PCSK9 Inhibitors Reduce PCSK9 and Early Atherogenic Biomarkers in Stimulated Human Coronary Artery Endothelial Cells

Abstract

Despite reports on the efficacy of proprotein convertase subtilisin-Kexin type 9 (PCSK9) inhibitors as a potent lipid-lowering agent in various large-scale clinical trials, the anti-atherogenic properties of PCSK9 inhibitors in reducing PCSK9 and atherogenesis biomarkers via the NF-ĸB and eNOS pathway has yet to be established. This study aimed to investigate the effects of PCSK9 inhibitors on PCSK9, targeted early atherogenesis biomarkers, and monocyte binding in stimulated human coronary artery endothelial cells (HCAEC). HCAEC were stimulated with lipopolysaccharides (LPS) and incubated with evolocumab and alirocumab. The protein and gene expression of PCSK9, interleukin-6 (IL-6), E-selectin, intercellular adhesion molecule 1 (ICAM-1), nuclear factor kappa B (NF-ĸB) p65, and endothelial nitric oxide synthase (eNOS) were measured using ELISA and QuantiGene plex, respectively. The binding of U937 monocytes to endothelial cell capacity was measured by the Rose Bengal method. The anti-atherogenic effects of evolocumab and alirocumab were contributed to by the downregulation of PCSK9, early atherogenesis biomarkers, and the significant inhibition of monocyte adhesion to the endothelial cells via the NF-ĸB and eNOS pathways. These suggest the beyond cholesterol-lowering beneficial effects of PCSK9 inhibitors in impeding atherogenesis during the initial phase of atherosclerotic plaque development, hence their potential role in preventing atherosclerosis-related complications.

PCSK9 inhibitors ameliorate arterial stiffness in ACS patients: evidences from mendelian randomization, cohort studies and basic experiments

“…According to published methods, 10 mmol/L β-gp, 20ug/m dexamethasone and 50ug/mL Lascorbic acid were added to the routine medium to induce osteogenesis of VSMCs for 7 days, the medium were changed every 2/3 days [23][24][25][26][27][28] . To mimic the effects of PCSK9i in vivo, 100 µg/mL of reagent-grade puri ed evolocumab (Selleck, America) was used to co-incubate with VSMCs for 7 days during osteogenesis induction [29][30][31] . To mimic the stimulation of VSMCs by PCSK9 protein in vivo, refer to previous research [32] , 0 µg/ml, 0.55 µg/ml, 1.1 µg/ml, 2.2 µg/ml, and 4.4 µg/ml of recombinant human PCSK9 (MedChemExpress, USA) were co-incubated with VSMCs for 7 days, and the concentration exhibiting the highest intracellular calcium content was chosen for subsequent experimental procedures.…”

PCSK9 inhibitors ameliorate arterial stiffness in ACS patients: evidences from Mendelian randomization, a retrospective study and basic experiments

“…According to published methods, 10 mmol/L β-gp, 20ug/m dexamethasone and 50ug/ mL L-ascorbic acid were added to the routine medium to induce osteogenesis of VSMCs for 7 days, the medium were changed every 2/3 days (24-29). To mimic the effects of PCSK9i in vivo, 100 μg/mL of reagent-grade purified evolocumab (Selleck, America) was used to co-incubate with VSMCs for 7 days during osteogenesis induction (30)(31)(32). To mimic the stimulation of VSMCs by PCSK9 protein in vivo, refer to previous research (33), 0 μg/mL, 0.55 μg/mL, 1.1 μg/mL, 2.2 μg/mL, and 4.4 μg/mL of recombinant human PCSK9 (MedChemExpress, USA) were co-incubated with VSMCs for 7 days, and the concentration exhibiting the highest intracellular calcium content was chosen for subsequent experimental procedures.…”

The Anti-Thrombotic Effects of PCSK9 Inhibitors

“…The in vitro study by authors Zulkapli et al showed that the co-incubation of alirocumab and evolocumab at specific concentrations resulted in an upregulation of the production of the eNOS protein. Only evolocumab caused an increase, although not significant, in eNOS mRNA [ 75 ]. Treatment with a PCSK9 inhibitor given to type 2 diabetes (T2D) patients (110 individuals) along with SGLT2i treatment improved FMD and the bioavailability of NO.…”