Aerobic exercise regulates FGF21 and NLRP3 inflammasome-mediated pyroptosis and inhibits atherosclerosis in mice

Abstract

Fibroblast growth factor 21 (FGF21), a known risk factor for atherosclerosis, is readily regulated by exercise, and it can inhibit NOD-like receptor protein 3 (NLRP3)-mediated pyroptosis. However, it is not clear whether aerobic exercise inhibits atherosclerosis via these pathways. Eight-week-old apolipoprotein E-deficient (ApoE-/-) mice on a high-fat diet were randomly divided into 1-h post-exercise (EX-1h), 24-h post-exercise (EX-24h), and sedentary (SED) groups. C57BL/6J wild-type mice fed normal chow served as controls (WT group). Mice in the EX-1h and EX-24h groups were subjected to treadmill exercise training for 12 weeks. Aerobic exercise reduced body weight; blood glucose, lipid, and inflammation levels; and aortic plaque area proportion. Aerobic exercise increased the sensitivity of FGF21 by upregulating the expression of the downstream receptor adiponectin (ApN); the serum FGF21 level after exercise increased initially, and then decreased. Aerobic exercise downregulated the expression of NLRP3 inflammasome-mediated pyroptosis-related markers in the aorta, and FGF21 may participate in the above process. Meanwhile, the liver may be the tissue source of serum FGF21 during aerobic exercise. In conclusion, aerobic exercise may inhibit atherogenesis by regulating FGF21 and NLRP3 inflammasome-mediated pyroptosis. Our study provides new information on the atherosclerosis-preventing mechanism of aerobic exercise.

Fig 1. Aerobic exercise reduces body weight without changing dietary intake.

(A) Body weight at the end of the experiment. (B) Average daily dietary intake. Data are presented as mean ± SD (n = 20). **p < 0.01 vs. WT group; ^## p < 0.01 vs. SED group.

Fig 2. Aerobic exercise inhibits atherosclerosis.

(A) Representative images of the whole aortic with oil red O staining. Scale bar = 10 mm; (B) Comparison of the proportion of aortic plaque area. Data are presented as mean ± SD (n = 5). **p <0.01 vs. WT group; ^## p <0.01 vs. SED group. (C) Representative images of hematoxylin and eosin (H&E) staining of cross-section of aortic root. Scale bar = 50 μm.

Fig 3. The effect of aerobic exercise on serum FGF21, FFAs, ApN levels.

(A) Aerobic exercise affects serum FGF21 levels. (B) Aerobic exercise affects serum FFAs levels. (C) Aerobic exercise affects serum ApN levels (n = 8). Data are expressed as mean ± SD. * p < 0.05, ** p < 0.01 vs. WT group; ^## p < 0.01 vs. SED group; ^&& p < 0.01 vs. EX-1h group.

Fig 4. The effect of aerobic exercise on the expression of liver and aorta FGF21 mRNA.

(A) Aerobic exercise affects FGF21 mRNA expression in liver; (B) Aerobic exercise did not alter the expression of FGF21 mRNA in the aorta (n = 7). Data are expressed as mean ± SD of three experiments. * p < 0.05, ** p < 0.01 vs. WT group; ^## p < 0.01 vs. SED group; ^&& p < 0.01 vs. EX-1h group.

Fig 5. Aerobic exercise activates liver FGF21 protein expression.

(A) Immunofluorescence detects liver FGF21 protein expression level. Scale bar = 100 μm. (B) Mean fluorescence intensity analysis of FGF21 in liver. The data are expressed as mean ± SD (n = 5, three slices taken from each mouse for staining), ** p < 0.01 vs. WT group; ^## p < 0.01 vs. SED group; ^&& p < 0.01 vs. EX-1h group.

Fig 6. Aerobic exercise does not change the expression of FGF21 protein in the aorta.

(A) The expression level of FGF21 protein in the aorta was detected by immunofluorescence. Scale bar = 100 μm. (B) Analysis of the average fluorescence intensity of FGF21 in the aorta (n = 5, three slices taken from each mouse for staining). (C) The protein levels of FGF21 in the aorta were detected by western blotting (n = 7). Data are presented as mean ± SD of three experiments.

Fig 7. Aerobic exercise inhibits NLRP3 inflammasome-mediated pyroptosis-related factor mRNA expression in the aorta.

(A-E) The mRNA expression of NLRP3, caspase-1, GSDMD, IL-1β, and IL-18 in the aorta were detected by real-time polymerase chain reaction (n = 7). Data are presented as mean ± SD of three experiments, **p < 0.01 vs. WT group; ^##p < 0.01 vs. SED group.

Fig 8. Aerobic exercise inhibits NLRP3 inflammasome-mediated pyroptosis-related factor protein expression in the aorta.

(A) The protein expression of NLRP3, caspase-1, GSDMD, IL-1β, and IL-18 in the aorta were detected by western blotting. (B-F) Comparison of protein expression in each group (n = 7). Data are presented as mean ± SD of three experiments, ** p < 0.01 vs. WT group; ^## p < 0.01 vs. SED group.

Fig 9. Aerobic exercise inhibits the expression of NLRP3 protein in the aorta.

(A) The expression of NLRP3 protein in the aorta was detected by the immunofluorescence method. Scale bar = 100 μm. (B) Analysis of the average fluorescence intensity of NLRP3 in the aorta (n = 5, three slices taken from each mouse for staining). The data are expressed as mean ± SD. ** p < 0.01 vs. WT group; ^## p < 0.01 vs. SED group.