Age-Related Shift in Cardiac and Metabolic Phenotyping Linked to Inflammatory Cytokines and Antioxidant Status in Mice

Abstract

Age-related alterations in cardiac function, metabolic, inflammatory and antioxidant profiles are associated with an increased risk of cardiovascular mortality and morbidity. Here, we examined cardiac and metabolic phenotypes in relation to inflammatory status and antioxidant capacity in young, middle-aged and old mice. Real-time reverse transcription-polymerase chain reactions were performed on myocardium and immunoassays on plasma. Left ventricular (LV) structure and function were assessed by echocardiography using high-frequency ultrasound. Middle-aged mice exhibited an altered metabolic profile and antioxidant capacity compared to young mice, whereas myocardial expression of inflammatory factors (TNFα, IL1β, IL6 and IL10) remained unchanged. In contrast, old mice exhibited increased expression of inflammatory cytokines and plasma levels of resistin compared to young and middle-aged mice (p < 0.05). The pro-inflammatory signature of aged hearts was associated with alterations in glutathione redox homeostasis and elevated contents of 4-hydroxynonenal (4-HNE), a marker of lipid peroxidation and oxidative stress. Furthermore, echocardiographic parameters of LV systolic and diastolic functions were significantly altered in old mice compared to young mice. Taken together, these findings suggest age-related shifts in cardiac phenotype encompass the spectrum of metabo-inflammatory abnormalities and altered redox homeostasis.

Keywords: aging; antioxidant; cardiac function; inflammation and resistin; metabolic status.

Figure 1

Age-dependent profile of resistin expression in mice. Plasma level (a) and quantitative RT-PCR analysis of mRNA expression (b) of resistin in young, middle-aged and old mice. * p < 0.05 and *** p < 0.001 between the indicated groups, ns, not significant, one-way ANOVA.

Figure 2

Parasternal short-axis M-mode images and age-associated comparisons of the left ventricular dimensions. Representative M-mode images of the parasternal short-axis view with measurements of the left ventricular dimensions in young (a), middle-aged (b) and old mice (c). Measurements of the left ventricular internal diameter in diastole (LVIDd) (d), left ventricular internal diameter in systole (LVIDs) (e), and left ventricular mass (f) in young, middle-aged and old mice. The echocardiographic examinations were performed and analyzed using the Vevo 2100 Imaging System (VisualSonics, Toronto, ON, Canada). * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 between the indicated groups, ns, not significant, one-way ANOVA.

Figure 3

Parasternal long-axis images and LV systolic function in young, middle-aged and old mice. Representative images from parasternal long-axis view with the left ventricular tracing in systole (green) in young (a), middle-aged (b), and old mice (c). Measurements of the left ventricular ejection fraction (EF) (d) and left ventricular fractional shortening (FS) (e) in young, middle-aged and old mice. The echocardiographic examinations were performed and analyzed using the Vevo 2100 Imaging System (VisualSonics, Toronto, ON, Canada). **** p < 0.0001 between the indicated groups, ns, not significant, one-way ANOVA.

Figure 4

Pulse Doppler waveform of mitral inflow velocity and LV diastolic function in young, middle-aged and old mice. Representative images of pulse Doppler waveforms of mitral inflow velocity (white wave) and tissue Doppler imaging (yellow wave) in young (a), middle-aged (b), and old mice (c). Measurements of mitral valve inflow E wave (MV E) (d) and tissue Doppler-derived septal mitral annular E’ wave (E’) (e) in young, middle-aged and old mice. The echocardiographic examinations were performed and analyzed using the Vevo 2100 Imaging System (VisualSonics, Toronto, ON, Canada). * p < 0.05, *** p < 0.001, and **** p < 0.0001 between the indicated groups, ns, not significant, one-way ANOVA.

Figure 5

Age-dependent antioxidant capacity in mice. Total SOD (a) and catalase (b) activities in young, middle-aged and old mice. Plasma levels of SOD and catalase were measured using ELISA kits. * p < 0.05 and ** p < 0.01 between the indicated groups, ns, not significant, one-way ANOVA.

Figure 6

Profile of oxidative stress markers in young, middle-aged and old mice. Quantification analysis (a) and Western blot image (b) of 4-HNE in cardiac tissue from young, middle-aged and old mice. Ratio of reduced glutathione to oxidized glutathione (GSH/GSSG) in plasma from young, middle-aged and old mice (c). * p < 0.05 between the indicated groups, ns, not significant, one-way ANOVA.

Figure 7

Age-dependent mRNA expression of inflammatory factors in mice. Quantitative RT-PCR analysis of mRNA expression levels of TNFα (a), IL1β (b), IL6 (c), and IL10 (d). * p < 0.05, ** p < 0.01, and *** p < 0.001 between the indicated groups, ns, not significant, one-way ANOVA.

Figure 8

NF-κB-p65 Ser276 phosphorylation in young, middle-aged and old mice. Heart total protein extracts were resolved by SDS-PAGE and probed with antibody against p-NF-κB-p65-Ser276. Western blot image (a) and quantification analysis from a (b). * p < 0.05 between the indicated groups, ns, not significant, one-way ANOVA.