title: Regulation of P2X7-dependent inflammatory functions by P2X4 receptor in mouse macrophages.
abstract: Activation of the P2X7 receptor of macrophages plays an important role in inflammation. We recently reported that co-expression of P2X4 receptor with P2X7 receptor facilitates P2X7 receptor-mediated cell death via Ca(2+) influx. However, it remained unclear whether P2X4 receptor is involved in P2X7 receptor-mediated inflammatory responses, such as cytokine production. Here, we present evidence that P2X4 receptor modulates P2X7 receptor-dependent inflammatory functions. Treatment of mouse macrophage RAW264.7 cells with 1mM ATP induced high mobility group box 1 (HMGB1) release and IL-1β production via activation of P2X7 receptor. Knockdown of P2X4 receptor or removal of extracellular Ca(2+) suppressed ATP-induced release of both HMGB1 and IL-1β. On the other hand, knockdown of P2X4 receptor or removal of extracellular Ca(2+) enhanced P2X7-dependent LC3-II expression (an index of autophagy), suggesting that P2X4 receptor suppresses P2X7-mediated autophagy. Since LC3-II expression was inhibited by pretreatment with antioxidant and NADPH oxidase inhibitor, we examined P2X7-mediated production of reactive oxygen species (ROS). We found that activation of P2X7 receptor-mediated production of ROS was significantly facilitated in P2X4-knockdown cells, suggesting that co-expression of P2X4 receptor with P2X7 receptor may suppress anti-inflammatory function-related autophagy via suppression of ROS production. We conclude that co-expression of P2X4 receptor with P2X7 receptor enhances P2X7-mediated inflammation through both facilitation of release of cytokines and suppression of autophagy.
authors.affiliation: Department of Radiation Biosciences, Faculty of Pharmaceutical Sciences, Tokyo University of Science, 2641 Yamazaki, Noda-shi, Chiba, Japan.
authors.forename: Ayumi
authors.initials: A
authors.lastname: Kawano
authors.forename: Mitsutoshi
authors.initials: M
authors.lastname: Tsukimoto
authors.forename: Daisuke
authors.initials: D
authors.lastname: Mori
authors.forename: Taisei
authors.initials: T
authors.lastname: Noguchi
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Harada
authors.forename: Takato
authors.initials: T
authors.lastname: Takenouchi
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Kitani
authors.forename: Shuji
authors.initials: S
authors.lastname: Kojima
mesh_terms: D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D055785:Gene Knockdown Techniques; D024243:HMGB1 Protein / Q000378:metabolism; D007249:Inflammation / Q000378:metabolism*; D053583:Interleukin-1beta / Q000378:metabolism; D008264:Macrophages / Q000378:metabolism*; D051379:Mice; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors; D017382:Reactive Oxygen Species / Q000378:metabolism; D058848:Receptors, Purinergic P2X4 / Q000235:genetics / Q000502:physiology*; D058486:Receptors, Purinergic P2X7 / Q000378:metabolism*; D019323:omega-N-Methylarginine / Q000494:pharmacology
chemical_list: D000975:Antioxidants; D024243:HMGB1 Protein; D053583:Interleukin-1beta; C480860:Map1lc3b protein, mouse; D008869:Microtubule-Associated Proteins; D017382:Reactive Oxygen Species; D058848:Receptors, Purinergic P2X4; D058486:Receptors, Purinergic P2X7; D019323:omega-N-Methylarginine; D019255:NADPH Oxidases; D002118:Calcium