Interleukin-6 knockout reverses macrophage differentiation imbalance and alleviates cardiac dysfunction in aging mice

Figures

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  Figure 1. Effect of aging on cardiac IL-6 expression. (A) The expression of IL-6 in mice at 3 months, 15 months, 20 months, and 25 months of age was measured by Western blotting. N=5 in each group. * p<0.05 vs. the 3-month group. # p<0.05 vs. the previous group. (B) Plasma sIL-6R and sgp130 levels were detected in each group. N=5 in each group. * p<0.05 vs. the 3-month group. # p<0.05 vs. the previous group. (C) Effects of DOX treatment on IL-6 mRNA expression in WT Møs, lymphocytes, DCs, MCs, and CFs. N=5 in each group. * p<0.05 vs. the group. (D) The cardiac expression of both IL-6 and Mø markers in mice of different ages was detected in 25-month-old mice by immunofluorescence staining. N=5 in each group. (E) Double immunofluorescence staining for anti-F4/80 and anti-IL-6. N=5 in each group.

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  Figure 2. Effects of IL-6 KO on cardiac dysfunction. (A) Survival rates of WT mice and IL-6 KO mice. (B–F) LVPWT, LVEDD, LVESD, LVEF, and LVFS were determined at different time points by ultrasonic cardiography. (G–I) The HRs of young mice and aging mice were measured. N=10-16 in each group. * p<0.05 vs. the young WT group; # p<0.05 vs. the aging WT group.

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  Figure 3. Effects of IL-6 KO on cardiac remodeling and aging-related protein levels. (A–C) Body weights (BWs), heart weights (HWs), and HW/BW ratios in the four groups. (D) MC CSA and cardiac fibrosis area in each group. (E) Lipofuscin density in each group. (F) p16, p19, p21, p53, and Sirt1 mRNA expression levels were analyzed by RT-PCR. (G) Cardiac p53 protein expression was detected by immunofluorescence staining. N=5-8 in each group. * p<0.05 vs. the young WT group; # p<0.05 vs. the aging WT group.

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  Figure 4. Effect of IL-6 KO on cardiac Mø differentiation. (A) STAT1 and p65 phosphorylation was detected by Western blotting. (B) Cardiac iNOS and Arg-1 levels were measured by immunofluorescence staining. (C) mRNA expression of Mø1- and Mø2-related markers in each group. N=5-8 in each group. * p<0.05 vs. the young WT group; # p<0.05 vs. the aging WT group.

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  Figure 5. Effects on MC apoptosis. (A) The expression of apoptosis-related proteins was detected in hearts. (B) TUNEL staining was performed to mark apoptotic cells, and the number of TUNEL-positive cells in each group was determined. N=5-8 in each group. * p<0.05 vs. the young WT group; # p<0.05 vs. the aging WT group.

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  Figure 6. Effects of IL-6 KO on Mø differentiation and MC apoptosis in vitro. (A). The levels of p65 phosphorylation in DOX-treated WT Møs and IL-6 KO Møs were measured. (B, C). The mRNA expression of Mø1- and Mø2-related markers was measured in DOX-treated Møs. (D). The effects of DOX-induced Mø differentiation on MC apoptosis and MC aging were determined by RT-PCR. N=5 in each group. * p<0.05 vs. the young WT group; # p<0.05 vs. the aging WT group.