title: Cardiomyocyte contractile status is associated with differences in fibronectin and integrin interactions.
abstract: Integrins link the extracellular matrix (ECM) with the intracellular cytoskeleton and other cell adhesion-associated signaling proteins to function as mechanotransducers. However, direct quantitative measurements of the cardiomyocyte mechanical state and its relationship to the interactions between specific ECM proteins and integrins are lacking. The purpose of this study was to characterize the interactions between the ECM protein fibronectin (FN) and integrins in cardiomyocytes and to test the hypothesis that these interactions would vary during contraction and relaxation states in cardiomyocytes. Using atomic force microscopy, we quantified the unbinding force (adhesion force) and adhesion probability between integrins and FN and correlated these measurements with the contractile state as indexed by cell stiffness on freshly isolated mouse cardiomyocytes. Experiments were performed in normal physiological (control), high-K(+) (tonically contracted), or low-Ca(2+) (fully relaxed) solutions. Under control conditions, the initial peak of adhesion force between FN and myocyte alpha(3)beta(1)- and/or alpha(5)beta(1)-integrins was 39.6 +/- 1.3 pN. The binding specificity between FN and alpha(3)beta(1)- and alpha(5)beta(1)-integrins was verified by using monoclonal antibodies against alpha(3)-, alpha(5)-, alpha(3) + alpha(5)-, or beta(1)-integrin subunits, which inhibited binding by 48%, 65%, 70%, or 75%, respectively. Cytochalasin D or 2,3-butanedione monoxime (BDM), to disrupt the actin cytoskeleton or block myofilament function, respectively, significantly decreased the cell stiffness; however, the adhesion force and binding probability were not altered. Tonic contraction with high-K(+) solution increased total cell adhesion (1.2-fold) and cell stiffness (27.5-fold) compared with fully relaxed cells with low-Ca(2+) solution. However, it could be partially prevented by high-K(+) bath solution containing BDM, which suppresses contraction by inhibiting the actin-myosin interactions. Thus, our results demonstrate that integrin binding to FN is modulated by the contractile state of cardiac myocytes.
authors.affiliation: Dept. of Systems Biology and Translational Medicine, Texas A&M Health Science Center College of Medicine, College Station, TX 77843-1114, USA.
authors.forename: Xin
authors.initials: X
authors.lastname: Wu
authors.forename: Zhe
authors.initials: Z
authors.lastname: Sun
authors.forename: Andrea
authors.initials: A
authors.lastname: Foskett
authors.forename: Jerome P
authors.initials: JP
authors.lastname: Trzeciakowski
authors.forename: Gerald A
authors.initials: GA
authors.lastname: Meininger
authors.forename: Mariappan
authors.initials: M
authors.lastname: Muthuchamy
mesh_terms: D000199:Actins / Q000502:physiology; D000818:Animals; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000502:physiology; D005109:Extracellular Matrix / Q000502:physiology; D005353:Fibronectins / Q000502:physiology*; D016023:Integrins / Q000502:physiology*; D008297:Male; D051379:Mice; D008815:Mice, Inbred Strains; D023421:Models, Animal; D009200:Myocardial Contraction / Q000502:physiology*; D032383:Myocytes, Cardiac / Q000502:physiology*; D009218:Myosins / Q000502:physiology
chemical_list: D000199:Actins; D005353:Fibronectins; D016023:Integrins; D009218:Myosins
references.citation: Am J Cardiol. 1999 Jun 17;83(12A):38H-43H
references.pmid: 10750585
references.citation: J Cell Sci. 2008 Jun 1;121(11):1785-91
references.pmid: 18492792
references.citation: Am J Physiol Heart Circ Physiol. 2005 Dec;289(6):H2291-301
references.pmid: 16284104
references.citation: Curr Opin Cell Biol. 1991 Oct;3(5):786-91
references.pmid: 1931080
references.citation: Proc Natl Acad Sci U S A. 1983 Feb;80(4):1008-12
references.pmid: 6405378
references.citation: Annu Rev Cell Biol. 1987;3:57-85
references.pmid: 3318883
references.citation: Cell Motil. 1983;3(5-6):449-62
references.pmid: 6420066
references.citation: Soc Gen Physiol Ser. 1997;52:209-18
references.pmid: 9210231
references.citation: J Mol Cell Cardiol. 2001 Mar;33(3):521-32
references.pmid: 11181020
references.citation: Cardiovasc Res. 2004 Aug 1;63(2):283-92
references.pmid: 15249186
references.citation: Circulation. 2003 Feb 25;107(7):1046-52
references.pmid: 12600920
references.citation: Biophys J. 2003 Feb;84(2 Pt 1):1252-62
references.pmid: 12547805
references.citation: Nat Chem Biol. 2009 Jun;5(6):383-90
references.pmid: 19448607
references.citation: Circulation. 1997 Dec 2;96(11):4065-82
references.pmid: 9403633
references.citation: J Biol Chem. 1997 Feb 14;272(7):4500-8
references.pmid: 9020175
references.citation: Nat Struct Biol. 2001 May;8(5):412-6
references.pmid: 11323715
references.citation: Science. 1995 Feb 10;267(5199):883-5
references.pmid: 7846531
references.citation: Nat Rev Mol Cell Biol. 2001 Nov;2(11):793-805
references.pmid: 11715046
references.citation: J Cell Biol. 1995 Nov;131(3):791-805
references.pmid: 7593197
references.citation: Am J Physiol. 1996 Sep;271(3 Pt 2):H1250-5
references.pmid: 8853365
references.citation: Curr Hypertens Rep. 2004 Dec;6(6):424-9
references.pmid: 15527685
references.citation: Circ Res. 2001 Jun 8;88(11):1112-9
references.pmid: 11397776
references.citation: J Pathol. 1996 Jul;179(3):321-5
references.pmid: 8774490
references.citation: J Physiol. 2004 Oct 15;560(Pt 2):403-11
references.pmid: 15297568
references.citation: Biophys J. 2005 Dec;89(6):4320-9
references.pmid: 16183875
references.citation: Heart Fail Rev. 2000 Oct;5(3):239-50
references.pmid: 16228907
references.citation: J Biol Chem. 1998 Feb 27;273(9):5081-5
references.pmid: 9478959
references.citation: Cell. 2003 Jan 24;112(2):147-50
references.pmid: 12553903
references.citation: Science. 1993 May 21;260(5111):1124-7
references.pmid: 7684161
references.citation: Circ Res. 1991 Nov;69(5):1280-92
references.pmid: 1934358
references.citation: Curr Opin Cell Biol. 1991 Oct;3(5):841-8
references.pmid: 1931084
references.citation: J Clin Invest. 1968 Sep;47(9):1973-99
references.pmid: 5675423
references.citation: Lab Invest. 1991 Jan;64(1):65-75
references.pmid: 1824954
references.citation: J Biol Chem. 2006 May 19;281(20):14015-25
references.pmid: 16554304
references.citation: Cardiovasc Res. 2006 Jun 1;70(3):422-33
references.pmid: 16466704
references.citation: Nat Cell Biol. 2000 Jun;2(6):313-7
references.pmid: 10854320
references.citation: J Mol Cell Cardiol. 2001 Mar;33(3):487-501
references.pmid: 11181017
references.citation: Cell Biol Int. 1997 Nov;21(11):685-96
references.pmid: 9817809
references.citation: Am J Physiol Cell Physiol. 2008 Jul;295(1):C268-78
references.pmid: 18495809
references.citation: Biophys J. 2005 Oct;89(4):2888-98
references.pmid: 16055535
references.citation: Biochem Biophys Res Commun. 1999 Jun 16;259(3):645-50
references.pmid: 10364472
references.citation: J Cell Biol. 1993 May;121(3):673-8
references.pmid: 8387531
references.citation: Cardiovasc Res. 2004 Aug 15;63(3):381-90
references.pmid: 15276463
references.citation: Am J Physiol Heart Circ Physiol. 2001 Apr;280(4):H1427-33
references.pmid: 11247750
references.citation: Proc Natl Acad Sci U S A. 1990 Jan;87(1):404-8
references.pmid: 2296596
references.citation: Cardiovasc Pathol. 2004 Jan-Feb;13(1):11-9
references.pmid: 14761780
references.citation: Exp Biol Med (Maywood). 2004 Jun;229(6):538-45
references.pmid: 15169973
references.citation: Am J Physiol Heart Circ Physiol. 2005 Dec;289(6):H2526-35
references.pmid: 16100245
references.citation: J Mol Cell Cardiol. 1997 Sep;29(9):2533-43
references.pmid: 9299376
references.citation: J Clin Invest. 1992 Feb;89(2):392-401
references.pmid: 1531344
references.citation: J Physiol. 2005 Apr 15;564(Pt 2):603-17
references.pmid: 15718258
references.citation: J Physiol. 2008 Mar 15;586(6):1699-713
references.pmid: 18218680
references.citation: Arterioscler Thromb Vasc Biol. 1998 Sep;18(9):1363-70
references.pmid: 9743223
references.citation: J Biol Chem. 2001 Aug 10;276(32):30285-92
references.pmid: 11382763
references.citation: Cardiovasc Res. 2000 Jul;47(1):23-37
references.pmid: 10869527
references.citation: Diabetes. 2006 Nov;55(11):3104-11
references.pmid: 17065349
references.citation: Physiol Genomics. 2004 Aug 11;18(3):273-83
references.pmid: 15306692
references.citation: Dev Dyn. 2004 Sep;231(1):179-89
references.pmid: 15305298
references.citation: J Clin Invest. 1991 Nov;88(5):1737-46
references.pmid: 1834701
references.citation: J Biol Chem. 2003 Apr 18;278(16):13591-4
references.pmid: 12556452
references.citation: Circ Res. 1997 Feb;80(2):281-9
references.pmid: 9012750
references.citation: Cell. 1992 Apr 3;69(1):11-25
references.pmid: 1555235
references.citation: J Mol Cell Cardiol. 1998 May;30(5):947-55
references.pmid: 9618235
references.citation: J Cell Biol. 1998 Oct 5;143(1):241-52
references.pmid: 9763435
references.citation: Annu Rev Physiol. 1997;59:551-71
references.pmid: 9074777
references.citation: Circ Res. 2006 Dec 8;99(12):1403-10
references.pmid: 17095723
references.citation: J Clin Invest. 1992 Apr;89(4):1060-8
references.pmid: 1556175
references.citation: Cardiovasc Res. 1996 Sep;32(3):433-42
references.pmid: 8881506
references.citation: Trends Pharmacol Sci. 2001 May;22(5):254-60
references.pmid: 11339977
references.citation: J Cell Physiol. 1988 Jul;136(1):43-53
references.pmid: 3294238
references.citation: Am J Physiol. 1995 Jul;269(1 Pt 1):C286-92
references.pmid: 7631757
references.citation: Sci Am. 1986 Jun;254(6):42-51
references.pmid: 3010451
references.citation: Biochemistry. 2001 Nov 20;40(46):13915-24
references.pmid: 11705381
references.citation: Am J Physiol. 1992 Jun;262(6 Pt 2):H1861-6
references.pmid: 1535758
references.citation: J Cell Physiol. 2001 Oct;189(1):1-13
references.pmid: 11573199
references.citation: J Histochem Cytochem. 1984 Apr;32(4):383-8
references.pmid: 6707462
references.citation: Diabetes. 2005 Jun;54(6):1615-25
references.pmid: 15919781
references.citation: Circ Res. 2002 Jan 11;90(1):11-3
references.pmid: 11786511