Advertisement

Modulation of vascular endothelial inflammatory response by proprotein convertase subtilisin-kexin type 9

      Highlights

      • LPS uptake in HUVECs is dose-dependent on LDL concentration.
      • PCSK9 dose-dependently inhibits LDLR-mediated endocytosis of LPS in HUVECs.
      • Decreased PCSK9 increases endothelial cell LPS uptake and proinflammatory response.

      Abstract

      Aims

      Endotoxins carried within LDL are cleared from the circulation via hepatic LDL receptor (LDLR)-mediated endocytosis. Proprotein convertase subtilisin-kexin type 9 (PCSK9) reduces this clearance by down-regulating LDLR density on hepatocytes. In addition to hepatocytes, vascular endothelial cells also express receptor targets of PCSK9, including LDLR. Therefore we hypothesized that PCSK9 may regulate vascular endothelial cell uptake of lipopolysaccharide (LPS) and alter the vascular endothelial cell inflammatory response.

      Methods and results

      We found that LPS is internalized by human umbilical vein vascular endothelial cells (HUVECs) and LPS uptake dose-dependently increased with increasing LDL concentration. Intracellular LPS co-localized with LDL. PCSK9 and, separately, blocking antibodies against LDLR, dose-dependently decreased the vascular endothelial cell uptake of LPS and, furthermore, inhibition of endocytosis using Dynasore blocked LPS uptake. In contrast, blocking antibodies against TLR4 did not alter LPS uptake. PCSK9 decreased the LPS-induced proinflammatory response (IL-6 and IL-8 gene expression and protein secretion, and VCAM-1/ICAM-1 expression) in vascular endothelial cells. In addition, a decrease in PCSK9 and increase in LDLR, mediated by triciribine or siPCSK9, increased LPS uptake and the LPS-induced proinflammatory response. Similar results were also found in aortic vascular tissue from Pcsk9−/− mice after LPS injection.

      Conclusions

      Our data suggest that, similar to PCSK9 treatment in hepatocytes, PCSK9 reduces vascular endothelial cell uptake of LPS via LDLR-mediated endocytosis. Consequently, PCSK9 decreases the LPS-induced proinflammatory response in vascular endothelial cells. These results raise the possibility that PCSK9 inhibition may have additional effects on vascular endothelial inflammation via this alternative pathway, beyond the known effects of PCSK9 inhibition on LDL lowering and hepatic endotoxin clearance.

      Graphical abstract

      Keywords

      To read this article in full you will need to make a payment

      Purchase one-time access:

      Academic & Personal: 24 hour online accessCorporate R&D Professionals: 24 hour online access
      One-time access price info
      • For academic or personal research use, select 'Academic and Personal'
      • For corporate R&D use, select 'Corporate R&D Professionals'

      Subscribe:

      Subscribe to Atherosclerosis
      Already a print subscriber? Claim online access
      Already an online subscriber? Sign in
      Institutional Access: Sign in to ScienceDirect

      References

        • Allard D.
        • Amsellem S.
        • Abifadel M.
        • Trillard M.
        • Devillers M.
        • Luc G.
        • et al.
        Novel mutations of the PCSK9 gene cause variable phenotype of autosomal dominant hypercholesterolemia.
        Hum. Mutat. 2005; 26: 497
        • Grefhorst A.
        • McNutt M.C.
        • Lagace T.A.
        • Horton J.D.
        Plasma PCSK9 preferentially reduces liver LDL receptors in mice.
        J. Lipid Res. 2008; 49: 1303-1311
        • Lagace T.A.
        PCSK9 and LDLR degradation: regulatory mechanisms in circulation and in cells.
        Curr. Opin. Lipidol. 2014; 25: 387-393
        • Goldstein J.L.
        • Brown M.S.
        • Anderson R.G.
        • Russell D.W.
        • Schneider W.J.
        Receptor-mediated endocytosis: concepts emerging from the LDL receptor system.
        Annu. Rev. Cell Biol. 1985; 1: 1-39
        • Lagace T.A.
        • Curtis D.E.
        • Garuti R.
        • McNutt M.C.
        • Park S.W.
        • Prather H.B.
        • et al.
        Secreted PCSK9 decreases the number of LDL receptors in hepatocytes and in livers of parabiotic mice.
        J. Clin. Invest. 2006; 116: 2995-3005
        • Cohen J.C.
        • Boerwinkle E.
        • Mosley Jr., T.H.
        • Hobbs H.H.
        Sequence variations in PCSK9, low LDL, and protection against coronary heart disease.
        N. Engl. J. Med. 2006; 354: 1264-1272
        • Kathiresan S.
        • Melander O.
        • Guiducci C.
        • Surti A.
        • Burtt N.P.
        • Rieder M.J.
        • et al.
        Six new loci associated with blood low-density lipoprotein cholesterol, high-density lipoprotein cholesterol or triglycerides in humans.
        Nat. Genet. 2008; 40: 189-197
        • Sabatine M.S.
        • Giugliano R.P.
        • Keech A.C.
        • Honarpour N.
        • Wiviott S.D.
        • Murphy S.A.
        • et al.
        Evolocumab and clinical outcomes in patients with cardiovascular disease.
        N. Engl. J. Med. 2017; 376: 1713-1722
        • Levels J.H.
        • Abraham P.R.
        • van den Ende A.
        • van Deventer S.J.
        Distribution and kinetics of lipoprotein-bound endotoxin.
        Infect. Immun. 2001; 69: 2821-2828
        • Levels J.H.
        • Lemaire L.C.
        • van den Ende A.E.
        • van Deventer S.J.
        • van Lanschot J.J.
        Lipid composition and lipopolysaccharide binding capacity of lipoproteins in plasma and lymph of patients with systemic inflammatory response syndrome and multiple organ failure.
        Crit. Care Med. 2003; 31: 1647-1653
        • Levels J.H.
        • Marquart J.A.
        • Abraham P.R.
        • van den Ende A.E.
        • Molhuizen H.O.
        • van Deventer S.J.
        • et al.
        Lipopolysaccharide is transferred from high-density to low-density lipoproteins by lipopolysaccharide-binding protein and phospholipid transfer protein.
        Infect. Immun. 2005; 73: 2321-2326
        • Walley K.R.
        • Thain K.R.
        • Russell J.A.
        • Reilly M.P.
        • Meyer N.J.
        • Ferguson J.F.
        • et al.
        PCSK9 is a critical regulator of the innate immune response and septic shock outcome.
        Sci. Transl. Med. 2014; 6: 258ra143
        • Topchiy E.
        • Cirstea M.
        • Kong H.J.
        • Boyd J.H.
        • Wang Y.
        • Russell J.A.
        • et al.
        Lipopolysaccharide is cleared from the circulation by hepatocytes via the low density lipoprotein receptor.
        PLoS One. 2016; 11e0155030
        • Wyne K.L.
        • Pathak K.
        • Seabra M.C.
        • Hobbs H.H.
        Expression of the VLDL receptor in endothelial cells.
        Arterioscler. Thromb. Vasc. Biol. 1996; 16: 407-415
        • Hu B.
        • Li D.
        • Sawamura T.
        • Mehta J.L.
        Oxidized LDL through LOX-1 modulates LDL-receptor expression in human coronary artery endothelial cells.
        Biochem. Biophys. Res. Commun. 2003; 307: 1008-1012
        • Zhang X.
        • Sessa W.C.
        • Fernandez-Hernando C.
        Endothelial transcytosis of lipoproteins in atherosclerosis.
        Front Cardiovasc Med. 2018; 5: 130
        • Bjune K.
        • Wierod L.
        • Naderi S.
        Triciribine increases LDLR expression and LDL uptake through stabilization of LDLR mRNA.
        Sci. Rep. 2018; 816174
        • Walley K.R.
        • Francis G.A.
        • Opal S.M.
        • Stein E.A.
        • Russell J.A.
        • Boyd J.H.
        The central role of Proprotein convertase subtilisin/kexin type 9 in septic pathogen lipid transport and clearance.
        Am. J. Respir. Crit. Care Med. 2015; 192: 1275-1286
        • Dwivedi D.J.
        • Grin P.M.
        • Khan M.
        • Prat A.
        • Zhou J.
        • Fox-Robichaud A.E.
        • et al.
        Differential expression of PCSK9 modulates infection, inflammation, and coagulation in a murine model of sepsis.
        Shock. 2016; 46: 672-680
        • Frank-Kamenetsky M.
        • Grefhorst A.
        • Anderson N.N.
        • Racie T.S.
        • Bramlage B.
        • Akinc A.
        • et al.
        Therapeutic RNAi targeting PCSK9 acutely lowers plasma cholesterol in rodents and LDL cholesterol in nonhuman primates.
        Proc. Natl. Acad. Sci. U. S. A. 2008; 105: 11915-11920
        • Vita J.A.
        Endothelial function.
        Circulation. 2011; 124: e906-e912
        • Sima A.V.
        • Stancu C.S.
        • Simionescu M.
        Vascular endothelium in atherosclerosis.
        Cell Tissue Res. 2009; 335: 191-203
        • Szeto C.C.
        • Kwan B.C.
        • Chow K.M.
        • Lai K.B.
        • Chung K.Y.
        • Leung C.B.
        • et al.
        Endotoxemia is related to systemic inflammation and atherosclerosis in peritoneal dialysis patients.
        Clin. J. Am. Soc. Nephrol. 2008; 3: 431-436
        • Kaynar A.M.
        • Yende S.
        • Zhu L.
        • Frederick D.R.
        • Chambers R.
        • Burton C.L.
        • et al.
        Effects of intra-abdominal sepsis on atherosclerosis in mice.
        Crit. Care. 2014; 18: 469
        • Biedron R.
        • Perun A.
        • Jozefowski S.
        CD36 differently regulates macrophage responses to smooth and rough lipopolysaccharide.
        PLoS One. 2016; 11e0153558
        • Miyake K.
        Innate recognition of lipopolysaccharide by Toll-like receptor 4-MD-2.
        Trends Microbiol. 2004; 12: 186-192
        • Shi J.
        • Zhao Y.
        • Wang Y.
        • Gao W.
        • Ding J.
        • Li P.
        • et al.
        Inflammatory caspases are innate immune receptors for intracellular LPS.
        Nature. 2014; 514: 187-192
        • Kayagaki N.
        • Wong M.T.
        • Stowe I.B.
        • Ramani S.R.
        • Gonzalez L.C.
        • Akashi-Takamura S.
        • et al.
        Noncanonical inflammasome activation by intracellular LPS independent of TLR4.
        Science. 2013; 341: 1246-1249
        • Vanaja S.K.
        • Russo A.J.
        • Behl B.
        • Banerjee I.
        • Yankova M.
        • Deshmukh S.D.
        • et al.
        Bacterial outer membrane vesicles mediate cytosolic localization of LPS and caspase-11 activation.
        Cell. 2016; 165: 1106-1119
        • Leung A.K.K.
        • Genga K.R.
        • Topchiy E.
        • Cirstea M.
        • Shimada T.
        • Fjell C.
        • et al.
        Reduced Proprotein convertase subtilisin/kexin 9 (PCSK9) function increases lipoteichoic acid clearance and improves outcomes in Gram positive septic shock patients.
        Sci. Rep. 2019; 910588