The protective role and mechanism of liriodendrin in rats with myocardial infarction

Figures

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  Figure 1. The value of troponin T was the highest in the control group, compared to the other two groups. The value of troponin T decreased significantly in the liriodendrin group. *: control group vs. sham group, P<0.05; #: control group vs. liriodendrin group, P<0.05; **: liriodendrin group vs. sham group, P<0.05.

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  Figure 2. Parameters of cardiac function obtained from echocardiographic images (mean ± SEM). As shown in the figure, EF of the control group decreased significatively and decreased the least in the liriodendrin group. Compared to the control group, EDV, ESV, LVIDd, and LVIDs of the liriodendrin group expanded less significantly. (A) Sham group; (B) control group; and (C) liriodendrin group. *: control group vs. sham group; P<0.05. #: control group vs. liriodendrin group, P<0.05; **: liriodendrin group vs. sham group, P<0.05. LVIDd, left ventricular end-diastolic diameter; LVIDs, left ventricular end-systolic diameter; ESV, end-systolic volume; EDV, end-diastolic volume; EF, ejection fraction; SEM, standard error of the mean.

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  Figure 3. Hearts were stained with HE to observe tissue morphology. (A) The sham group. Liriodendrin alleviated MI damage in rats: (B) collagen deposition was greatest in the control group, myocardial tissue structure disappeared, and many inflammatory cells infiltrated in the damaged region; (C) the least damage was observed in the liriodendrin group. HE, hematoxylin and eosin; MI, myocardial infarction.

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  Figure 4. Effect of liriodendrin on anti-apoptosis in myocardium stained with TUNEL. Apoptotic cells are characterized by dark brown nuclei. (A) Sham group; (B) control group; (C) liriodendrin group; and (D) semiquantitative analysis of myocardial tissues by apoptosis index showed that apoptosis was suppressed by liriodendrin and that apoptosis was the most significant in the control group. *: control group vs. sham group, P<0.05; #: control group vs. liriodendrin group, P<0.05. TUNEL, transferase-mediated nick end labeling.

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  Figure 5. Treatment impact of liriodendrin on concentrations of TNF-α and IL-1β in myocardial tissue. Rat hearts were collected after euthanization. Left ventricular myocardial tissue was homogenized and sonicated. After the centrifuge, the supernatant was obtained to measure IL-1β (A) and TNF-α (B). Liriodendrin treatment remarkedly inhibited the release of inflammatory factors compared to the control group. *: control group vs. sham group, P<0.05; #: control group vs. liriodendrin group, P<0.05. IL-1β, interleukin-1β; TNF-α, tumor necrosis factor-α.

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  Figure 6. The anti-apoptosis ability of cardiomyocytes in rats with MI was enhanced after liriodendrin treatment. GAPDH was used as a loading control. (A,B) On the contrary, Bax and caspase-3 proteins were reduced in the liriodendrin group compared to the control group; (C) Bcl-2 protein was increased in the liriodendrin group. All results above suggested that apoptosis was inhibited after liriodendrin was administered. *: control group vs. sham group, P<0.05; #: control group vs. liriodendrin group, P<0.05. MI, myocardial infarction; Bcl-2, B-cell lymphoma-2; GAPDH, glyceraldehyde-3-phosphate dehydrogenase.

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  Figure 7. After Liriodendrin treatment, the transcriptional activity of apoptosis was down-regulated in rats with MI. Liriodendrin reduced the mRNA levels of Bax (A), Bcl-2 (B), caspase-3 (C). Real-time PCR assay was performed in duplicate, and GAPDH mRNA was used to normalize the relative amount of mRNA. *: control group vs. sham group, P<0.05; #: control group vs. liriodendrin group, P<0.05. MI, myocardial infarction; Bcl-2, B-cell lymphoma-2; GAPDH, glyceraldehyde-3-phosphate dehydrogenase.

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  Figure 8. Effect of liriodendrin on activation of NF-κB. (A) Proteins obtained from the myocardium were analyzed by western blot using an enhanced chemiluminescence kit. (B,D) Each band was normalized to that of GAPDH. Liriodendrin inhibited NF-κB and IκBα phosphorylation in the myocardium of rats. (C,E) Quantitated density of NF-κB and IκBα in the myocardium. *: control group vs. sham group, P<0.05; #: control group vs. liriodendrin group, P<0.05; **: liriodendrin group vs. sham group, P<0.05. NF-κB, nuclear factor kappa-B; GAPDH, glyceraldehyde-3-phosphate dehydrogenase.

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  Figure 9. After Liriodendrin treatment, the transcriptional activity of the NF-κB pathway was down-regulated in rats with MI. After RT-PCR assay was performed, we found that liriodendrin reduced the mRNA levels of NF-κB (A), IκBα (B), and IKK-β (C). *: control group vs. sham group, P<0.05; #: control group vs. liriodendrin group, P<0.05. NF-κB, nuclear factor kappa-B; MI, myocardial infarction; RT-PCR, real-time polymerase chain reaction.

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  Figure 10. Effects of liriodendrin treatment on upregulated expression of cytokines, TNF-α, IL-1β, and IL-6 in ischemia-hypoxia stimulated H9C2 cells. (A) Different concentrations of liriodendrin were given 24 h before ischemia-hypoxia exposure. Cell culture media were harvested 24 h after the process of ischemia-hypoxia, and the concentrations of TNF-α, IL-1β, and IL-6 were measured by ELISA assay. (B) Liriodendrin downregulated the NF-κB pathway and apoptosis in H9C2 cells, and the therapy effect was proportional to the concentration of liriodendrin. *P<0.05, compared with the DMSO group. IL, interleukin; DMSO, dimethyl sulfoxide; ELISA, enzyme-linked immunosorbent assay.