P2Y2 and P2X4 Receptors Mediate Ca2+ Mobilization in DH82 Canine Macrophage Cells
Purinergic receptors of the P2 subclass are commonly found in human and rodent macrophages where they can be activated by adenosine 5'-triphosphate (ATP) or uridine 5'-triphosphate (UTP) to mediate Ca2+ mobilization, resulting in downstream signalling to promote inflammation and pain. However, little is understood regarding these receptors in canine macrophages. To establish a macrophage model of canine P2 receptor signalling, the expression of these receptors in the DH82 canine macrophage cell line was determined by reverse transcription polymerase chain reaction (RT-PCR) and immunocytochemistry. P2 receptor function in DH82 cells was pharmacologically characterised using nucleotide-induced measurements of Fura-2 AM-bound intracellular Ca2+. RT-PCR revealed predominant expression of P2X4 receptors, while immunocytochemistry confirmed predominant expression of P2Y2 receptors, with low levels of P2X4 receptor expression. ATP and UTP induced robust Ca2+ responses in the absence or presence of extracellular Ca2+. ATP-induced responses were only partially inhibited by the P2X4 receptor antagonists, 2',3'-O-(2,4,6-trinitrophenyl)-ATP (TNP-ATP), paroxetine and 5-BDBD, but were strongly potentiated by ivermectin. UTP-induced responses were near completely inhibited by the P2Y2 receptor antagonists, suramin and AR-C118925. P2Y2 receptor-mediated Ca2+ mobilization was inhibited by U-73122 and 2-aminoethoxydiphenyl borate (2-APB), indicating P2Y2 receptor coupling to the phospholipase C and inositol triphosphate signal transduction pathway. Together this data demonstrates, for the first time, the expression of functional P2 receptors in DH82 canine macrophage cells and identifies a potential cell model for studying macrophage-mediated purinergic signalling in inflammation and pain in dogs.
Medienart: |
E-Artikel |
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Erscheinungsjahr: |
2020 |
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Erschienen: |
2020 |
Enthalten in: |
Zur Gesamtaufnahme - volume:21 |
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Enthalten in: |
International journal of molecular sciences - 21(2020), 22 vom: 13. Nov. |
Sprache: |
Englisch |
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Beteiligte Personen: |
Sophocleous, Reece Andrew [VerfasserIn] |
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Links: |
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Anmerkungen: |
Date Completed 03.03.2021 Date Revised 03.03.2021 published: Electronic Citation Status MEDLINE |
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doi: |
10.3390/ijms21228572 |
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funding: |
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Förderinstitution / Projekttitel: |
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PPN (Katalog-ID): |
NLM317702335 |
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520 | |a Purinergic receptors of the P2 subclass are commonly found in human and rodent macrophages where they can be activated by adenosine 5'-triphosphate (ATP) or uridine 5'-triphosphate (UTP) to mediate Ca2+ mobilization, resulting in downstream signalling to promote inflammation and pain. However, little is understood regarding these receptors in canine macrophages. To establish a macrophage model of canine P2 receptor signalling, the expression of these receptors in the DH82 canine macrophage cell line was determined by reverse transcription polymerase chain reaction (RT-PCR) and immunocytochemistry. P2 receptor function in DH82 cells was pharmacologically characterised using nucleotide-induced measurements of Fura-2 AM-bound intracellular Ca2+. RT-PCR revealed predominant expression of P2X4 receptors, while immunocytochemistry confirmed predominant expression of P2Y2 receptors, with low levels of P2X4 receptor expression. ATP and UTP induced robust Ca2+ responses in the absence or presence of extracellular Ca2+. ATP-induced responses were only partially inhibited by the P2X4 receptor antagonists, 2',3'-O-(2,4,6-trinitrophenyl)-ATP (TNP-ATP), paroxetine and 5-BDBD, but were strongly potentiated by ivermectin. UTP-induced responses were near completely inhibited by the P2Y2 receptor antagonists, suramin and AR-C118925. P2Y2 receptor-mediated Ca2+ mobilization was inhibited by U-73122 and 2-aminoethoxydiphenyl borate (2-APB), indicating P2Y2 receptor coupling to the phospholipase C and inositol triphosphate signal transduction pathway. Together this data demonstrates, for the first time, the expression of functional P2 receptors in DH82 canine macrophage cells and identifies a potential cell model for studying macrophage-mediated purinergic signalling in inflammation and pain in dogs | ||
650 | 4 | |a Journal Article | |
650 | 4 | |a DH82 | |
650 | 4 | |a P2X4 receptor | |
650 | 4 | |a P2Y2 receptor | |
650 | 4 | |a canine | |
650 | 4 | |a dog | |
650 | 4 | |a macrophage | |
650 | 4 | |a neuroinflammation | |
650 | 4 | |a pain | |
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650 | 7 | |a Calcium |2 NLM | |
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700 | 1 | |a Miles, Nicole Ashleigh |e verfasserin |4 aut | |
700 | 1 | |a Ooi, Lezanne |e verfasserin |4 aut | |
700 | 1 | |a Sluyter, Ronald |e verfasserin |4 aut | |
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