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Hydroxysafflor Yellow A Inhibits LPS-Induced NLRP3 Inflammasome Activation via Binding to Xanthine Oxidase in Mouse RAW264.7 Macrophages.

Xu X, Guo Y, Zhao J, Wang N, Ding J, Liu Q - Mediators Inflamm. (2016)

Bottom Line: In this study, we predicted the potential interaction between HSYA and xanthine oxidase (XO) via PharmMapper inverse docking and confirmed the binding inhibition via inhibitory test (IC50 = 40.04 μM).Computation docking illustrated that, in this HSYA-XO complex, HSYA was surrounded by Leu 648, Leu 712, His 875, Leu 873, Ser 876, Glu 879, Phe 649, and Asn 650 with a binding energy of -5.77 kcal/M and formed hydrogen bonds with the hydroxyl groups of HSYA at Glu 879, Asn 650, and His 875.Moreover, we proved that HSYA markedly inhibited LPS-induced cleaved caspase-1 activation via suppressing the sensitization of NLRP3 inflammasome and prevented the mature IL-1β formation from pro-IL-1β form.

View Article: PubMed Central - PubMed

Affiliation: Beijing Hospital of Traditional Chinese Medicine, Capital Medical University, Beijing 100010, China; Beijing Institute of Traditional Chinese Medicine, Beijing 100010, China; Beijing Key Laboratory of Basic Research with Traditional Chinese Medicine on Infectious Diseases, Beijing 100010, China.

ABSTRACT
Hydroxysafflor yellow A (HSYA) is an effective therapeutic agent for inflammatory diseases and autoimmune disorders; however, its regulatory effect on NLRP3 inflammasome activation in macrophages has not been investigated. In this study, we predicted the potential interaction between HSYA and xanthine oxidase (XO) via PharmMapper inverse docking and confirmed the binding inhibition via inhibitory test (IC50 = 40.04 μM). Computation docking illustrated that, in this HSYA-XO complex, HSYA was surrounded by Leu 648, Leu 712, His 875, Leu 873, Ser 876, Glu 879, Phe 649, and Asn 650 with a binding energy of -5.77 kcal/M and formed hydrogen bonds with the hydroxyl groups of HSYA at Glu 879, Asn 650, and His 875. We then found that HSYA significantly decreased the activity of XO in RAW264.7 macrophages and suppressed LPS-induced ROS generation. Moreover, we proved that HSYA markedly inhibited LPS-induced cleaved caspase-1 activation via suppressing the sensitization of NLRP3 inflammasome and prevented the mature IL-1β formation from pro-IL-1β form. These findings suggest that XO may be a potential target of HSYA via direct binding inhibition and the combination of HSYA-XO suppresses LPS-induced ROS generation, contributing to the depression of NLRP3 inflammasome and inhibition of IL-1β secretion in macrophages.

No MeSH data available.


Related in: MedlinePlus

HSYA inhibits LPS-induced IL-1β and IL-18 secretion. RAW264.7 cells were pretreated with HSYA at indicated doses (25, 50, and 100 μM) for 3 h and then treated with 1 μg/mL LPS for 12 h (Western Blot) or 24 h (ELISA). (a) The pro-IL-1β, IL-1β, and GAPDH proteins expressions were analyzed by Western Blotting. (b) The IL-1β and IL-18 secretion were measured by ELISA. Data represent the mean ± SEM of three independent experiments and differences between mean values were assessed by one-way ANOVA. ∗P < 0.05, ∗∗P < 0.01, #P < 0.05, and ##P < 0.01 indicate significant differences compared with the control group of indicated proteins, respectively.
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fig8: HSYA inhibits LPS-induced IL-1β and IL-18 secretion. RAW264.7 cells were pretreated with HSYA at indicated doses (25, 50, and 100 μM) for 3 h and then treated with 1 μg/mL LPS for 12 h (Western Blot) or 24 h (ELISA). (a) The pro-IL-1β, IL-1β, and GAPDH proteins expressions were analyzed by Western Blotting. (b) The IL-1β and IL-18 secretion were measured by ELISA. Data represent the mean ± SEM of three independent experiments and differences between mean values were assessed by one-way ANOVA. ∗P < 0.05, ∗∗P < 0.01, #P < 0.05, and ##P < 0.01 indicate significant differences compared with the control group of indicated proteins, respectively.

Mentions: To further estimate the inhibition of HSYA on NLRP3 inflammasome activation, we checked the expression of NLRP3 downstream cytokines IL-1β and IL-18. As shown in Figure 8(a), HSYA treatment at concentrations from 50 to 100 μM notably decreased the protein expression of IL-1β in a dose-dependent manner (P < 0.05). Results based on Western Blot also displayed that 100 μM HSYA possessed an observable negative regulatory effect on pro-IL-1β expression, but with no significant difference compared with LPS group. Moreover, data in Figure 8(b) showed that, as expected, HSYA treatment at concentrations from 50 to 100 μM markedly decreased the secretion level of IL-1β and IL-18 in supernatant (P < 0.05).


Hydroxysafflor Yellow A Inhibits LPS-Induced NLRP3 Inflammasome Activation via Binding to Xanthine Oxidase in Mouse RAW264.7 Macrophages.

Xu X, Guo Y, Zhao J, Wang N, Ding J, Liu Q - Mediators Inflamm. (2016)

HSYA inhibits LPS-induced IL-1β and IL-18 secretion. RAW264.7 cells were pretreated with HSYA at indicated doses (25, 50, and 100 μM) for 3 h and then treated with 1 μg/mL LPS for 12 h (Western Blot) or 24 h (ELISA). (a) The pro-IL-1β, IL-1β, and GAPDH proteins expressions were analyzed by Western Blotting. (b) The IL-1β and IL-18 secretion were measured by ELISA. Data represent the mean ± SEM of three independent experiments and differences between mean values were assessed by one-way ANOVA. ∗P < 0.05, ∗∗P < 0.01, #P < 0.05, and ##P < 0.01 indicate significant differences compared with the control group of indicated proteins, respectively.
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig8: HSYA inhibits LPS-induced IL-1β and IL-18 secretion. RAW264.7 cells were pretreated with HSYA at indicated doses (25, 50, and 100 μM) for 3 h and then treated with 1 μg/mL LPS for 12 h (Western Blot) or 24 h (ELISA). (a) The pro-IL-1β, IL-1β, and GAPDH proteins expressions were analyzed by Western Blotting. (b) The IL-1β and IL-18 secretion were measured by ELISA. Data represent the mean ± SEM of three independent experiments and differences between mean values were assessed by one-way ANOVA. ∗P < 0.05, ∗∗P < 0.01, #P < 0.05, and ##P < 0.01 indicate significant differences compared with the control group of indicated proteins, respectively.
Mentions: To further estimate the inhibition of HSYA on NLRP3 inflammasome activation, we checked the expression of NLRP3 downstream cytokines IL-1β and IL-18. As shown in Figure 8(a), HSYA treatment at concentrations from 50 to 100 μM notably decreased the protein expression of IL-1β in a dose-dependent manner (P < 0.05). Results based on Western Blot also displayed that 100 μM HSYA possessed an observable negative regulatory effect on pro-IL-1β expression, but with no significant difference compared with LPS group. Moreover, data in Figure 8(b) showed that, as expected, HSYA treatment at concentrations from 50 to 100 μM markedly decreased the secretion level of IL-1β and IL-18 in supernatant (P < 0.05).

Bottom Line: In this study, we predicted the potential interaction between HSYA and xanthine oxidase (XO) via PharmMapper inverse docking and confirmed the binding inhibition via inhibitory test (IC50 = 40.04 μM).Computation docking illustrated that, in this HSYA-XO complex, HSYA was surrounded by Leu 648, Leu 712, His 875, Leu 873, Ser 876, Glu 879, Phe 649, and Asn 650 with a binding energy of -5.77 kcal/M and formed hydrogen bonds with the hydroxyl groups of HSYA at Glu 879, Asn 650, and His 875.Moreover, we proved that HSYA markedly inhibited LPS-induced cleaved caspase-1 activation via suppressing the sensitization of NLRP3 inflammasome and prevented the mature IL-1β formation from pro-IL-1β form.

View Article: PubMed Central - PubMed

Affiliation: Beijing Hospital of Traditional Chinese Medicine, Capital Medical University, Beijing 100010, China; Beijing Institute of Traditional Chinese Medicine, Beijing 100010, China; Beijing Key Laboratory of Basic Research with Traditional Chinese Medicine on Infectious Diseases, Beijing 100010, China.

ABSTRACT
Hydroxysafflor yellow A (HSYA) is an effective therapeutic agent for inflammatory diseases and autoimmune disorders; however, its regulatory effect on NLRP3 inflammasome activation in macrophages has not been investigated. In this study, we predicted the potential interaction between HSYA and xanthine oxidase (XO) via PharmMapper inverse docking and confirmed the binding inhibition via inhibitory test (IC50 = 40.04 μM). Computation docking illustrated that, in this HSYA-XO complex, HSYA was surrounded by Leu 648, Leu 712, His 875, Leu 873, Ser 876, Glu 879, Phe 649, and Asn 650 with a binding energy of -5.77 kcal/M and formed hydrogen bonds with the hydroxyl groups of HSYA at Glu 879, Asn 650, and His 875. We then found that HSYA significantly decreased the activity of XO in RAW264.7 macrophages and suppressed LPS-induced ROS generation. Moreover, we proved that HSYA markedly inhibited LPS-induced cleaved caspase-1 activation via suppressing the sensitization of NLRP3 inflammasome and prevented the mature IL-1β formation from pro-IL-1β form. These findings suggest that XO may be a potential target of HSYA via direct binding inhibition and the combination of HSYA-XO suppresses LPS-induced ROS generation, contributing to the depression of NLRP3 inflammasome and inhibition of IL-1β secretion in macrophages.

No MeSH data available.


Related in: MedlinePlus