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Advisor(s)
Abstract(s)
Chronic musculoskeletal pain involves connective tissue remodeling triggered by inflammatory
mediators, such as bradykinin. Fibroblast cells signaling involve changes in intracellular Ca2+ ([Ca2+]i). ATP has
been related to connective tissue mechanotransduction, remodeling and chronic inflammatory pain, via P2
purinoceptors activation. Here, we investigated the involvement of ATP in bradykinin-induced Ca2+ signals in
human subcutaneous fibroblasts.
Bradykinin, via B2 receptors, caused an abrupt rise in [Ca2+]i to a peak that declined to a plateau, which
concentration remained constant until washout. The plateau phase was absent in Ca2+-free medium; [Ca2+]i
signal was substantially reduced after depleting intracellular Ca2+ stores with thapsigargin. Extracellular ATP
inactivation with apyrase decreased the [Ca2+]i plateau. Human subcutaneous fibroblasts respond to
bradykinin by releasing ATP via connexin and pannexin hemichannels, since blockade of connexins, with 2-
octanol or carbenoxolone, and pannexin-1, with 10Panx, attenuated bradykinin-induced [Ca2+]i plateau,
whereas inhibitors of vesicular exocytosis, such as brefeldin A and bafilomycin A1, were inactive. The kinetics
of extracellular ATP catabolism favors ADP accumulation in human fibroblast cultures. Inhibition of
ectonucleotidase activity and, thus, ADP formation from released ATP with POM-1 or by Mg2+ removal from
media reduced bradykinin-induced [Ca2+]i plateau. Selective blockade of the ADP-sensitive P2Y12 receptor with
AR-C66096 attenuated bradykinin [Ca2+]i plateau, whereas the P2Y1 and P2Y13 receptor antagonists,
respectively MRS 2179 and MRS 2211, were inactive. Human fibroblasts exhibited immunoreactivity against
connexin-43, pannexin-1 and P2Y12 receptor.
Bradykinin induces ATP release from human subcutaneous fibroblasts via connexin and
pannexin-1-containing hemichannels leading to [Ca2+]i mobilization through the cooperation of B2 and P2Y12
receptors.
Description
Keywords
ADP ATP release Bradykinin Cx43 Hemichannels Human subcutaneous fibroblasts Panx1 P2 purinoceptors
Citation
Publisher
BioMed Central