Cardiology Division Dichek Cardiovascular Research Laboratory

Contact Information:

David A. Dichek, M.D.
Professor of Medicine
John L. Locke, Jr. Family Endowed Chair in Medicine
Associate Director for Research, Division of Cardiology
Department of Medicine
University of Washington

U.S. Mail
1959 N.E. Pacific Street
Box 357710
Seattle, WA 98195-7710

Courier (FEDEX, UPS, etc.):
Rm K143A Health Sciences Bldg.
1959 NE Pacific St.
Seattle, WA 98195

Tel (206) 685-6959
FAX (206) 221-6346

Email: ddichek@u.washington.edu

Overview of the Laboratory:

Our major projects share a common goal: to discover the mechanisms that cause vascular disease and to translate these discoveries into novel therapies that can prevent or reverse vascular disease.  There are 3 major projects:

• Development of gene-transfer vectors capable of mediating long-term recombinant gene expression in blood vessels and use of these vectors to prevent or reverse atherosclerosis.

• Investigation of the role of the urokinase plasminogen activator (uPA)/plasminogen system in the development of atherosclerosis and plaque rupture.

• Investigation of the role of transforming growth factor β (TGF-β) signaling in the development and prevention of vascular disease and in the formation of blood vessels during embryogenesis.

Significance:

Cardiovascular disease, especially atherosclerosis, is a leading cause of death and disability throughout the world.  During the past twenty years, medical therapy and life-style changes have decreased the prevalence of cardiovascular disease in the “Western” world.  However, cardiovascular disease is far from being eliminated and is increasing in the developing world.  Targeted molecular interventions—that are based on a detailed understanding of the pathogenesis of cardiovascular diseases—hold great promise for preventing and treating cardiovascular disease. 

Approaches, Contributions, and Current Work:

We use techniques and approaches of molecular biology, biochemistry, genetics, virology, immunology, developmental biology, and whole-animal physiology.

Vascular gene therapy:

Since 1988, (1) our laboratory has worked on improving gene-transfer vectors and evaluating their ability to express genes in the artery wall.  We clarified the promise of adenoviral vectors to express genes in injured and uninjured arteries and we also identified their limitations including brevity of expression and proinflammatory effects. (2-7)  Recently, we discovered that helper-dependent adenoviral vectors (which lack all viral genes) can express therapeutic genes stably in the artery wall, with minimal associated inflammation. (8, 9)  Clinical applications of this approach include gene transfer to blood vessels that prevents or reverses atherosclerosis and prevents venous bypass graft disease.

Currently we use helper-dependent adenoviral vectors to express genes in rabbit carotid arteries and test whether expression of these genes can prevent atherosclerosis. (9-11)  We found that expression of apolipoprotein A-I in arterial endothelium using a helper-dependent adenoviral vector can prevent early atherosclerosis in cholesterol-fed rabbits. (12)  New animal models are under development that will allow more extensive testing of this therapeutic approach in arteries as well as grafted veins.  We are also constructing and testing new expression cassettes that we anticipate will allow durable high-level therapeutic gene expression in endothelium. (13) 

The uPA/plasminogen system and vascular disease:

Urokinase-type plasminogen activator (uPA) is a serine protease that converts the zymogen plasminogen to plasmin (a broadly active protease).  uPA circulates in plasma and is expressed by cells in the artery wall.  We became interested in uPA because of its ability to act as a therapeutic agent by promoting lysis of occlusive intravascular thrombi.  We first proposed using uPA (and the related enzyme tPA) for antithrombotic gene therapy delivered either directly to the vessel wall or from the surface of thrombogenic intravascular devices. (1, 14-16)  We then discovered that uPA—when expressed at increased levels in the artery wall of hyperlipidemic rabbits or mice—increases atherosclerosis and causes arterial constriction. (17, 18)  More recently we discovered that increased expression of uPA in mouse plaque macrophages causes plaque rupture. (19)  These results suggest that uPA expression in the vessel wall contributes to the progression of vascular disease.

Currently our work is aimed at defining the mechanisms through which uPA accelerates atherosclerosis and causes plaque rupture.  Insights into the mechanisms of uPA-accelerated atherosclerosis and plaque rupture may reveal new strategies for preventing atherosclerosis and its complications.

TGF-β signaling in vascular disease and development:

Transforming growth factor beta-1 (TGF-β) is a pleiotropic cytokine that is expressed in the artery wall and circulates in plasma.  We use mouse models of gene transfer, germ-line transgenesis, and gene knockout to uncover the roles of TGF-β signaling in the vasculature. 

We used gene transfer to discover that increased TGF-β expression in the artery wall of rodents promotes intimal growth, and we identified the mechanisms through which TGF-β acts. (20-22)  We used germ-line transgenesis to discover that increased TGF-β expression in the vasculature during embryogenesis disrupts vasculogenesis, and causes embryonic death. (23)  To bypass this embryonic lethality and determine the role of TGF-β signaling in adult vascular disease, we used the “tet” system to obtain regulated postnatal cardiovascular overexpression of TGF-β. (24)  These experiments revealed that increased expression of TGF-β in adults retards atherosclerosis and slows aneurysm progression. (25) 

To define the roles played by normal levels of vascular TGF-β signaling we used the “Cre-Lox” system to achieve cardiovasculature-specific deletion of the type II TGF-β receptor during mouse development.  This deletion eliminates TGF-β signaling in vascular smooth muscle.  Embryonic lethality caused by this deletion revealed an essential role for TGF-β signaling in vascular morphogenesis, smooth muscle cell differentiation, and matrix synthesis during mid-gestation. (26, 27)

Currently we are using inducible knockout technology to uncover the role of vascular TGF-β signaling in adult mice.  We anticipate that insights from these experiments will clarify the role of TGF-β signaling in vascular homeostasis, atherosclerosis, and aneurysm formation during adult life and will reveal novel strategies for preserving vascular health.

Publications cited above:

1.         Dichek DA, Neville RF, Zwiebel JA, Freeman SM, Leon MB, Anderson WF. Seeding of intravascular stents with genetically engineered endothelial cells. Circulation. 1989;80:1347–1353.
2.         Lee SW, Trapnell BC, Rade JJ, Virmani R, Dichek DA. In vivo adenoviral vector-mediated gene transfer into balloon-injured rat carotid arteries. Circ. Res. 1993;73:797–807.
3.         Schulick AH, Dong G, Newman KD, Virmani R, Dichek DA. Endothelium-specific in vivo gene transfer. Circ. Res. 1995;77:475–485.
4.         Schulick AH, Newman KD, Virmani R, Dichek DA. In vivo gene transfer into injured carotid arteries. Optimization and evaluation of acute toxicity. Circulation. 1995;91:2407–2414.
5.         Newman KD, Dunn PF, Owens JW, Schulick AH, Virmani R, Sukhova G, Libby P, Dichek DA. Adenovirus-mediated gene transfer into normal rabbit arteries results in prolonged vascular cell activation, inflammation, and neointimal hyperplasia. J. Clin. Invest. 1995;96:2955–2965.
6.         Schulick AH, Vassalli G, Dunn PF, Dong G, Rade JJ, Zamarron C, Dichek DA. Established immunity precludes adenovirus-mediated gene transfer in rat carotid arteries. Potential for immunosuppression and vector engineering to overcome barriers of immunity. J. Clin. Invest. 1997;99:209–219.
7.         Rade JJ, Schulick AH, Virmani R, Dichek DA. Local adenoviral-mediated expression of recombinant hirudin reduces neointimal formation after arterial injury. Nat. Med. 1996;2:293–298.
8.         Wen S, Graf S, Massey PG, Dichek DA. Improved vascular gene transfer with a helper-dependent adenoviral vector. Circulation. 2004;110:1484-1491.
9.         Jiang B, Qian K, Du L, Luttrell I, Chitaley K, Dichek DA. Helper-dependent adenovirus is superior to first-generation adenovirus for expressing transgenes in atherosclerosis-prone arteries. Arterioscler. Thromb. Vasc. Biol. 2011;31:1317–1325.
10.       Schneider DB, Vassalli G, Wen S, Driscoll RM, Sassani AB, DeYoung MB, Linnemann R, Virmani R, Dichek DA. Expression of Fas ligand in arteries of hypercholesterolemic rabbits accelerates atherosclerotic lesion formation. Arterioscler. Thromb. Vasc. Biol. 2000;20:298–308.
11.       Du L, Dronadula N, Tanaka S, Dichek DA. Helper-dependent adenoviral vector achieves prolonged, stable expression of IL-10 in rabbit carotid arteries but does not limit early atherogenesis. Hum Gen Ther. 2011;22:959–968.
12.       Flynn R, Qian K, Tang C, Dronadula N, Buckler J, Jiang B, Wen S, Dichek H, Dichek D. Expression of apolipoprotein A-I in rabbit carotid endothelium protects against atherosclerosis. Mol Ther. 2011;19:1833–1841.
13.       Dronadula N, Du L, Flynn R, Buckler JM, Kho J, Jiang Z, Tanaka S, Dichek DA. Construction of a novel expression cassette for increasing transgene expression in vivo in endothelial cells of large blood vessels. Gene Ther. 2011;18:501–508.
14.       Dichek DA, Lee SW, Nguyen NH. Characterization of recombinant plasminogen activator production by primate endothelial cells transduced with retroviral vectors. Blood. 1994;84:504–516.
15.       Dichek DA, Anderson J, Kelly AB, Hanson SR, Harker LA. Enhanced in vivo antithrombotic effects of endothelial cells expressing recombinant plasminogen activators transduced with retroviral vectors. Circulation. 1996;93:301–309.
16.       Shayani V, Newman KD, Dichek DA. Optimization of recombinant t-PA secretion from seeded vascular grafts. J. Surg. Res. 1994;57:495–504.
17.       Cozen AE, Moriwaki H, Kremen M, DeYoung MB, Dichek HL, Slezicki KI, Young SG, Veniant M, Dichek DA. Macrophage-targeted overexpression of urokinase causes accelerated atherosclerosis, coronary artery occlusions, and premature death. Circulation. 2004;109:2129–2135.
18.       Falkenberg M, Tom C, DeYoung MB, Wen S, Linnemann R, Dichek DA. Increased expression of urokinase during atherosclerotic lesion development causes arterial constriction and lumen loss, and accelerates lesion growth. Proc. Natl. Acad. Sci. U. S. A. 2002;99:10665–10670.
19.       Hu JH, Du L, Chu T, Otsuka G, Dronadula N, Jaffe M, Gill SE, Parks WC, Dichek DA. Overexpression of urokinase by plaque macrophages causes histologic features of plaque rupture and increases vascular MMP activity in aged apo E-Null mice. Circulation. 2010;121:1637–1644.
20.       Schulick AH, Taylor AJ, Zuo W, Qiu C-B, Dong G, Woodward RN, Agah R, Roberts AB, Virmani R, Dichek DA. Overexpression of transforming growth factor b1 in arterial endothelium causes hyperplasia, apoptosis, and cartilaginous metaplasia. Proc. Natl. Acad. Sci. U. S. A. 1998;95:6983–6988.
21.       Otsuka G, Agah R, Frutkin AD, Wight TN, Dichek DA. Transforming growth factor beta 1 induces neointima formation through plasminogen activator inhibitor-1-dependent pathways. Arterioscler. Thromb. Vasc. Biol. 2006;26:737–743.
22.       Otsuka G, Stempien-Otero A, Frutkin AD, Dichek DA. Mechanisms of TGF-beta1–induced intimal growth: Plasminogen-independent activities of plasminogen activator inhibitor-1 and heterogeneous origin of intimal cells. Circ. Res. 2007;100:1300–1307.
23.       Agah R, Prasad KSS, Linnemann R, Firpo MT, Quertermous T, Dichek DA. Cardiovascular overexpression of transforming growth factor-b1 causes abnormal yolk sac vasculogenesis and early embryonic death. Circ. Res. 2000;86:1024–1030.
24.       Lee S, Agah R, Xiao M, Frutkin AD, Kremen M, Shi H, Dichek DA. In vivo expression of a conditional TGF-beta1 transgene: no evidence for TGF-beta1 transgene expression in SM22alpha-tTA transgenic mice. J. Mol. Cell. Cardiol. 2006;40:148–156.
25.       Frutkin AD, Otsuka G, Stempien-Otero A, Sesti C, Du L, Jaffe M, Dichek HL, Pennington CJ, Edwards DR, Nieves-Cintron M, Minter D, Preusch M, Hu JH, Marie JC, Dichek DA. TGF-beta1 limits plaque growth, stabilizes plaque structure, and prevents aortic dilation in Apolipoprotein E-null mice. Arterioscler. Thromb. Vasc. Biol. 2009;29:1251–1257.
26.       Frutkin AD, Shi H, Otsuka G, Leveen P, Karlsson S, Dichek DA. A critical developmental role for tgfbr2 in myogenic cell lineages is revealed in mice expressing SM22-Cre, not SMMHC-Cre. J. Mol. Cell. Cardiol. 2006;41:724–731.
27.       Jaffe M, Sesti C, Washington I, Du L, Dronadula N, Chin MT, Stolz DB, Davis EC, Dichek DA. Transforming growth factor beta signaling in myogenic cells regulates vascular morphogenesis, differentiation, and matrix synthesis. Arterioscler. Thromb. Vasc. Biol. 2012 (Epub ahead of print: Oct 6, 2011).