HIIT and MICT mitigate endothelial dysfunction in early atherosclerotic mice via PCSK9 inhibition

Abstract

Atherosclerosis (AS), driven by vascular endothelial dysfunction and poses a global health threat. This study compared the therapeutic effects of high-intensity interval training (HIIT) and moderate-intensity continuous training (MICT) on vascular endothelial function in early-stage AS mice, specifically investigating PCSK9 modulation and the TRX/TXNIP/NLRP3/GSDMD-N pathway. ApoE^-/- mice (n = 6/group) fed a high-fat diet for 12 weeks were randomized into sedentary (AS-S), HIIT (AS-HIIT), and MICT (AS-MICT) groups, with wild-type mice as control. Training lasted 12 weeks. Outcomes included body weight, lipid profiles (TG, TC, LDL-C, HDL-C), oxidative stress markers (T-SOD, GSH-Px, MDA), vascular function (eNOS expression, ACh-induced vasorelaxation), and TRX/TXNIP/NLRP3/GSDMD-N pathway activity. Both HIIT and MICT reduced body weight (p < 0.05) and improved lipid profile. Exercise groups showed reduced oxidative stress and inflammation pathways (p < 0.05). HIIT and MICT ameliorate early AS by reducing PCSK9 and oxidative/inflammatory pathway levels (p < 0.05), but HIIT demonstrates superior efficacy in improving endothelial function and pathway activation. These findings show HIIT and MICT mitigate endothelial dysfunction in early atherosclerotic mice via PCSK9 inhibition and advocate for HIIT as a prioritized strategy in early AS management.

Keywords: Atherosclerosis; Endothelial dysfunction; Exercise; Pyroptosis.

Fig. 1

Effects of MCIT and HIIT on basic data and vascular function in atherosclerotic mice. The groups included wild-type mice (Control), ApoE KO HFD sedentary mice (AS-S), ApoE KO HFD mice undergoing moderate-intensity continuous training (AS-MICT), and ApoE KO HFD mice undergoing high-intensity interval training (AS-HIIT). Mice were trained on a treadmill following specific exercise protocols for 12 weeks. The following parameters were measured. Changes of (A) body weight were recorded. (B) T-SOD activity, (C) MDA, (D) GSH-Px, (E) TG, (F) TC, (G) HDL-C, and (H) LDL-C in the serum of mice were tested by kits. (I) Representative hematoxylin and eosin (HE)-stained images of aortas from each group. (J) and (K) The protein expression of eNOS in aortas was determined by western blotting. (L) ACh-induced relaxation and (M) SNP-induced relaxation of aortas were measured by vascular reactivity experiment. *P < 0.05, AS-S compared with Control; #P < 0.05, AS-MICT and AS-HIIT compared with AS-S. Data are presented as means ± SD.

Fig. 2

MICT and HIIT improve vascular function through inhibiting PCSK9 level. (A and B) PCSK9 expression in the liver, kidney, skeletal muscle, heart, and aorta was measured using western blotting. (C) PCSK9 levels in the serum were quantified via ELISA. *P < 0.05, AS-S compared with Control; #P < 0.05, AS-MICT and AS-HIIT compared with AS-S. Data are presented as means ± SD. (D and E) Protein expression of eNOS in aortic tissue, either Vehicle- or PCSK9-treated in vitro, was analyzed. (F) ACh-induced and (G) SNP-induced vasodilation function of in vitro-cultured aortas treated with or without PCSK9. *P < 0.05, Vehicle group compared with PCSK9 group. Data are presented as means ± SD.

Fig. 3

MICT and HIIT inhibit the TRX/TXNIP/NLRP3/GSDMD-N signaling pathway in the aortas of atherosclerotic mice. (A and B) The protein expressions of TRX/TXNIP and NLRP3/GSDMD-N were detected by western blot. (C and D) Immunohistochemical staining was performed to detect NLRP3 and GSDMD-N in the aortas. *P < 0.05, AS-S compared with Control; #P < 0.05, AS-MICT and AS-HIIT compared with AS-S. Data are presented as means ± SD.

Fig. 4

PCSK9 impairs endothelial function through activating the TRX/TXNIP/NLRP3/GSDMD-N signaling pathway. (A and B) Protein expressions of TRX, TXNIP, NLRP3, and GSDMD-N were analyzed by western blotting in Vehicle- or PCSK9-treated aortas in vitro. MCC950 (a specific NLRP3 inhibitor), TXNIP-IN-1 (a specific TXNIP inhibitor), and Ruscogenin (another TXNIP inhibitor) were applied, with a final intervention concentration of 10 μg/ml for all inhibitors. (C and D) The protein expressions of TRX, TXNIP, NLRP3 and GSDMD-N were detected by western blot. (E) ACh-induced and (F) SNP-induced vasodilation in vitro of aortas treated with or without PCSK9. *P < 0.05, Vehicle group compared with PCSK9 group. Data are presented as means ± SD.

Fig. 5

MICT and HIIT can alleviate endothelial-dependent vasodilation dysfunction in mice with atherosclerosis (AS) by suppressing the levels of PCSK9 in serum and blood vessels. These protective effects may be related to the blockade of the PCSK9-induced TRX/TXNIP/NLRP3/GSDMD-N pathway activation. Both types of training can effectively prevent the progression of endothelial dysfunction in the early stages of AS.