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Thromb Haemost 2018; 118(08): 1450-1460
DOI: 10.1055/s-0038-1661353
Atherosclerosis and Ischaemic Disease
Georg Thieme Verlag KG Stuttgart · New York

Deletion of Extra Domain A of Fibronectin Reduces Acute Myocardial Ischaemia/Reperfusion Injury in Hyperlipidaemic Mice by Limiting Thrombo-Inflammation

Mehul R. Chorawala
1   Department of Internal Medicine, University of Iowa, Iowa City, Iowa, United States
,
Prem Prakash
1   Department of Internal Medicine, University of Iowa, Iowa City, Iowa, United States
,
Prakash Doddapattar
1   Department of Internal Medicine, University of Iowa, Iowa City, Iowa, United States
,
Manish Jain
1   Department of Internal Medicine, University of Iowa, Iowa City, Iowa, United States
,
Nirav Dhanesha
1   Department of Internal Medicine, University of Iowa, Iowa City, Iowa, United States
,
Anil K. Chauhan
1   Department of Internal Medicine, University of Iowa, Iowa City, Iowa, United States
› Author Affiliations Funding A.K.C. is supported by grants from the National Heart, Lung and Blood Institute of the National Institutes of Health grants (R35HL139926) and by Established Investigator Award 18EIA33900009 from American Heart Association.
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Publication History

18 March 2018

09 May 2018

Publication Date:
30 June 2018 (online)

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Abstract

Background Fibronectin splicing variant containing extra domain A (Fn-EDA), which is an endogenous ligand for Toll-like receptor 4 (TLR4), is present in negligible amounts in the plasma of healthy humans, but markedly elevated in patients with co-morbid conditions including diabetes and hyperlipidaemia, which are risk factors for myocardial infarction (MI). Very little is known about the role of Fn-EDA in the pathophysiology of acute MI under these co-morbid conditions.

Materials and Methods We determined the role of Fn-EDA in myocardial ischaemia/reperfusion (I/R) injury in the hyperlipidaemic apolipoprotein E-deficient (ApoE−/−) mice. Infarct size, plasma cardiac troponin I (cTnI) levels, intravascular thrombosis (CD41-positive), neutrophil infiltration (Ly6 B.2-positive), neutrophil extracellular traps (citrullinated H3-positive) and myocyte apoptosis (terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling-positive) were assessed in myocardial I/R injury model (1-hour ischaemia/23 hours of reperfusion).

Results Irrespective of gender, Fn-EDA−/−ApoE−/− mice exhibited smaller infarct size and decreased cTnI levels concomitant with reduced post-ischaemic intra-vascular thrombi, neutrophils influx, neutrophil extracellular traps and myocyte apoptosis (p < 0.05 vs. ApoE−/− mice). Genetic deletion of TLR4 attenuated myocardial I/R injury in ApoE−/− mice (p < 0.05 vs. ApoE−/− mice), but did not further reduce in Fn-EDA−/− ApoE−/− mice suggesting that Fn-EDA requires TLR4 to mediate myocardial I/R injury. Bone marrow transplantation experiments revealed that Fn-EDA exacerbates myocardial I/R injury through TLR4 expressed on the haematopoietic cells. Infusion of a specific inhibitor of Fn-EDA, 15 minutes post-reperfusion, into ApoE−/− mice attenuated myocardial I/R injury.

Conclusion Fn-EDA exacerbates TLR4-dependent myocardial I/R injury by promoting post-ischaemic thrombo-inflammatory response. Targeting Fn-EDA may reduce cardiac damage following coronary artery re-canalization after acute MI.

Keywords

fibronectins - thrombosis - inflammation - myocardial ischaemia - hyperlipidaemia

Supplementary Material

 

  • References

  • 1 Mozaffarian D, Benjamin EJ, Go AS. , et al; Writing Group Members; American Heart Association Statistics Committee; Stroke Statistics Subcommittee. Heart disease and stroke statistics-2016 update: a report from the American Heart Association. Circulation 2016; 133 (04) e38-e360
    CrossrefPubMedSearch in Google Scholar
  • 2 Yellon DM, Hausenloy DJ. Myocardial reperfusion injury. N Engl J Med 2007; 357 (11) 1121-1135
    CrossrefPubMedSearch in Google Scholar
  • 3 Turer AT, Hill JA. Pathogenesis of myocardial ischemia-reperfusion injury and rationale for therapy. Am J Cardiol 2010; 106 (03) 360-368
    CrossrefPubMedSearch in Google Scholar
  • 4 Frangogiannis NG. The extracellular matrix in myocardial injury, repair, and remodeling. J Clin Invest 2017; 127 (05) 1600-1612
    CrossrefPubMedSearch in Google Scholar
  • 5 Chauhan AK, Kisucka J, Cozzi MR. , et al. Prothrombotic effects of fibronectin isoforms containing the EDA domain. Arterioscler Thromb Vasc Biol 2008; 28 (02) 296-301
    CrossrefPubMedSearch in Google Scholar
  • 6 Wang Y, Reheman A, Spring CM. , et al. Plasma fibronectin supports hemostasis and regulates thrombosis. J Clin Invest 2014; 124 (10) 4281-4293
    CrossrefPubMedSearch in Google Scholar
  • 7 Prakash P, Kulkarni PP, Lentz SR, Chauhan AK. Cellular fibronectin containing extra domain A promotes arterial thrombosis in mice through platelet Toll-like receptor 4. Blood 2015; 125 (20) 3164-3172
    CrossrefPubMedSearch in Google Scholar
  • 8 Wang Y, Gallant RC, Ni H. Extracellular matrix proteins in the regulation of thrombus formation. Curr Opin Hematol 2016; 23 (03) 280-287
    CrossrefPubMedSearch in Google Scholar
  • 9 White ES, Baralle FE, Muro AF. New insights into form and function of fibronectin splice variants. J Pathol 2008; 216 (01) 1-14
    CrossrefPubMedSearch in Google Scholar
  • 10 Okamura Y, Watari M, Jerud ES. , et al. The extra domain A of fibronectin activates Toll-like receptor 4. J Biol Chem 2001; 276 (13) 10229-10233
    CrossrefPubMedSearch in Google Scholar
  • 11 Kanters SD, Banga JD, Algra A, Frijns RC, Beutler JJ, Fijnheer R. Plasma levels of cellular fibronectin in diabetes. Diabetes Care 2001; 24 (02) 323-327
    CrossrefPubMedSearch in Google Scholar
  • 12 Konieczyńska M, Bryk AH, Malinowski KP, Draga K, Undas A. Interplay between elevated cellular fibronectin and plasma fibrin clot properties in type 2 diabetes. Thromb Haemost 2017; 117 (09) 1671-1678
    Thieme ConnectPubMedSearch in Google Scholar
  • 13 Carlyle WC, Jacobson AW, Judd DL. , et al. Delayed reperfusion alters matrix metalloproteinase activity and fibronectin mRNA expression in the infarct zone of the ligated rat heart. J Mol Cell Cardiol 1997; 29 (09) 2451-2463
    CrossrefPubMedSearch in Google Scholar
  • 14 Knowlton AA, Connelly CM, Romo GM, Mamuya W, Apstein CS, Brecher P. Rapid expression of fibronectin in the rabbit heart after myocardial infarction with and without reperfusion. J Clin Invest 1992; 89 (04) 1060-1068
    CrossrefPubMedSearch in Google Scholar
  • 15 Arslan F, Smeets MB, Riem Vis PW. , et al. Lack of fibronectin-EDA promotes survival and prevents adverse remodeling and heart function deterioration after myocardial infarction. Circ Res 2011; 108 (05) 582-592
    CrossrefPubMedSearch in Google Scholar
  • 16 Dhanesha N, Ahmad A, Prakash P, Doddapattar P, Lentz SR, Chauhan AK. Genetic ablation of extra domain A of fibronectin in hypercholesterolemic mice improves stroke outcome by reducing thrombo-inflammation. Circulation 2015; 132 (23) 2237-2247
    CrossrefPubMedSearch in Google Scholar
  • 17 Doddapattar P, Gandhi C, Prakash P. , et al. Fibronectin splicing variants containing extra domain A promote atherosclerosis in mice through Toll-like receptor 4. Arterioscler Thromb Vasc Biol 2015; 35 (11) 2391-2400
    CrossrefPubMedSearch in Google Scholar
  • 18 Gondokaryono SP, Ushio H, Niyonsaba F. , et al. The extra domain A of fibronectin stimulates murine mast cells via toll-like receptor 4. J Leukoc Biol 2007; 82 (03) 657-665
    CrossrefPubMedSearch in Google Scholar
  • 19 Khan MM, Gandhi C, Chauhan N. , et al. Alternatively-spliced extra domain A of fibronectin promotes acute inflammation and brain injury after cerebral ischemia in mice. Stroke 2012; 43 (05) 1376-1382
    CrossrefPubMedSearch in Google Scholar
  • 20 Tan MH, Sun Z, Opitz SL, Schmidt TE, Peters JH, George EL. Deletion of the alternatively spliced fibronectin EIIIA domain in mice reduces atherosclerosis. Blood 2004; 104 (01) 11-18
    CrossrefPubMedSearch in Google Scholar
  • 21 Doddapattar P, Jain M, Dhanesha N, Lentz SR, Chauhan AK. Fibronectin containing extra domain A induces plaque destabilization in the innominate artery of aged apolipoprotein E-deficient mice. Arterioscler Thromb Vasc Biol 2018; 38 (03) 500-508
    CrossrefPubMedSearch in Google Scholar
  • 22 Muro AF, Chauhan AK, Gajovic S. , et al. Regulated splicing of the fibronectin EDA exon is essential for proper skin wound healing and normal lifespan. J Cell Biol 2003; 162 (01) 149-160
    CrossrefPubMedSearch in Google Scholar
  • 23 Michael LH, Entman ML, Hartley CJ. , et al. Myocardial ischemia and reperfusion: a murine model. Am J Physiol 1995; 269 (6 Pt 2): H2147-H2154
    PubMedSearch in Google Scholar
  • 24 Gandhi C, Motto DG, Jensen M, Lentz SR, Chauhan AK. ADAMTS13 deficiency exacerbates VWF-dependent acute myocardial ischemia/reperfusion injury in mice. Blood 2012; 120 (26) 5224-5230
    CrossrefPubMedSearch in Google Scholar
  • 25 Dhanesha N, Doddapattar P, Chorawala MR. , et al. ADAMTS13 retards progression of diabetic nephropathy by inhibiting intrarenal thrombosis in mice. Arterioscler Thromb Vasc Biol 2017; 37 (07) 1332-1338
    CrossrefPubMedSearch in Google Scholar
  • 26 van Keulen JK, de Kleijn DP, Nijhuis MM. , et al. Levels of extra domain A containing fibronectin in human atherosclerotic plaques are associated with a stable plaque phenotype. Atherosclerosis 2007; 195 (01) e83-e91
    CrossrefPubMedSearch in Google Scholar
  • 27 de Boer OJ, Li X, Teeling P. , et al. Neutrophils, neutrophil extracellular traps and interleukin-17 associate with the organisation of thrombi in acute myocardial infarction. Thromb Haemost 2013; 109 (02) 290-297
    Search in Google Scholar
  • 28 Savchenko AS, Borissoff JI, Martinod K. , et al. VWF-mediated leukocyte recruitment with chromatin decondensation by PAD4 increases myocardial ischemia/reperfusion injury in mice. Blood 2014; 123 (01) 141-148
    CrossrefPubMedSearch in Google Scholar
  • 29 Oyama J, Blais Jr C, Liu X. , et al. Reduced myocardial ischemia-reperfusion injury in toll-like receptor 4-deficient mice. Circulation 2004; 109 (06) 784-789
    CrossrefPubMedSearch in Google Scholar
  • 30 Spinelli SL, Casey AE, Pollock SJ. , et al. Platelets and megakaryocytes contain functional nuclear factor-kappaB. Arterioscler Thromb Vasc Biol 2010; 30 (03) 591-598
    CrossrefPubMedSearch in Google Scholar
  • 31 Guan JL, Trevithick JE, Hynes RO. Retroviral expression of alternatively spliced forms of rat fibronectin. J Cell Biol 1990; 110 (03) 833-847
    CrossrefPubMedSearch in Google Scholar
  • 32 Mehta J, Dinerman J, Mehta P. , et al. Neutrophil function in ischemic heart disease. Circulation 1989; 79 (03) 549-556
    CrossrefPubMedSearch in Google Scholar
  • 33 Litt MR, Jeremy RW, Weisman HF, Winkelstein JA, Becker LC. Neutrophil depletion limited to reperfusion reduces myocardial infarct size after 90 minutes of ischemia. Evidence for neutrophil-mediated reperfusion injury. Circulation 1989; 80 (06) 1816-1827
    CrossrefPubMedSearch in Google Scholar
  • 34 Dreyer WJ, Michael LH, West MS. , et al. Neutrophil accumulation in ischemic canine myocardium. Insights into time course, distribution, and mechanism of localization during early reperfusion. Circulation 1991; 84 (01) 400-411
    CrossrefPubMedSearch in Google Scholar
  • 35 Jordan JE, Zhao ZQ, Vinten-Johansen J. The role of neutrophils in myocardial ischemia-reperfusion injury. Cardiovasc Res 1999; 43 (04) 860-878
    CrossrefPubMedSearch in Google Scholar
  • 36 Clark SR, Ma AC, Tavener SA. , et al. Platelet TLR4 activates neutrophil extracellular traps to ensnare bacteria in septic blood. Nat Med 2007; 13 (04) 463-469
    CrossrefPubMedSearch in Google Scholar
  • 37 Fliss H, Gattinger D. Apoptosis in ischemic and reperfused rat myocardium. Circ Res 1996; 79 (05) 949-956
    CrossrefPubMedSearch in Google Scholar
  • 38 Nian M, Lee P, Khaper N, Liu P. Inflammatory cytokines and postmyocardial infarction remodeling. Circ Res 2004; 94 (12) 1543-1553
    CrossrefPubMedSearch in Google Scholar
  • 39 Nakamura M, Wang NP, Zhao ZQ. , et al. Preconditioning decreases Bax expression, PMN accumulation and apoptosis in reperfused rat heart. Cardiovasc Res 2000; 45 (03) 661-670
    CrossrefPubMedSearch in Google Scholar