Fat-1 Transgenic Mice Rich in Endogenous Omega-3 Fatty Acids are Protected from Lipopolysaccharide-Induced Cardiac Dysfunction

Figures

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  Figure 1. The elevation of the Fat‐1 transgene expression enhances mice survival after endotoxic shock induced by exogenous LPS. Log‐rank test was used to determine the median survival time. *P < 0.05 vs. WT control treated with LPS.

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  Figure 2. Fat‐1 transgene improved cardiac function in LPS‐treated mice. (A) Representative photographs of M‐mode echocardiography and quantitative analysis of LV dimension and systolic/diastolic function in WT and fat‐1 mice exposed to LPS or vehicle. (B) Representative image of haematoxylin and eosin showing myocardial morphology in different groups. Original magnification 400× (scale bar = 40 μm). (C) Myocardial levels of BNP and cTNT in WT and fat‐1 mice exposed to LPS or vehicle. *P < 0.05 vs. WT control treated with vehicle, # P < 0.05 vs. WT treated with LPS.

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  Figure 3. Proteomic analysis reveals myocardial protein alterations in fat‐1 mice after LPS challenge. (A) With the use of a fold change >2 and an adjusted P value of <0.05, 91 proteins were found to be differentially expressed (32 up‐regulated and 59 down‐regulated) in the left ventricle from fat‐1 mice compared with control mice at 12 h after LPS challenge as shown in the volcano plots. (B) KEGG pathways analysis of the differentially expressed proteins. (C) Major biological processes altered in the left ventricle from fat‐1 mice were identified using GO analysis according to biological process ranked by enrichment score.

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  Figure 4. Fat‐1 transgene reduced LPS‐induced myocardial inflammatory cytokine production. (A) Myocardial TNF‐α, IL‐1β, and IL‐6 mRNA expression were assayed by quantitative real‐time PCR in the vehicle‐ and LPS‐treated WT and fat‐1 mice. (B) Myocardial TNF‐α, IL‐1β, and IL‐6 protein levels in the WT and fat‐1 mice treated with or without LPS were measured by ELISA. (C) Representative photomicrographs of immunohistochemical staining for TNF‐α, IL‐1β, and IL‐6 (brown) in cardiac sections from vehicle‐, and LPS‐treated WT and fat‐1 mice (scale bar = 40 μm). Quantitation of these cytokines immunoreactivity is shown lower panel. *P < 0.05 vs. WT control treated with vehicle, # P < 0.05 vs. WT treated with LPS.

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  Figure 5. Fat‐1 transgene attenuated oxidative stress in LPS‐treated mice. (A) Fat‐1 transgene reduces malondialdehyde (MDA) content and enhances anti‐oxidant enzyme system including superoxide dismutase (SOD) and glutathione (GSH) levels in hearts of LPS‐treated rats. (B) Myocardial COX2 mRNA levels in the vehicle‐ and LPS‐treated WT and fat‐1 mice were measured by quantitative real‐time PCR. (C) COX2 activity was analysed using COX activity assay kit. *P < 0.05 vs. WT control treated with vehicle, # P < 0.05 vs. WT treated with LPS.

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  Figure 6. Fat‐1 transgene enhances autophagic activity protein expression in LPS‐treated mice. (A) Autophagy‐associated protein LC3, p62, and Atg7 levels were analysed by immunoblotting at 72 h after LPS (10 mg/kg) intraperitoneal injected into WT and fat‐1 transgenic mice. (B) Representative confocal images showed the LC3 immunoreactivity was colocalized with myogoblin, a marker of cardiomyocytes. Red, LC3; blue, DAPI‐stained nuclei; Green α‐actinin‐positive cardiacmyocytes; original magnification ×600; scale bar = 20 μm. Quantitative analysis of LC3 dots in cardiacmyocytes is shown in right panel. (C) Representative electron micrographs in left‐ventricular myocardium from WT and fat‐1 mice (scale bar = 1 μm). Quantitative analysis of autophagosomes is shown in right panel. *P < 0.05 vs. WT control treated with vehicle, # P < 0.05 vs. WT treated with LPS.

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  Figure 7. Fat‐1 transgene regulates NFκB and Sirt1 in LPS‐treated mice. (A) Myocardial IκB and nuclear NFκB levels were analysed by immunoblotting at 72 h after LPS (10 mg/kg) intraperitoneal injected into WT and fat‐1 transgenic mice. Data were normalized using tubulin. (B) Immunoblotting and quantification myocardial Sirt1 and PPARγ levels in the WT and fat‐1 mice treated with or without LPS. Data were normalized using tubulin. Quantification of densitometry of these proteins was shown in right panel. *P < 0.05 vs. WT control treated with vehicle, # P < 0.05 vs. WT treated with LPS.