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

Mehul R. Chorawala; Prem Prakash; Prakash Doddapattar; Manish Jain; Nirav Dhanesha; Anil K. Chauhan

doi:10.1055/s-0038-1661353

Thrombosis and Haemostasis, Table of Contents

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

¹Department of Internal Medicine, University of Iowa, Iowa City, Iowa, United States

,

Prem Prakash

¹Department of Internal Medicine, University of Iowa, Iowa City, Iowa, United States

,

Prakash Doddapattar

¹Department of Internal Medicine, University of Iowa, Iowa City, Iowa, United States

,

Manish Jain

¹Department of Internal Medicine, University of Iowa, Iowa City, Iowa, United States

,

Nirav Dhanesha

¹Department of Internal Medicine, University of Iowa, Iowa City, Iowa, United States

,

Anil K. Chauhan

¹Department of Internal Medicine, University of Iowa, Iowa City, Iowa, United States

› Author Affiliations

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

Full Text

References

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
2 Yellon DM, Hausenloy DJ. Myocardial reperfusion injury. N Engl J Med 2007; 357 (11) 1121-1135
3 Turer AT, Hill JA. Pathogenesis of myocardial ischemia-reperfusion injury and rationale for therapy. Am J Cardiol 2010; 106 (03) 360-368
4 Frangogiannis NG. The extracellular matrix in myocardial injury, repair, and remodeling. J Clin Invest 2017; 127 (05) 1600-1612
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
6 Wang Y, Reheman A, Spring CM. , et al. Plasma fibronectin supports hemostasis and regulates thrombosis. J Clin Invest 2014; 124 (10) 4281-4293
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
8 Wang Y, Gallant RC, Ni H. Extracellular matrix proteins in the regulation of thrombus formation. Curr Opin Hematol 2016; 23 (03) 280-287
9 White ES, Baralle FE, Muro AF. New insights into form and function of fibronectin splice variants. J Pathol 2008; 216 (01) 1-14
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
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
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
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
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
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
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
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
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
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
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
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
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
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
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
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
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
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
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
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
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
31 Guan JL, Trevithick JE, Hynes RO. Retroviral expression of alternatively spliced forms of rat fibronectin. J Cell Biol 1990; 110 (03) 833-847
32 Mehta J, Dinerman J, Mehta P. , et al. Neutrophil function in ischemic heart disease. Circulation 1989; 79 (03) 549-556
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
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
35 Jordan JE, Zhao ZQ, Vinten-Johansen J. The role of neutrophils in myocardial ischemia-reperfusion injury. Cardiovasc Res 1999; 43 (04) 860-878
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
37 Fliss H, Gattinger D. Apoptosis in ischemic and reperfused rat myocardium. Circ Res 1996; 79 (05) 949-956
38 Nian M, Lee P, Khaper N, Liu P. Inflammatory cytokines and postmyocardial infarction remodeling. Circ Res 2004; 94 (12) 1543-1553
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

Supplementary Material

Supplementary Material