Lycium barbarum polysaccharides inhibit ischemia/reperfusion-induced myocardial injury via the Nrf2 antioxidant pathway

Figures

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  Fig. 1. Pretreatment with LBP reduced myocardial I/R injury. (A) Representative images of the infarct size in all groups. (B) Quantitative analysis of the myocardial infarct size. (C, D) Quantitative analysis of serum creatine kinase (CK) and lactate dehydrogenase (LDH) activity. (E-H) Hemodynamic parameters (HR, heart rate; MAP, mean arterial pressure; LVSP, left ventricular systolic pressure; LVEDP, left ventricular end-diastolic pressure; +dp/dt and −dp/dt, the maximum slope of systolic and diastolic pressure decrements) in all rats. Data are Means ± SD (n = 10). *P < 0.05, **P < 0.01 versus sham group, #P < 0.05 versus I/R group.

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  Fig. 2. LBP inhibited myocardial oxidative stress damage in I/R group. (A) Representative images using dihydroethidium (DHE) staining in myocardial tissue; scale bar =25 μm. (B) Relative fluorescence intensity of reactive oxygen species (ROS). (C-F) Malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GPX) and catalase (CAT) levels in serum from all rats. (G) Representative images by propidium iodide/Annexin V staining; White triangle represents TUNEL-positive cells. (H) Quantification of the percentage of apoptosis in myocardial tissue; scale bar =50 μm. (I, J) Tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) levels in myocardial tissue in all rats. Data are Means ± SD (n = 10). *P < 0.05, **P < 0.01 versus sham group, #P < 0.05 versus I/R group.

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  Fig. 3. LBP activated Nrf2 antioxidant signal pathway. (A)Western blots of nuclear Nrf2, histone3 (H3), cytosolic Nrf2, heme oxygenase 1(HO1), NADPH dehydrogenase quinone 1 (NQO1) and GAPDH proteins. (B-E) Western blot analysis of nuclear Nrf2, cytosolic Nrf2, HO1 and NQO1. Data are Means ± SD (n = 10). *P < 0.05 versus sham group, #P < 0.05 versus I/R group.

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  Fig. 4. LBP administration decreased hypoxia/reoxygenation (H/R) induced injury in H9c2 cells. (A-C) Viability of H9c2 cells was measured by MTT in H/R pretreated with or without LBP. (D, E) Quantitative analysis of creatine kinase (CK) and lactate dehydrogenase (LDH) activies. Data are Means ± SD (n = 6). *P < 0.05, **P < 0.01 versus Con group, #P < 0.05 versus H/R group.

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  Fig. 5. LBP suppressed oxidative stress induced by hypoxia/reoxygenation (H/R) in H9c2 cells. (A) Representative images using 2, 7-dichlorodihydrofluorescein diacetate (DCHF-DA) staining; scale bar =25 μm. (B) Relative fluorescence intensity of reactive oxygen species (ROS). (C-F) Malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GPX) and catalase (CAT) levels in supernatant from all groups. (G) Representative images by terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) staining; White triangles represent TUNEL-positive cells. (H) Quantification of the percentage of apoptotic H9c2 cells; scale bar =50 μm. (I, J) Tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) levels in cell supernatant. Data are Means ± SD (n = 6). *P < 0.05, **P < 0.01 versus Con group, #P < 0.05 versus H/R group.

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  Fig. 6. LBP upregulated the expression of anti-oxidative molecular in Nrf2 signal pathway H/R in H9c2 cells. (A)Western blots of nuclear Nrf2, histone3 (H3), cytosolic Nrf2, heme oxygenase 1(HO1), NADPH dehydrogenase quinone 1 (NQO1) and GAPDH proteins. (B-E) Western blot analysis of nuclear Nrf2, cytosolic Nrf2, HO1 and NQO1. (F) Representative images of Nrf2 translocation by propidium iodide/Annexin V staining in all groups. (G) Relative fluorescence intensity of nuclear Nrf2. Data are Means ± SD (n = 6). *P < 0.05 versus Con group, #P < 0.05 versus H/R group.

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  Fig. 7. The protective effects of LBP were abolished by Nrf2 knockdown. (A) Western blots of Nrf2, heme oxygenase 1(HO1), NADPH dehydrogenase quinone 1 (NQO1) and GAPDH proteins. (B-D) Western blot analysis of Nrf2, HO1 and NQO1. (E, F) The production of reactive oxygen species (ROS) in H9c2 cells; scale bar =25 μm. (G-J) Quantification of malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GPX) and catalase (CAT) levels in Nrf2 knockdown cells. (K, L) Apoptotic index in Ctr siRNA and Nrf2 siRNA group; White triangle represents TUNEL-positive cells; scale bar =50 μm. (M, N) Tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) levels in cell supernatant. Data are Means ± SD (n = 6). **P < 0.01, ***P < 0.001, versus Ctr siRNA group.