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Rhein antagonizes P2X7 receptor in rat peritoneal macrophages.

Hu F, Xing F, Zhu G, Xu G, Li C, Qu J, Lee I, Pan L - Sci Rep (2015)

Bottom Line: These two inhibitory effects of rhein were also observed in rat peritoneal macrophages.Meanwhile, rhein reduced ATP/BzATP-induced IL-1β release in lipopolysaccharide-activated macrophages.Together, our results demonstrate that rhein inhibit ATP/BzATP-induced [Ca(2+)]c increase, pore formation, ROS production, phagocytosis attenuation, IL-1β release and cell apoptosis by antagonizing the P2X7 receptor in rat peritoneal macrophages.

View Article: PubMed Central - PubMed

Affiliation: The Key Laboratory of Weak-Light Nonlinear Photonics of Education Ministry, School of Physics and TEDA Applied Physics Institute, Nankai University, Tianjin, China.

ABSTRACT
P2X7 receptor plays important roles in inflammation and immunity, and thereby it serves as a potential therapeutic target for inflammatory diseases. Rhein, an anthraquinone derivative, exhibits significant anti-inflammatory and immunosuppressive activities in therapy. However, the underlying mechanisms are largely unclear. Here, we aimed to investigate the effects of rhein on P2X7 receptor-mediated responses in vitro. In HEK293 cells expressing rat P2X7 receptor, we first found that rhein concentration-dependently blocked ATP-induced cytosolic calcium concentration ([Ca(2+)]c) elevation and pore formation of the plasma membrane, two hallmarks of the P2X7 receptor activation. These two inhibitory effects of rhein were also observed in rat peritoneal macrophages. Furthermore, rhein counteracted macrophage phagocytosis attenuation and suppressed reactive oxygen species (ROS) production triggered by ATP/BzATP. Meanwhile, rhein reduced ATP/BzATP-induced IL-1β release in lipopolysaccharide-activated macrophages. Prolonged application of ATP caused macrophage apoptosis, while the presence of rhein suppressed this cell cytotoxicity. Such ATP/BzATP-induced cellular reactions were also inhibited by a well-known rat P2X7 receptor antagonist, brilliant blue G, in a similar way to rhein. Together, our results demonstrate that rhein inhibit ATP/BzATP-induced [Ca(2+)]c increase, pore formation, ROS production, phagocytosis attenuation, IL-1β release and cell apoptosis by antagonizing the P2X7 receptor in rat peritoneal macrophages.

No MeSH data available.


Related in: MedlinePlus

Rhein reduced ATP-induced macrophage cell death.(a) Cell viability in control (no ATP, BBG and rhein) and cells exposed to ATP (5 mM) in the presence of rhein (0.1, 0.3, 1, 3, 10 μM) and BBG (0.1, 1, 10 μM), respectively. Data represent the mean ± SD (n = 15 wells for each case). *P < 0.05 compared to control group; #P < 0.05, compared to ATP alone group. (b) The statistic data of the percentage of dead cells in rhein group (ATP alone group was taken as 100%). The smooth curve represents the fit to the Boltzmann equation with an IC50 of 0.40 μM. *P < 0.05 compared to ATP alone group. (c) Cell survival in cells treated with rhein at 0.1, 1, 10 μM, respectively. There was no significant difference between treated cells and control (no rhein) (n = 15 wells for each case).
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f7: Rhein reduced ATP-induced macrophage cell death.(a) Cell viability in control (no ATP, BBG and rhein) and cells exposed to ATP (5 mM) in the presence of rhein (0.1, 0.3, 1, 3, 10 μM) and BBG (0.1, 1, 10 μM), respectively. Data represent the mean ± SD (n = 15 wells for each case). *P < 0.05 compared to control group; #P < 0.05, compared to ATP alone group. (b) The statistic data of the percentage of dead cells in rhein group (ATP alone group was taken as 100%). The smooth curve represents the fit to the Boltzmann equation with an IC50 of 0.40 μM. *P < 0.05 compared to ATP alone group. (c) Cell survival in cells treated with rhein at 0.1, 1, 10 μM, respectively. There was no significant difference between treated cells and control (no rhein) (n = 15 wells for each case).

Mentions: We tested whether rhein prevented macrophage death induced by high extracellular ATP. Stimulation with ATP (5 mM) evidently reduced macrophage viability as summarized in Fig. 7a. However, rhein reversed ATP-induced macrophage death in a concentration-dependent manner, with significant inhibition at concentrations of ≥0.1 μM. Fitting the mean data with the Boltzmann equation derived an IC50 of 0.4 μM (Fig. 7b). In contrast, there was no cytolytic action with concentrations up to 10 μM rhein (Fig. 7c). Besides, the ATP-induced cell death was suppressed by BBG in a similar fashion. These data suggested that rhein inhibited ATP-induced cell death through blocking P2X7 receptors in rat peritoneal macrophages.


Rhein antagonizes P2X7 receptor in rat peritoneal macrophages.

Hu F, Xing F, Zhu G, Xu G, Li C, Qu J, Lee I, Pan L - Sci Rep (2015)

Rhein reduced ATP-induced macrophage cell death.(a) Cell viability in control (no ATP, BBG and rhein) and cells exposed to ATP (5 mM) in the presence of rhein (0.1, 0.3, 1, 3, 10 μM) and BBG (0.1, 1, 10 μM), respectively. Data represent the mean ± SD (n = 15 wells for each case). *P < 0.05 compared to control group; #P < 0.05, compared to ATP alone group. (b) The statistic data of the percentage of dead cells in rhein group (ATP alone group was taken as 100%). The smooth curve represents the fit to the Boltzmann equation with an IC50 of 0.40 μM. *P < 0.05 compared to ATP alone group. (c) Cell survival in cells treated with rhein at 0.1, 1, 10 μM, respectively. There was no significant difference between treated cells and control (no rhein) (n = 15 wells for each case).
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4564849&req=5

f7: Rhein reduced ATP-induced macrophage cell death.(a) Cell viability in control (no ATP, BBG and rhein) and cells exposed to ATP (5 mM) in the presence of rhein (0.1, 0.3, 1, 3, 10 μM) and BBG (0.1, 1, 10 μM), respectively. Data represent the mean ± SD (n = 15 wells for each case). *P < 0.05 compared to control group; #P < 0.05, compared to ATP alone group. (b) The statistic data of the percentage of dead cells in rhein group (ATP alone group was taken as 100%). The smooth curve represents the fit to the Boltzmann equation with an IC50 of 0.40 μM. *P < 0.05 compared to ATP alone group. (c) Cell survival in cells treated with rhein at 0.1, 1, 10 μM, respectively. There was no significant difference between treated cells and control (no rhein) (n = 15 wells for each case).
Mentions: We tested whether rhein prevented macrophage death induced by high extracellular ATP. Stimulation with ATP (5 mM) evidently reduced macrophage viability as summarized in Fig. 7a. However, rhein reversed ATP-induced macrophage death in a concentration-dependent manner, with significant inhibition at concentrations of ≥0.1 μM. Fitting the mean data with the Boltzmann equation derived an IC50 of 0.4 μM (Fig. 7b). In contrast, there was no cytolytic action with concentrations up to 10 μM rhein (Fig. 7c). Besides, the ATP-induced cell death was suppressed by BBG in a similar fashion. These data suggested that rhein inhibited ATP-induced cell death through blocking P2X7 receptors in rat peritoneal macrophages.

Bottom Line: These two inhibitory effects of rhein were also observed in rat peritoneal macrophages.Meanwhile, rhein reduced ATP/BzATP-induced IL-1β release in lipopolysaccharide-activated macrophages.Together, our results demonstrate that rhein inhibit ATP/BzATP-induced [Ca(2+)]c increase, pore formation, ROS production, phagocytosis attenuation, IL-1β release and cell apoptosis by antagonizing the P2X7 receptor in rat peritoneal macrophages.

View Article: PubMed Central - PubMed

Affiliation: The Key Laboratory of Weak-Light Nonlinear Photonics of Education Ministry, School of Physics and TEDA Applied Physics Institute, Nankai University, Tianjin, China.

ABSTRACT
P2X7 receptor plays important roles in inflammation and immunity, and thereby it serves as a potential therapeutic target for inflammatory diseases. Rhein, an anthraquinone derivative, exhibits significant anti-inflammatory and immunosuppressive activities in therapy. However, the underlying mechanisms are largely unclear. Here, we aimed to investigate the effects of rhein on P2X7 receptor-mediated responses in vitro. In HEK293 cells expressing rat P2X7 receptor, we first found that rhein concentration-dependently blocked ATP-induced cytosolic calcium concentration ([Ca(2+)]c) elevation and pore formation of the plasma membrane, two hallmarks of the P2X7 receptor activation. These two inhibitory effects of rhein were also observed in rat peritoneal macrophages. Furthermore, rhein counteracted macrophage phagocytosis attenuation and suppressed reactive oxygen species (ROS) production triggered by ATP/BzATP. Meanwhile, rhein reduced ATP/BzATP-induced IL-1β release in lipopolysaccharide-activated macrophages. Prolonged application of ATP caused macrophage apoptosis, while the presence of rhein suppressed this cell cytotoxicity. Such ATP/BzATP-induced cellular reactions were also inhibited by a well-known rat P2X7 receptor antagonist, brilliant blue G, in a similar way to rhein. Together, our results demonstrate that rhein inhibit ATP/BzATP-induced [Ca(2+)]c increase, pore formation, ROS production, phagocytosis attenuation, IL-1β release and cell apoptosis by antagonizing the P2X7 receptor in rat peritoneal macrophages.

No MeSH data available.


Related in: MedlinePlus