Ellagic acid prevents monocrotaline-induced pulmonary artery hypertension via inhibiting NLRP3 inflammasome activation in rats Bing Tang, Guang-xian.

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Ellagic acid prevents monocrotaline-induced pulmonary artery hypertension via inhibiting NLRP3 inflammasome activation in rats Bing Tang, Guang-xian Chen, Meng-ya Liang, Jian-ping Yao, Zhong-kai Wu International Journal of Cardiology Volume 180, Pages 134-141 (February 2015) DOI: 10.1016/j.ijcard.2014.11.161 Copyright © 2014 Terms and Conditions

Fig. 1 EA alleviates hemodynamic changes and right ventricular hypertrophy in MCT-induced PAH 4weeks after MCT exposure. EA decreased RVSP in MCT-treated rats. Further a dose-dependent manner presented, the RVSP of EA-high group was lower than EA-MCT group (p<0.05). EA also reduced the RV/(LV+S) ratio in MCT-treated rats. Representative hemodynamic data (RVSP) from BIOPAC MP100 data acquisition system were showed in Fig. 1C. Data represent means±SD. *p<0.05 versus Control group; ▲p<0.05 versus MCT group; #p<0.05 versus Control group. n=8 per group. MCT, monocrotaline; PAH, pulmonary artery hypertension; EA, ellagic acid; RVSP, right ventricle systolic pressure; RV/LV+S ratio, the right ventricular weight to left ventricular plus septal weight ratio. International Journal of Cardiology 2015 180, 134-141DOI: (10.1016/j.ijcard.2014.11.161) Copyright © 2014 Terms and Conditions

Fig. 2 EA prevents pulmonary vascular remodeling in MCT-treated rats. A through D, Representative hematoxylin and eosin (HE) staining images compare the arteriole wall thickness in rats. Original magnification ×400, scale bars=50μm. The pulmonary arteriole walls in the Control group were thin and single layered. The intima, media and extima were difficult to distinguish. Conspicuous pulmonary vascular remodeling could be observed in MCT group, characterized by pulmonary arteriole media muscularization and intima and extima thickness. After EA treatment, medial hypertrophy decreased. E, Quantification analysis of percent medial thickness for arteriole (25–100μm in external diameter) showed that EA reduced the medial wall thickness in MCT-treated rats compared with Controls. Data represent means±SD. *p<0.05 versus Control group; ▲p<0.05 versus MCT group; #p<0.05 versus Control group. n=8 per group. EA, ellagic acid; MCT, monocrotaline. International Journal of Cardiology 2015 180, 134-141DOI: (10.1016/j.ijcard.2014.11.161) Copyright © 2014 Terms and Conditions

Fig. 3 EA decreases serum BNP levels in MCT-induced PAH rats. Enzyme-linked immunosorbent assay showed that MCT-treated rats had higher BNP levels compared with Controls, whereas EA supplementation reduced the MCT-induced increase in BNP levels. Data represent means±SD. n.s., not significant; *p<0.05 versus Control group; ▲p<0.05 versus MCT group. n=8 per group. EA, ellagic acid; MCT, monocrotaline; PAH, pulmonary artery hypertension; BNP, B-type natriuretic peptide. International Journal of Cardiology 2015 180, 134-141DOI: (10.1016/j.ijcard.2014.11.161) Copyright © 2014 Terms and Conditions

Fig. 4 EA suppresses OS in the lungs of MCT-treated rats. Data represent means±SD. SOD activity in the MCT group was lower than that in the Control group, but in the EA group, it was higher than in the MCT group. The MDA level in the lungs of the MCT group was higher than that in the Control group, and in the EA group, it was lower than in the MCT group. n.s., not significant; *p<0.05 versus Control group; ▲p<0.05 versus MCT group. n=8 rats per group. EA, ellagic acid; MCT, monocrotaline; SOD, superoxide dismutase; MDA, malondialdehyde. International Journal of Cardiology 2015 180, 134-141DOI: (10.1016/j.ijcard.2014.11.161) Copyright © 2014 Terms and Conditions

Fig. 5 EA reduces inflammatory cytokine levels of serum in MCT-induced PAH rats. Selected cytokines with divergent expression in the MCT group compared with the EA-treated group. Expression levels in normal lungs are shown for comparison. Cytokines induced during the progression of MCT-induced PAH that were suppressed by EA treatment include the inflammatory cytokines IL-1β, IL-2, IL-4, IL-6, IL-10, IFN-γ and MIP-1α. Analysis was performed by the Bio-Plex Pro™ rat cytokine immunoassay. Data represent means±SD. n.s., not significant; *p<0.05 versus Control group; ▲p<0.05 versus MCT group; #p<0.05 versus Control group. n=8 per group. EA, ellagic acid; MCT, monocrotaline; PAH, pulmonary artery hypertension; IL-1β, interleukin-1beta; TNF-α, tumor necrosis factor alpha; IFN-γ, interferon gamma; MIP-1α, macrophage inflammatory protein 1 alpha. International Journal of Cardiology 2015 180, 134-141DOI: (10.1016/j.ijcard.2014.11.161) Copyright © 2014 Terms and Conditions

Fig. 6 EA inhibits lung NALP3 inflammasome activation in MCT-treated rats. Representative Western blot results and graph showed lung NALP3, caspase-1 and IL-1β expression 4weeks after MCT injection in different groups of rats. Relative protein levels of NALP3, caspase-1(p20) and IL-1β(p17) were determined after normalization with β-actin. Data are the means±SD. The NLRP3 inflammasome pathway was activated in MCT-induced PAH rats, further EA treatment can down-regulate this signal pathway. *p<0.05 versus Control group; ▲p<0.05 versus MCT group; #p<0.05 versus Control group. n=6 per group. EA, ellagic acid; MCT, monocrotaline; IL-1β, interleukin-1beta. International Journal of Cardiology 2015 180, 134-141DOI: (10.1016/j.ijcard.2014.11.161) Copyright © 2014 Terms and Conditions