Impact and mechanism of proprotein convertase subtilisin/kexin type 9 on cholesterol efflux in human monocyte-derived foam cells

Liao Fujun; Bao Hailong; Gong Caiwei; Liu Danan

doi:10.12307/2026.623

Abstract：BACKGROUND:Human proprotein convertase subtilisin/kexin type 9(PCSK9),a key regulator of cholesterol metabolism,modulates cholesterol uptake and plasma lipid levels by binding to and promoting the degradation of the low-density lipoprotein receptor.Cholesterol efflux is mediated by transporters such as ATP-binding cassette transporter A1(ABCA1)and scavenger receptor class B type Ⅰ(SR-BI),with trafficking kinesin protein 2(TRAK2)potentially acting as an adaptor protein in this process.Investigating whether PCSK9 regulates cholesterol efflux through the TRAK2/ABCA1 pathway provides novel insights into its role in atherosclerosis. OBJECTIVE:To explore the effect of PCSK9 on cholesterol efflux in human monocyte-derived macrophages and its underlying mechanisms. METHODS:THP-1 cells were centrifuged and added with phorbol ester for 48 hours to induce differentiation into THP-1-derived macrophages,and then incubated with serum-free medium containing oxidized low-density lipoprotein for 24 hours to induce foam cell formation.(1)The foam cells were divided into four groups:control group,PCSK9 protein group,negative control group(non-targeted siRNA transfected foam cells),and PCSK9 siRNA transfection group.RT-qPCR and western blot assay were used to detect the mRNA and protein expressions of TRAK2,ABCA1,ABCG1,and SR-BI.(2)The foam cells were divided into three groups:negative control group,TRAK2 siRNA transfection group,and(TRAK2+PCSK9)siRNA co-transfection group.RT-qPCR and western blot assay were used to detect the mRNA and protein expressions of ABCA1.(3)The foam cells were divided into eight groups:control group,PCSK9 protein group,PCSK9 siRNA transfection group,TRAK2 siRNA transfection group,ABCA1 siRNA transfection group,(TRAK2+PCSK9)siRNA co-transfection group,(PCSK9+ABCA1)siRNA co-transfection group,and(TRAK2+ABCA1)siRNA co-transfection group.25-NBD-cholesterol was used to analyze the cholesterol efflux rate of foam cells. RESULTS AND CONCLUSION:(1)The TRAK2 mRNA and protein expression levels in the PCSK9 group were higher than those in the control group(P<0.05),whereas the expression levels of ABCA1,ABCG1,and SR-BI showed no significant difference compared to the control group.In the PCSK9 siRNA transfection group,TRAK2 mRNA and protein expression levels were lower than in the negative control group(P<0.001 and P<0.05,respectively),while ABCA1 mRNA and protein expression levels were higher than in the negative control group(P<0.001);no significant difference was observed in the mRNA and protein expression levels of ABCG1 and SR-BI compared to the negative control group.(2)In the TRAK2 siRNA transfection group,ABCA1 mRNA and protein expression levels were higher than those in the negative control group(P<0.001 and P<0.05,respectively),and in the(TRAK2+PCSK9)siRNA group,ABCA1 mRNA and protein expression levels were higher than those in the negative control group(P<0.001 and P<0.05,respectively).There was no significant difference in ABCA1 mRNA and protein expression levels between the(TRAK2+PCSK9)siRNA group and the TRAK2 siRNA group.(3)There was no significant difference in cholesterol efflux between the PCSK9 group and the control group.Cholesterol efflux in the PCSK9 siRNA,TRAK2 siRNA,and PCSK9+TRAK2 siRNA groups was higher than in the control group(P<0.001),with no significant difference in cholesterol efflux between these groups.Cholesterol efflux in the ABCA1 siRNA group was lower than in the control group(P<0.001);cholesterol efflux in the(PCSK9+ABCA1)siRNA group was lower than in the PCSK9 siRNA group(P<0.001),and cholesterol efflux in the(TRAK2+ABCA1)siRNA group was lower than in the TRAK2 siRNA group(P<0.001).These findings indicate that PCSK9 may regulate cholesterol efflux via the TRAK2/ABCA1 pathway.