Aporocactus flagelliformis water extract and limonin suppresses P2Y purinoceptor 14-mediated proinflammatory features in 3D4/31 porcine alveolar macrophages

Respiratory diseases have been recognized as a significant cause of reduced livestock productivity since 1995. Respiratory diseases in the swine industry caused by both biological and non-biological factors are collectively referred to as porcine respiratory disease complex (PRDC). However, there is a lack of eco-friendly anti-inflammatory drugs (AIDs) that can effectively control lung inflammation caused by PRDC. Additionally, P2Y purinoceptor 14 (P2Y₁₄) has been identified as a major regulator of macrophage inflammatory responses, and it has been reported that porcine-P2Y₁₄ recognizes uridine-5'-diphosphate-glucose (UDPG), similar to human-P2Y₁₄. However, the regulatory role of P2Y₁₄ in porcine inflammation remains unclear. In this study, we demonstrated the development of AIDs applicable to PRDC using the Mexican medicinal plant <italic>Aporocactus flagelliformis</italic> and 3D4/31 porcine alveolar macrophages (PAMs). We measured intracellular reactive oxygen species (ROS) generation and autophagic activity using flow cytometry. Protein fold changes were measured by immunoblotting, and inflammation-related gene expression was quantified by real-time polymerase chain reaction. <italic>Aporocactus flagelliformis</italic> water extract (AFWE) reduced inflammatory features, including ROS generation, autophagy, and proinflammatory cytokine expression in 3D4/31-PAMs. Glycogen accumulation and <italic>signal transducer and activator of transcription 1</italic> (<italic>STAT1</italic>) expression reported in P2Y₁₄-mediated inflammatory responses were also observed in 3D4/31-PAMs, and were reduced by AFWE treatment. Limonin, a major anti-inflammatory compound identified in AFWE, reduced <italic>P2RY14</italic> and proinflammatory gene expression induced by the P2Y₁₄ ligand UDPG in 3D4/31-PAMs, demonstrating an inhibitory effect on porcine-P2Y₁₄-mediated inflammation. These results suggest that P2Y₁₄ is a proinflammatory receptor in PAMs and an effective target for AID development. We propose that AFWE and limonin are AID candidates for inhibiting P2Y₁₄-mediated inflammation in PAMs.