, both receptors were functional. AR 2B was observed to be the dominant receptor under high μM concentrations of agonism leading to increased pro-inflammatory and profibrotic mediators. Specifically, agonism of AR2B lead to increased levels of αSMA and other inflammatory and fibrotic mediators, indicating a transdifferentiation of fibroblasts to a more activated myofibroblasts stage under strong and sustained adenosine signaling. The key fibrotic mediator TGF-β1 was observed to be increased with adenosine signaling while antagonism of AR2B suppressed TGF-β1 mediated increases in αSMA and fibronectin, suggesting complementary signaling pathways with significant crosstalk. It was further observed that AR2B receptor antagonism suppressed TGF-β receptor mediated SMAD binding element associated transcriptional activity the suppression of TGF-β receptor signaling and transcriptional activity was associated with decreased SMAD4, a critical protein necessary for proper formation of the SMAD complex transcription factor. Taken together these data highlight the impact of adenosine signaling on proinflammatory and profibrotic endpoints in renal fibroblasts and describe for the first time direct inhibition of TGF-β receptor signaling by antagonism of the AR2B receptor.
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|Advisor:||Murphy, Suzanne K.|
|Commitee:||Farrell, Francis, Law, Willam R., Morel, Diane|
|School:||University of the Sciences in Philadelphia|
|Department:||Cell and Molecular Biology|
|School Location:||United States -- Pennsylvania|
|Source:||DAI-B 78/11(E), Dissertation Abstracts International|
|Subjects:||Cellular biology, Pharmacology|
|Keywords:||Adenosine, Adenosine receptor, Diabetic nephropathy, Fibrosis, Inflammation, TGF-B1|
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