Advertisement

LDL accelerates monocyte to macrophage differentiation: Effects on adhesion and anoikis

  • Author Footnotes
    1 Both authors contributed equally to this work.
    R. Escate
    Footnotes
    1 Both authors contributed equally to this work.
    Affiliations
    Cardiovascular Research Center (CSIC-ICCC), IIB-Sant Pau, Barcelona, Spain
    Search for articles by this author
  • Author Footnotes
    1 Both authors contributed equally to this work.
    T. Padro
    Footnotes
    1 Both authors contributed equally to this work.
    Affiliations
    Cardiovascular Research Center (CSIC-ICCC), IIB-Sant Pau, Barcelona, Spain
    Search for articles by this author
  • L. Badimon
    Correspondence
    Corresponding author. Cardiovascular Research Center, c/Sant Antoni Mª Claret 167, 08025 Barcelona, Spain.
    Affiliations
    Cardiovascular Research Center (CSIC-ICCC), IIB-Sant Pau, Barcelona, Spain

    Cardiovascular Research Chair, UAB, Barcelona, Spain
    Search for articles by this author
  • Author Footnotes
    1 Both authors contributed equally to this work.

      Highlights

      • Low-density-lipoproteins (LDL) accelerate monocyte-macrophage differentiation.
      • LDL upregulate expression of integrins in monocytes.
      • LDL reduce anoikis by affecting the caspase-cascade extrinsic pathway.
      • LDL link lipids, innate immunity and atherosclerosis.

      Abstract

      Background and aims

      High LDL triggers dyslipidemia and atherosclerosis, a chronic inflammatory disease with participation of the innate immunity system. Monocytes are recruited to areas of LDL-induced endothelial damage and initiate differentiation. This study was aimed to investigate the effects of LDL on the early transitional stages of monocyte differentiation into macrophages.

      Methods

      Blood monocytes, isolated from healthy donors by their adhesion properties, were exposed to native-LDL (1.80 mg/mL) for 48-h. Monocyte phenotype was assessed at transcript and miRNA levels by real-time PCR. Protein-expression was determined by western-blot and flow-cytometry.

      Results

      CD14 time-dependently decreased in adhered monocytes, reaching a >4fold decrease at transcript- and protein-levels after 7-days in culture when cells were already differentiated into macrophages. At 4-days differentiation, monocytes exposed to LDL reduced CD14-transcrition >1.5fold in mRNA (p = 0.002) and 34% CD14-protein (p = 0.039), whereas increased in CD16-expression (p = 0.019). Besides, LDL induced a significant increase in integrin CD49c (α3-subunit) at mRNA (>2fold, p = 0.008) and protein (>3fold, p = 0.045) level and a decrease in the apoptosis-effectors CASP8 and CASP3 (p = 0.002 and p = 0.035, respectively) as well as in the precursor form of the death-receptor DR5 (p = 0.045) without affecting its mRNA-expression level, suggesting a LDL-dependent post-transcriptional regulation of DR5. In silico prediction analysis indicated miR-126-3p as a candidate to regulate DR5-expression and miR-126-3p was shown affected by LDL reaching a significant increase (p = 0.033).

      Conclusions

      In differentiating human monocytes, LDL stimulates expression of cell-adhesion molecules and downregulates apoptosis-effectors, regulating anoikis and survival programs in the early stage macrophages.

      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

        • Ross R.
        Atherosclerosis–an inflammatory disease.
        N. Engl. J. Med. 1999; 340: 115-126
        • Behrendt D.
        • Ganz P.
        Endothelial function. From vascular biology to clinical applications.
        Am. J. Cardiol. 2002; 90: 40L-48L
        • Libby P.
        • Ridker P.M.
        • Maseri A.
        Inflammation and atherosclerosis.
        Circulation. 2002; 105: 1135-1143
        • Altman R.
        Risk factors in coronary atherosclerosis athero-inflammation: the meeting point.
        Thromb. J. 2003; 1: 4
        • Badimon L.
        • Padró T.
        • Vilahur G.
        Atherosclerosis, platelets and thrombosis in acute ischaemic heart disease.
        Eur. Heart J. Acute Cardiovasc. Care. 2012; 1: 60-74
        • Pritchard K.A.
        • Tota R.R.
        • Lin J.H.
        • et al.
        Native low density lipoprotein. Endothelial cell recruitment of mononuclear cells.
        Arterioscler. Thromb. Vasc. Biol. 1991; 11: 1175-1181
        • Han K.H.
        • Tangirala R.K.
        • Green S.R.
        • Quehenberger O.
        Chemokine receptor CCR2 expression and monocyte chemoattractant protein-1-mediated chemotaxis in human monocytes. A regulatory role for plasma LDL.
        Arterioscler. Thromb. Vasc. Biol. 1998; 18: 1983-1991
        • Smalley D.M.
        • Lin J.H.-C.
        • Curtis M.L.
        • Kobari Y.
        • Stemerman M.B.
        • Pritchard K.A.
        Native LDL increases endothelial cell adhesiveness by inducing intercellular adhesion molecule 1.
        Arterioscler. Thromb. Vasc. Biol. 1996; 16: 585-590
        • Serrano C.V.
        • Pesaro A.E.
        • de Lemos J.A.
        • et al.
        Native LDL-cholesterol mediated monocyte adhesion molecule overexpression is blocked by simvastatin.
        Cardiovasc. Drugs Ther. 2009; 23: 215-220
        • Margadant C.
        • Monsuur H.N.
        • Norman J.C.
        • Sonnenberg A.
        Mechanisms of integrin activation and trafficking.
        Curr. Opin. Cell Biol. 2011; 23: 607-614
        • Gilmore A.P.
        Anoikis.
        Cell Death Differ. 2005; 12: 1473-1477
        • Beauséjour M.
        • Thibodeau S.
        • Demers M.-J.
        • et al.
        Suppression of anoikis in human intestinal epithelial cells: differentiation state-selective roles of α2β1, α3β1, α5β1, and α6β4 integrins.
        BMC Cell Biol. 2013; 14: 53
        • Ammon C.
        • Meyer S.P.
        • Schwarzfischer L.
        • Krause S.W.
        • Andreesen R.
        • Kreutz M.
        Comparative analysis of integrin expression on monocyte-derived macrophages and monocyte-derived dendritic cells.
        Immunology. 2000; 100: 364-369
        • Gargiulo S.
        • Gamba P.
        • Testa G.
        • et al.
        Molecular signaling involved in oxysterol-induced β1-integrin over-expression in human macrophages.
        Int. J. Mol. Sci. 2012; 13: 14278-14293
        • Cha S.S.
        • Sung B.J.
        • Kim Y.A.
        • et al.
        Crystal structure of TRAIL-DR5 complex identifies a critical role of the unique frame insertion in conferring recognition specificity.
        J. Biol. Chem. 2000; 275: 31171-31177
        • Elmore S.
        Apoptosis: a review of programmed cell death.
        Toxicol. Pathol. 2007; 35: 495-516
        • Kothakota S.
        • Azuma T.
        • Reinhard C.
        • et al.
        Caspase-3-generated fragment of gelsolin: effector of morphological change in apoptosis.
        Science. 1997; 278: 294-298
        • Kamada S.
        • Kusano H.
        • Fujita H.
        • et al.
        A cloning method for caspase substrates that uses the yeast two-hybrid system: cloning of the antiapoptotic gene gelsolin.
        Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 8532-8537
        • Laguinge L.M.
        • Samara R.N.
        • Wang W.
        • et al.
        DR5 receptor mediates anoikis in human colorectal carcinoma cell lines.
        Cancer Res. 2008; 68: 909-917
        • Haselmann V.
        • Kurz A.
        • Bertsch U.
        • et al.
        Nuclear death receptor TRAIL-R2 inhibits maturation of let-7 and promotes proliferation of pancreatic and other tumor cells.
        Gastroenterology. 2014; 146: 278-290
        • Alexiou P.
        • Vergoulis T.
        • Gleditzsch M.
        • et al.
        miRGen 2.0: a database of microRNA genomic information and regulation.
        Nucleic Acids Res. 2010; 38: D137-D141
        • Borrell-Pagès M.
        • Romero J.C.
        • Badimon L.
        LRP5 negatively regulates differentiation of monocytes through abrogation of Wnt signalling.
        J. Cell. Mol. Med. 2014; 18: 314-325
        • Havel R.J.
        • Eder H.A.
        • Bragdon J.H.
        The distribution and chemical composition of ultracentrifugally separated lipoproteins in human serum.
        J. Clin. Investig. 1955; 34: 1345-1353
        • Ohkawa H.
        • Ohishi N.
        • Yagi K.
        Assay for lipid peroxides in animal tissues by thiobarbituric acid reaction.
        Anal. Biochem. 1979; 95: 351-358
        • Cal R.
        • Castellano J.
        • Revuelta-López E.
        • et al.
        Low-density lipoprotein receptor-related protein 1 mediates hypoxia-induced very low density lipoprotein-cholesteryl ester uptake and accumulation in cardiomyocytes.
        Cardiovasc. Res. 2012; 94: 469-479
        • Cornelissen M.
        • Philippé J.
        • De Sitter S.
        • De Ridder L.
        Annexin V expression in apoptotic peripheral blood lymphocytes: an electron microscopic evaluation.
        Apoptosis. 2002; 7: 41-47
        • Padró T.
        • Peña E.
        • García-Arguinzonis M.
        • Llorente-Cortes V.
        • Badimon L.
        Low-density lipoproteins impair migration of human coronary vascular smooth muscle cells and induce changes in the proteomic profile of myosin light chain.
        Cardiovasc Res. 2008; 77: 211-220
        • Griffiths-Jones S.
        • Saini H.K.
        • van Dongen S.
        • Enright A.J.
        miRBase: tools for microRNA genomics.
        Nucleic Acids Res. 2008; 36: D154-D158
        • Wagner J.
        • Riwanto M.
        • Besler C.
        • et al.
        Characterization of levels and cellular transfer of circulating lipoprotein-bound microRNAs.
        Arterioscler. Thromb. Vasc. Biol. 2013; 33: 1392-1400
        • Krüger J.
        • Rehmsmeier M.
        RNAhybrid: microRNA target prediction easy, fast and flexible.
        Nucleic Acids Res. 2006; 34: W451-W454
        • Ziegler-Heitbrock L.
        • Ancuta P.
        • Crowe S.
        • et al.
        Nomenclature of monocytes and dendritic cells in blood.
        Blood. 2010; 116: e74-80
        • Tanaka M.
        • Honda J.
        • Imamura Y.
        • Shiraishi K.
        • Tanaka K.
        • Oizumi K.
        Surface phenotype analysis of CD16+ monocytes from leukapheresis collections for peripheral blood progenitors.
        Clin. Exp. Immunol. 1999; 116: 57-61
        • Gleissner C.A.
        • Shaked I.
        • Erbel C.
        • Böckler D.
        • Katus H.A.
        • Ley K.
        CXCL4 downregulates the atheroprotective hemoglobin receptor CD163 in human macrophages.
        Circ. Res. 2010; 106: 203-211
        • Walczak H.
        • Degli-Esposti M.A.
        • Johnson R.S.
        • et al.
        TRAIL-R2: a novel apoptosis-mediating receptor for TRAIL.
        EMBO J. 1997; 16: 5386-5397
        • Lee M.-S.
        • Cherla R.P.
        • Lentz E.K.
        • Leyva-Illades D.
        • Tesh V.L.
        Signaling through C/EBP homologous protein and death receptor 5 and calpain activation differentially regulate THP-1 cell maturation-dependent apoptosis induced by Shiga toxin type 1.
        Infect. Immun. 2010; 78: 3378-3391
        • Lipsy R.J.
        Effective management of patients with dyslipidemia.
        Am. J. Manag. Care. 2003; 9: S39-S58
        • Perk J.
        • De Backer G.
        • Gohlke H.
        • et al.
        European guidelines on cardiovascular disease prevention in clinical practice (version 2012)” the fifth joint task force of the european society of cardiology and other societies on cardiovascular disease prevention in clinical practice (constituted by representatives of nine societies and by invited experts)..
        Eur. Heart J. 2012; 33: 1635-1701
        • Ziegler-Heitbrock H.W.
        • Ulevitch R.J.
        CD14: cell surface receptor and differentiation marker.
        Immunol. Today. 1993; 14: 121-125
        • Rothe G.
        • Gabriel H.
        • Kovacs E.
        • et al.
        Peripheral blood mononuclear phagocyte subpopulations as cellular markers in hypercholesterolemia.
        Arterioscler. Thromb. Vasc. Biol. 1996; 16: 1437-1447
        • Krychtiuk K.A.
        • Kastl S.P.
        • Pfaffenberger S.
        • et al.
        Association of small dense LDL serum levels and circulating monocyte subsets in stable coronary artery disease.
        PLoS One. 2015; 10: e0123367
        • Rogacev K.S.
        • Ulrich C.
        • Blömer L.
        • et al.
        Monocyte heterogeneity in obesity and subclinical atherosclerosis.
        Eur. Heart J. 2010; 31: 369-376
        • Kashiwagi M.
        • Imanishi T.
        • Tsujioka H.
        • et al.
        Association of monocyte subsets with vulnerability characteristics of coronary plaques as assessed by 64-slice multidetector computed tomography in patients with stable angina pectoris.
        Atherosclerosis. 2010; 212: 171-176
        • Merino A.
        • Buendia P.
        • Martin-Malo A.
        • Aljama P.
        • Ramirez R.
        • Carracedo J.
        Senescent CD14+CD16+ monocytes exhibit proinflammatory and proatherosclerotic activity.
        J. Immunol. 2011; 186: 1809-1815
        • Häkkinen T.
        • Karkola K.
        • Ylä-Herttuala S.
        Macrophages, smooth muscle cells, endothelial cells, and T-cells express CD40 and CD40L in fatty streaks and more advanced human atherosclerotic lesions. Colocalization with epitopes of oxidized low-density lipoprotein, scavenger receptor, and CD16 (Fc gammaRIII).
        Virchows Arch. 2000; 437: 396-405
        • Ziegler-Heitbrock L.
        The CD14+ CD16+ blood monocytes: their role in infection and inflammation.
        J. Leukoc. Biol. 2007; 81: 584-592
        • Schaer C.A.
        • Schoedon G.
        • Imhof A.
        • Kurrer M.O.
        • Schaer D.J.
        Constitutive endocytosis of CD163 mediates hemoglobin-heme uptake and determines the noninflammatory and protective transcriptional response of macrophages to hemoglobin.
        Circ. Res. 2006; 99: 943-950
        • Munn D.H.
        • Beall A.C.
        • Song D.
        • Wrenn R.W.
        • Throckmorton D.C.
        Activation-induced apoptosis in human macrophages: developmental regulation of a novel cell death pathway by macrophage colony-stimulating factor and interferon gamma.
        J. Exp. Med. 1995; 181: 127-136
        • Adams J.M.
        • Cory S.
        The Bcl-2 protein family: arbiters of cell survival.
        Science. 1998; 281: 1322-1326
        • Valley C.C.
        • Lewis A.K.
        • Mudaliar D.J.
        • et al.
        Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces death receptor 5 networks that are highly organized.
        J. Biol. Chem. 2012; 287: 21265-21278
        • Wilson N.S.
        • Dixit V.
        • Ashkenazi A.
        Death receptor signal transducers: nodes of coordination in immune signaling networks.
        Nat. Immunol. 2009; 10: 348-355
        • Baetu T.M.
        • Hiscott J.
        On the TRAIL to apoptosis.
        Cytokine Growth Factor Rev. 2002; 13: 199-207
        • Sun X.
        • Zhang M.
        • Sanagawa A.
        • et al.
        Circulating microRNA-126 in patients with coronary artery disease: correlation with LDL cholesterol.
        Thromb. J. 2012; 10: 16
        • Stupack D.G.
        • Cheresh D.A.
        Get a ligand, get a life: integrins, signaling and cell survival.
        J. Cell Sci. 2002; 115: 3729-3738
        • Hemler M.E.
        VLA proteins in the integrin family: structures, functions, and their role on leukocytes.
        Annu. Rev. Immunol. 1990; 8: 365-400