development of cardiac malformations : The involvement of the
endothelin-1 pathway
Groenendijk, B.C.W.
Citation
Groenendijk, B. C. W. (2006, March 23). Influence of blood flow on shear stress responsive
genes in the development of cardiac malformations : The involvement of the endothelin-1
pathway. Retrieved from https://hdl.handle.net/1887/4346
Version:
Corrected Publisher’s Version
License:
Licence agreement concerning inclusion of doctoral thesis in the
Institutional Repository of the University of Leiden
Chapter 1
1.1
Hemodynamics and Cardiac Development
In this thesis, the role of fluid shear stress is investigated in the chicken embryonic heart, to determine whether through altered gene expression it has an effect on cardiac development. Therefore, background information is provided for fluid dynamics, normal human and chicken heart development, cardiac malformations, cardiac and vitelline blood flow, and the chicken model that we use to generate cardiovascular anomalies.
1.1.1 Fluid
Dynamics
Blood flow is an important epigenetic factor in heart development1-3, as will be described below. Blood flow causes a tangential frictional force on the vessel wall, which is called wall shear stress. Wall shear stress is important, since this is the direct effect of blood flow on the cells. Besides shear stress, pressure is also an important hemodynamic trigger for a biological response. Volumetric fluid flow rate is one of the parameters determining shear stress, as can be demonstrated with the Hagen-Poiseuille law:
3
4
R
Q
π
μ
τ
=
Flow with a Re above approximately 2100 is turbulent. Laminar flow exists when Re is below approximately 2100, but in these cases inertia still exists and laminar vortices may occur. In the embryonic situation, Re of blood is even lower (Re<1), indicating that it is a very viscous flow, where inertia forces become small and blood flow stops when propulsion (heart beat and windkessel effect) terminates4. Flow with a Reynolds number smaller than 1 is also called ‘Stokes flow’. Due to the low Re, embryonic blood will have its maximum velocity shifted to the inner curvature in a vessel. The Fåhraeus-Lindqvist effect (plasma-skimming)5 also needs to be taken into account, which is a decrease in apparent viscosity that occurs when a suspension, such as blood, is made to flow through a tube of smaller diameter (<200μm). In these vessels, due to the particulate nature of blood because of the red blood cells, an almost cell-free layer of plasma is present near the vessel wall with reduced viscosity, and hence shear stress6. The shear stress is expected to be highest in the inner curvature of embryonic vessels6. In adults Re is higher and blood velocity will peak closer to the outer curvature of a vessel. Laminar vortices may also occur, e.g., after valves and bifurcations, resulting in areas with shear gradients with low mean shear and the subsequent formation of atherosclerotic plaques7-9.
In the Hagen-Poiseuille law, the volumetric blood flow stands in the numerator, indicating that an increase in blood flow causes an increase in shear stress. The lumen radius is in the denominator, which signifies that the smaller the lumen the higher the shear stress. The heart has a complex geometry with various diameters, and during development it changes constantly, as will be described below, implying that flow and shear patterns are intricate.
1.1.2 Cardiac
Development
relatively close proximity, and occurs in the human within the first 23 days after fertilisation, in the chicken from approximately HH10 (day 2) to HH16 (day 3)15. In the second phase of looping the outflow tract is brought in front of the atria, which is eventually followed by the wedging of the aorta in between the atrioventricular orifices, which requires among others the remodelling of the inner curvature17. Between 22 and 28 days after conception (human; HH11-HH15 in the chicken), the heart is a single chambered pump. At the end of this time-window the endocardial cushions develop, which begin the separation of the heart into the four chambers17,18. While the endocardial cushions develop to form the atrioventricular and semilunar valves, the atrial (HH19) and ventricular (HH20) septa begin to form. The ventricular septum is completed at HH3419 (day 46 in human). The atrial septum is completely developed at approximately HH33-34 (day 45), except for the foramen ovale, that closes after birth together with the ductus arteriosus and ductus venosus. The septation of the common outflow tract into the aorta and the pulmonary trunk17,18 is finished around HH33-35 (day 51). The pharyngeal arch artery system in chicken embryos develops symmetrical at first with the paired first, second and third pharyngeal arch arteries. However, differences in the diameters of left and right exist. At approximately HH17 the first pair disappears and the fourth pair develops, of which the left will regress around HH28, resulting in an asymmetrical system. Around HH21 the second arch arteries disappear and the 6th develop. Finally, the arch arteries will encompass the two brachiocephalic arteries (left and right 3rd), an aortic arch (right 4th), and the pulmonary arteries and ductus arteriosus (left and right 6th)20,21. Because of the tight regulation and intricacy, these looping and separation processes can easily be impaired, resulting in cardiac defects.
1.1.3 Cardiac
Malformations
defect (ASD)18. This is frequently associated with other cardiac and extracardiac abnormalities. A few other examples of CHD are transposition of the great arteries, coarctation of the aorta, interruption of the aortic arch (type B), cardiac valve anomalies, hypoplastic left heart syndrome, and tetralogy of Fallot. The etiology of congenital heart defects may be genetic, e.g., in relation with Down syndrome, DiGeorge syndrome, Turner XO syndrome, or Marfan syndrome24-27. Environmental factors can also play a role in the etiology, such as hormones28,29, teratogens30,31, and as recently shown in a chicken model, via altered blood flow2, but it is often multifactorial. The heart is functional early in development, and the cardiovascular system is, therefore, exquisitely sensitive to the intra- and extra-embryonic environment, and virtually any disturbance in the environment has cardiovascular consequences21. As mentioned above, the disturbances in environment may also include changes in blood flow, which, during heart development, moves through the cardiovascular system with all its complex loopings, indentations and separations.
1.1.4
Blood Flow in the Chicken Embryo
importance of blood flow in cardiac development has been demonstrated by Hove et al.3. A change in the normal alterations of the intracardiac flow patterns can, therefore, lead to cardiovascular anomalies.
1.1.5 Venous
Clip
Model
Because the intracardiac blood flow pattern is important for normal heart development33, a model was generated in which the right lateral vitelline vein of a chicken embryo is ligated. This is called the venous clip model1. With ligation, blood from the right yolk sac region reroutes immediately via a new vein through the caudal plexus to the left, preventing that the embryo is deprived of blood and oxygen2. The ligation results in altered blood flow patterns through the heart1 and eventually to a range of cardiovascular malformations, such as VSDs, semilunar valve anomalies and pharyngeal arch artery malformations2. Not only morphological defects were demonstrated, hemodynamics were also altered at HH34, showing a decrease in heart rate, and an increase in mean dorsal aortic blood flow35. It is now clear that blood flow is involved in the development of the heart, and that by disturbing it cardiovascular malformations will arise. This led to the question about the mechanism, how cells can sense changes in flow, and how they can respond to this biologically, so that it finally results in an altered cardiac morphology and function.
1.2
Shear Stress and Gene Expression
Endothelial cells comprise the inner layer of the cardiovasculature and are, therefore, the cells that are subjected to blood flow, and hence shear stress. We pose that shear stress is the factor that causes the cardiovascular malformations found in the venous clip model by means of alterations in gene expression. Therefore, some background information about shear sensing, the influence on gene expression, and three shear stress responsive genes is presented.
1.2.1 Shear
Sensing
cells39, and in the early embryonic epithelial cells of Hensen’s node40,41, primary cilia have been shown to act as fluid shear sensors. Primary cilia have also been described to be present on embryonic aortic endothelial cells42, on cultured senescent bovine aortic endothelial cells43, and on cultured human umbilical vein endothelial cells44. These studies demonstrate that primary cilia may very well be fluid shear stress sensors on endothelial and endocardial cells. In chapter 6 we elaborate on this.
1.2.2 Gene
Expression
Upon detecting a change in shear stress in endothelial cells, a signal is sent to the nucleus by means of the activation of protein kinase C (PKC), which in turn activates a second messenger cascade, involving the family of mitogen-activated protein (MAP) kinases, most importantly extracellular signal-regulated kinases (ERK1/2), resulting in gene transcription to be up- or down-regulated (reviewed by Gimbrone et al.45, and Traub and Berk9). Members of the MAP kinases have many potential substrates, including other protein kinases (Raf-1, MEK), transcription factors, enzymes (cPLA2) and cell surface proteins (EGF receptor). Transcription factors, such as c-fos, c-jun, Egr-1, SP1, and NFκB are activated and influence gene regulation. Although transcriptional activation may occur through oxidative stress46, the binding of transcription factors to a shear stress response element (SSRE) in the promoter sequence of a gene is the most probable mechanism for shear-mediated transcription. Transcription factors have been shown to bind to the SSRE of certain genes and regulate transcription, e.g., NFκB in platelet-derived growth factor B (PDGF-B) regulation47. SSREs can be positive or negative regulators of transcription. The TRE (AP-1) site for instance, binds the transcription factors fos and jun and inhibits transcription48, whereas the Egr-1/SP1 binding site stimulates transcription upon activation49. Changes in gene expression by shear stress have been demonstrated by several independent groups46,50,51. In this thesis the stress is put on three genes: endothelin-1 (ET-1), lung Krüppel-like factor (LKLF/KLF2), and endothelial nitric oxide synthase (eNOS/NOS-3).
Endothelin-1
receptors. Hitherto, the receptors have been grouped into two main classes: the endothelin-A (ETendothelin-A) and the endothelin-B (ETB) receptors. Besides being a vasoconstrictor, mediated by the action of ET-1 on both ETA and ETB receptors on smooth muscle cells53, ET-1 also has vasodilator properties mediated by nitric oxide (NO) and prostacyclin (PGI2) release through activation of the ETB receptor located in the endothelium54-57. Furthermore, ET-1 is a growth factor, involved in, e.g., the proliferation of fibroblasts and smooth muscle cells through the ETA receptor58-61, the proliferation of endothelial cells through the ETB receptor62,63, and hypertrophy of various cell types, including cardiomyocytes64,65. It is also implicated in collagen production66. In addition, it is involved, via the ETA receptor, in the closure of the ductus arteriosus after birth67.
What is most interesting, is that endothelin-1 knockout mice (Edn-1-/-), display similar cardiovascular defects as chicken embryos in the venous clip model2,68. Heterozygous mice (Edn+/-) show elevated blood pressure69. Likewise, knockout mice of genes from the ET-1 pathway, Ece-1-/- and Eta-/-, show similar cardiovascular malformations70,71. ET-1 mRNA and protein production are regulated by wall shear stress. In vitro studies have demonstrated that ET-1 gene expression, and protein levels are increased by low steady laminar shear stress levels (≤ 6 dyne/cm2)72,73 and decreased by high steady laminar shear (≥ 8 dyne/cm2)51,74-76. In addition, studies have shown that ET-1 is increased for up to one hour by high shear, followed by a chronic down-regulation50,77,78.
Lung Krüppel-Like Factor
Lung Krüppel-like factor is a member of the SP/XKLF family of Cys2/His2 zinc-finger transcription factors79,80. Knockout mice of KLF2 die between embryonic day 12.5 and 14.5 due to severe hemorrhages in the embryonic cardiac outflow tract region and in the abdomen81,82. The hemorrhaging is caused by rupturing of the vessels due to abnormal thinning of the tunica media and concomitant instability of the vessel wall81, demonstrating that KLF2 in endothelial cells regulates the assembly of the vascular tunica media and concomitant vessel wall stabilisation during mammalian embryogenesis. The embryos are also retarded in growth, and show craniofacial abnormalities and signs of anaemia82. The latter can be explained by the fact that KLF2 is essential for primitive erythropoiesis and that it regulates the β-like globin genes83. More important for our study is that KLF2 is up-regulated by increased shear stress in human umbilical vein endothelial cells46. In addition,
NOS-3 induction84. That KLF2 can induce NOS-3 expression and enzymatic activity, had already been shown by SenBanerjee et al.85, who also demonstrated that KLF2 regulates endothelial activation in response to proinflammatory stimuli. However, the latter is not of importance in early embryos, as the immune system has not yet been developed. Recently, Dekker et al.86 have confirmed that KLF2 induces NOS-3 expression, and also demonstrated that KLF2 decreases ET-1 expression. This identifies KLF2 as a molecular switch between the production of nitric oxide (NO) and ET-1 at sites of high shear stress.
Endothelial Nitric Oxide Synthase
Endothelial nitric oxide synthase is the major NOS isoform of the endothelium. It is the functional counterpart of ET-1, as it catalyses the conversion of L-arginine and molecular oxygen to L-citrullin and NO87, the latter being a vasodilator. Therefore, endothelial NO together with ET-1 are physiologically important for maintaining vascular homeostasis (reviewed by Lavallée et al.88).
Knockout mice for Nos-3 are hypertensive and lack NO-mediated, endothelium-dependent vasodilation, demonstrating that endothelial NO is an important systemic vasodilator89-91. However, NO is not only implicated in vasodilation, it also suppresses smooth muscle cell proliferation92,93, and has a negative inotropic effect94. Nos-3-deficient mice, furthermore, show mild pulmonary hypertension and hyper-responsiveness to hypoxia95,96. In addition, these mice were reported to display bicuspid aortic valves, heart failure, and atrial and ventricular septal defects97,98, implying the importance of NOS-3 in cardiac development. In contrast to the reported hypertension in Nos-3-/- mice, transgenic mice expressing large amounts of Nos-3 targeted to the vessel wall by the ET-1 promoter are hypotensive, and show a reduced vascular sensitivity to NO99.
Interestingly, the NOS-3 promoter contains a SSRE (GAGACC), besides numerous other binding sites for transcription factors100,101. In vitro studies have shown that steady laminar shear stress increases the expression of NOS-3 mRNA and NO formation75,102,103.
1.3
Setting of this Thesis
to cardiac (mal)development in chicken embryos, was investigated by our colleagues at the Department of Obstetrics and Gynaecology of the Erasmus Medical Centre, Rotterdam105-107. At the Delft University of Technology, our colleagues at the Department of Aero- and Hydrodynamics produced a computational fluid dynamics program with which the shear stress distribution can be mapped in the embryonic chicken heart108. By means of micro-Particle Image Velocimetry blood flow can be monitored very precisely and the wall shear stress can be calculated109.
1.4 Chapter
Outline
Chapter 2 describes the expression patterns of three shear stress responsive genes during chicken cardiovascular development. We show that the gene patterns can be linked to areas of high shear stress in the developing heart.
In Chapter 3 the expression patterns and levels of the three shear responsive genes are investigated after venous clip. Here, we demonstrate that changes in flow and shear stress indeed affect gene expression in specific ways.
Chapter 4 presents a study in which we infused endothelin and endothelin receptor antagonists into the blood stream of stage HH18 chicken embryos, to investigate the effects of ET-1 on ventricular function and cardiovascular morphology at HH24 and HH35 respectively. It is shown that the endothelin-1 pathway is involved in the disturbed diastolic ventricular function and in morphological anomalies in the venous clip model.
In Chapter 5 we demonstrate that infusion of endothelin-1 and endothelin receptor antagonists have direct effects on hemodynamics in vitelline veins measured with micro-Particle Image Velocimetry. With Doppler measurements we were not able to detect alterations in the embryonic hemodynamics.
In Chapter 6 a study to a possible shear sensor is shown. Monocilia, discovered on the endocardium, may be shear sensors and are present in the embryonic chicken heart at areas of low shear stress.
1.5 References
1. Hogers B, DeRuiter MC, Gittenberger-de Groot AC, Poelmann RE. Unilateral vitelline vein ligation alters intracardiac blood flow patterns and morphogenesis in the chick embryo. Circ Res. 1997;80:473-481.
2. Hogers B, DeRuiter MC, Gittenberger-de Groot AC, Poelmann RE. Extraembryonic venous obstructions lead to cardiovascular malformations and can be embryolethal. Cardiovasc Res. 1999;41:87-99.
3. Hove JR, Koster RW, Forouhar AS, Acevedo-Bolton G, Fraser SE, Gharib M. Intracardiac fluid forces are an essential epigenetic factor for embryonic cardiogenesis. Nature. 2003;421:172-177.
4. Purcell EM. Life at low Reynolds number. Am J Physiol. 1977;45:3-9.
5. Goldsmith HL, Cokelet GR, Gaehtgens P. Robin Fahraeus: evolution of his concepts in cardiovascular physiology. Am J Physiol. 1989;257:H1005-H1015.
6. Wang CY, Bassingthwaighte JB. Blood flow in small curved tubes. J Biomech Eng. 2003;125:910-913.
7. Moore JE, Xu C, Glagov S, Zarins CK, Ku DN. Fluid wall shear stress measurements in a model of the human abdominal aorta: oscillatory behavior and relationship to atherosclerosis. Atherosclerosis. 1994;110:225-240. 8. Topper JN, Gimbrone MA, Jr. Blood flow and vascular gene expression: fluid shear stress as a modulator of
endothelial phenotype. Mol Med Today. 1999;5:40-46.
9. Traub O, Berk BC. Laminar shear stress: mechanisms by which endothelial cells transduce an atheroprotective force. Arterioscler Thromb Vasc Biol. 1998;18:677-685.
10. Hamburger V, Hamilton HL. A series of normal stages in the development of the chick embryo. J Morphol. 1951;88:49-92.
11. DeRuiter MC, Poelmann RE, VanderPlas-de Vries I, Mentink MMT, Gittenberger-de Groot AC. The development of the myocardium and endocardium in mouse embryos. Fusion of two heart tubes? Anat Embryol. 1992;185:461-473.
12. Markwald RR, Fitzharris TP, Manasek FJ. Structural development of endocardial cushions. Am J Anat. 1977;148:85-120.
13. Männer J. On rotation, torsion, lateralization, and handedness of the embryonic heart loop: new insights from a simulation model for the heart loop of chick embryos. Anat Rec A Discov Mol Cell Evol Biol. 2004;278:481-492. 14. Gittenberger-de Groot AC, Bartelings MM, DeRuiter MC, Poelmann RE. Normal cardiac development. Chapter
1: 1-14. In: Wladimiroff JW, Pilu G, eds. Ultrasound and the fetal heart. The Parthenon Publishing Group, New York – London, 1996.
15. Männer J. Cardiac looping in the chick embryo: a morphological review with special reference to terminological and biomechanical aspects of the looping process. Anat Rec. 2000;259:248-262.
16. Olson EN, Srivastava D. Molecular pathways controlling heart development. Science. 1996;272:671-676.
17. Gittenberger-de Groot AC, DeRuiter MC, Bartelings MM, Poelmann RE. Embryology of congenital heart disease. Chapter 7: 2.1-2.10. In: Crawford MH, DiMarco JP, eds. Cardiology. Mosby International Limited, London, 2001. 18. Gittenberger-de Groot AC, Bartelings MM, DeRuiter MC, Poelmann RE. Basics of cardiac development for the
understanding of congenital heart malformations. Pediatr Res. 2005;57:169-176.
19. Hogers B, Gittenberger-de-Groot AC, DeRuiter MC, Mentink MMT, Poelmann RE. Cardiac inflow malformations are more lethal and precede cardiac outflow malformations. Chick embryonic venous clip model. Chapter 5: 79-100. In: Hogers B, ed. The role of blood flow in normal and abnormal heart development. Ponsen & Looijen BV, Wageningen, 1998.
21. Kirby ML, Waldo KL. Neural crest and cardiovascular patterning. Circ Res. 1995;77:211-215.
22. Morris CD. Epidemiology of congenital heart disease. Chapter 7: 1.1-1.8. In: Crawford MH, DiMarco JP, eds. Cardiology. Mosby International Limited, London, 2001.
23. Vriend JWJ, Van der Velde ET, Mulder BJM. Aangeboren hartafwijkingen in Nederland. Landelijke registratie en DNA-bank van aangeboren hartafwijkingen: het CONCOR-project. Chapter 5: 105-120. In: Van Leest LATM, Koek HL, Van Trijp MJCA, Van Dis SJ, Peters RJG, Bots ML, Verschuren WMM, eds. Hart- en vaatziekten in Nederland 2005. Nederlandse Hartstichting, Den Haag, 2005.
24. Barlow GM, Chen X-N, Lyons GE, Kurnit DM, Celle L, Spinner NB, Zackai EH, Mjaatvedt CH, VanRiper AJ, Vekemans MJ, Korenberg JR. Down syndrome congenital heart disease: a narrowed region and a candidate gene. Genet Med. 2001;3:91-101.
25. Baldini A. DiGeorge syndrome: an update. Curr Opin Cardiol. 2004;19:201-204. 26. Lin AE. The heart of Turner syndrome: small matters. Teratology. 2002;66:63-64.
27. Collod-Béroud G, Boileau C. Marfan syndrome in the third Millennium. Eur J Hum Genet. 2002;10:673-681. 28. Boot MJ, Steegers-Theunissen RP, Poelmann RE, van Iperen L, Gittenberger-de Groot AC. Cardiac outflow tract
malformations in chick embryos exposed to homocysteine. Cardiovasc Res. 2004;64:365-373.
29. Kapusta L, Haagmans ML, Steegers EA, Cuypers MH, Blom HJ, Eskes TK. Congenital heart defects and maternal derangement of homocysteine metabolism. J Pediatr. 1999;135:773-774.
30. Broekhuizen MLA, Wladimiroff JW, Tibboel D, Poelmann RE, Wenink ACG, Gittenberger-de Groot AC. Induction of cardiac anomalies with all-trans retinoic acid in the chick embryo. Cardiol Young. 1992;2:311-317. 31. Bouman HGA, Broekhuizen MLA, Baasten AMJ, Gittenberger-de Groot AC, Wenink ACG. Stereological study of
stage 34 chicken hearts with looping disturbances after retinoic acid treatment: disturbed growth of myocardium and atrioventricular cushion tissue. Anat Rec. 1997;248:242-250.
32. Fujii S, Hirota A, Kamino K. Optical indications of pace-maker potential and rhythm generation in early embryonic chick heart. J Physiol. 1981;312:253-263.
33. Hogers B, DeRuiter MC, Baasten AMJ, Gittenberger-de Groot AC, Poelmann RE. Intracardiac blood flow patterns related to the yolk sac circulation of the chick embryo. Circ Res. 1995;76:871-877.
34. Manasek FJ, Monroe RG. Early cardiac morphogenesis is independent of function. Dev Biol. 1972;27:584-588. 35. Broekhuizen MLA, Hogers B, DeRuiter MC, Poelmann RE, Gittenberger-de Groot AC, Wladimiroff JW. Altered
hemodynamics in chick embryos after extraembryonic venous obstruction. Ultrasound Obstet Gynecol. 1999;13:437-445.
36. Resnick N, Yahav H, Shay-Salit A, Shushy M, Schubert S, Zilberman LC, Wofovitz E. Fluid shear stress and the vascular endothelium: for better and for worse. Prog Biophys Mol Biol. 2003;81:177-199.
37. Florian JA, Kosky JR, Ainslie K, Pang ZY, Dull RO, Tarbell JM. Heparan sulfate proteoglycan is a mechanosensor on endothelial cells. Circ Res. 2003;93:E136-E142.
38. Mochizuki S, Vink H, Hiramatsu O, Kajita T, Shigeto F, Spaan JA, Kajiya F. Role of hyaluronic acid glycosaminoglycans in shear-induced endothelium-derived nitric oxide release. Am J Physiol Heart Circ Physiol. 2003;285:H722-H726.
39. Praetorius HA, Spring KR. Bending the MDCK cell primary cilium increases intracellular calcium. Journal of Membrane Biology. 2001;184:71-79.
40. McGrath J, Somlo S, Makova S, Tian X, Brueckner M. Two populations of node monocilia initiate left-right asymmetry in the mouse. Cell. 2003;114:61-73.
41. Yost HJ. Left-right asymmetry: Nodal cilia make and catch a wave. Current Biology. 2003;13:R808-R809.
43. Briffeuil P, Thibaut-Vercruyssen R, Ronveaux-Dupal MF. Ciliation of bovine aortic endothelial cells in culture. Atherosclerosis. 1994;106:75-81.
44. Iomini C, Tejada K, Mo W, Vaananen H, Piperno G. Primary cilia of human endothelial cells disassemble under laminar shear stress. J Cell Biol. 2004;164:811-817.
45. Gimbrone MA, Jr., Topper JN, Nagel T, Anderson KR, Garcia-Cardena G. Endothelial dysfunction, hemodynamic forces, and atherogenesis. Ann N Y Acad Sci. 2000;902:230-239.
46. Dekker RJ, Van Soest S, Fontijn RD, Salamanca S, de Groot PG, VanBavel E, Pannekoek H, Horrevoets AJG. Prolonged fluid shear stress induces a distinct set of endothelial cell genes, most specifically lung Krüppel-like factor (KLF2). Blood. 2002;100:1689-1698.
47. Khachigian LM, Resnick N, Gimbrone MA, Collins T. Nuclear factor-kB interacts functionally with the platelet-derived growth factor B-chain shear-stress response element in vascular endothelial cells exposed to fluid shear stress. J Clin Invest. 1995;96:1169-1175.
48. Shyy JY, Lin MC, Han J, Lu Y, Petrime M, Chien S. The cis-acting phorbol ester "12-O-tetradecanoylphorbol 13-acetate"-responsive element is involved in shear stress-induced monocyte chemotactic protein 1 gene expression. Proc Natl Acad Sci U S A. 1995;92:8069-8073.
49. Resnick N, Yahav H, Khachigian LM, Collins T, Anderson KR, Dewey FC, Gimbrone MA, Jr. Endothelial gene regulation by laminar shear stress. Adv Exp Med Biol. 1997;430:155-164.
50. Malek AM, Izumo S. Control of endothelial cell gene expression by flow. J Biomech. 1995;28:1515-1528.
51. McCormick SM, Eskin SG, McIntire LV, Teng CL, Lu CM, Russell CG, Chittur KK. DNA microarray reveals changes in gene expression of shear stressed human umbilical vein endothelial cells. Proc Natl Acad Sci U S A. 2001;98:8955-8960.
52. Yanagisawa M, Kurihara H, Kimura S, Tomobe Y, Kobayashi M, Mitsui Y, Yazaki Y, Goto K, Masaki T. A novel potent vasoconstrictor peptide produced by vascular endothelial cells. Nature. 1988;332411:411-415.
53. Haynes WG, Strachan FE, Webb DJ. Endothelin ETA and ETB receptors cause vasoconstriction of human resistance and capacitance vessels in vivo. Circulation. 1995;92:357-363.
54. de Nucci G, Thomas R, D'Orleans-Juste P, Antunes E, Walder C, Warner TD, Vane JR. Pressor effects of circulating endothelin are limited by its removal in the pulmonary circulation and by the release of prostacyclin and endothelium-derived relaxing factor. Proc Natl Acad Sci U S A. 1988;85:9797-9800.
55. Namiki A, Hirata Y, Ishikawa M, Moroi M, Aikawa J, Machii K. Endothelin-1- and endothelin-3-induced vasorelaxation via common generation of endothelium-derived nitric oxide. Life Sci. 1992;50:677-682.
56. Tsukahara H, Ende H, Magazine HI, Bahou WF, Goligorsky MS. Molecular and functional characterization of the non-isopeptide-selective ETB receptor in endothelial cells. Receptor coupling to nitric oxide synthase. J Biol Chem. 1994;269:21778-21785.
57. Verhaar MC, Strachan FE, Newby DE, Cruden NL, Koomans HA, Rabelink TJ, Webb DJ. Endothelin-A receptor antagonist-mediated vasodilatation is attenuated by inhibition of nitric oxide synthesis and by endothelin-B receptor blockade. Circulation. 1998;97:752-756.
58. Fujisaki H, Ito H, Hirata Y, Tanaka M, Hata M, Lin MH, Adachi S, Akimoto H, Marumo F, Hiroe M. Natriuretic Peptides Inhibit Angiotensin-Ii-Induced Proliferation of Rat Cardiac Fibroblasts by Blocking Endothelin-1 Gene-Expression. Journal of Clinical Investigation. 1995;96:1059-1065.
59. Yang Z, Krasnici N, Luscher TF. Endothelin-1 potentiates human smooth muscle cell growth to PDGF: effects of ETA and ETB receptor blockade. Circulation. 1999;100:5-8.
61. Zhang YM, Wang KQ, Zhou GM, Zuo J, Ge JB. Endothelin-1 promoted proliferation of vascular smooth muscle cell through pathway of extracellular signal-regulated kinase and cyclin D1. Acta Pharmacol Sin. 2003;24:563-568. 62. Morbidelli L, Orlando C, Maggi CA, Ledda F, Ziche M. Proliferation and migration of endothelial cells is
promoted by endothelins via activation of ETB receptors. Am J Physiol. 1995;269:H686-H695.
63. Dong F, Zhang X, Wold LE, Ren Q, Zhang Z, Ren J. Endothelin-1 enhances oxidative stress, cell proliferation and reduces apoptosis in human umbilical vein endothelial cells: role of ETB receptor, NADPH oxidase and caveolin-1. Br J Pharmacol. 2005;145:323-333.
64. Shubeita HE, McDonough PM, Harris AN, Knowlton KU, Glembotski CC, Brown JH, Chien KR. Endothelin induction of inositol phospholipid hydrolysis, sarcomere assembly, and cardiac gene expression in ventricular myocytes. A paracrine mechanism for myocardial cell hypertrophy. J Biol Chem. 1990;265:20555-20562.
65. Yamazaki T, Komuro I, Kudoh S, Zou Y, Shiojima I, Hiroi Y, Mizuno T, Maemura K, Kurihara H, Aikawa R, Takano H, Yazaki Y. Endothelin-1 is involved in mechanical stress-induced cardiomyocyte hypertrophy. J Biol Chem. 1996;271:3221-3228.
66. Guarda E, Katwa LC, Myers PR, Tyagi SC, Weber KT. Effects of endothelins on collagen turnover in cardiac fibroblasts. Cardiovasc Res. 1993;27:2130-2134.
67. Taniguchi T, Muramatsu I. Pharmacological knockout of endothelin ET(A) receptors. Life Sci. 2003;74:405-409. 68. Kurihara Y, Kurihara H, Oda H, Maemura K, Nagai R, Ishikawa T, Yazaki Y. Aortic arch malformations and
ventricular septal defect in mice deficient in endothelin-1. J Clin Invest. 1995;96:293-300.
69. Kurihara Y, Kurihara H, Suzuki H, Kodama T, Maemura K, Nagai R, Oda H, Kuwaki T, Cao WH, Kamada N, Jishage K, Ouchi Y, Azuma S, Toyoda Y, Ishikawa T, Kumada M, Yazaki Y. Elevated blood pressure and craniofacial abnormalities in mice deficient in endothelin -1. Nature. 1994;368:703-710.
70. Yanagisawa H, Yanagisawa M, Kapur RP, Richardson JA, Williams SC, Clouthier DE, de Wit D, Emoto N, Hammer RE. Dual genetic pathways of endothelin-mediated intercellular signaling revealed by targeted disruption of endothelin converting enzyme-1 gene. Development. 1998;125:825-836.
71. Clouthier DE, Hosoda K, Richardson JA, Williams SC, Yanagisawa H, Kuwaki T, Kumada M, Hammer RE, Yanagisawa M. Cranial and cardiac neural crest defects in endothelin-A receptor-deficient mice. Development. 1998;125:813-824.
72. Yoshizumi M, Kurihara H, Sugiyama T, Takaku F, Yanagisawa M, Masaki T, Yasaki Y. Hemodynamic shear stress stimulates endothelin production by cultured endothelial cells. Biochem Biophys Res Commun. 1989;161:859-864.
73. Mattart M, Mazzolai L, Chambaz C, Hayoz D, Brunner HR, Silacci P. ET-1 and NOSIII gene expression regulation by plaque-free and plaque-prone hemodynamic conditions. Biorheology. 2003;40:289-297.
74. Sharefkin JB, Diamond SL, Eskin SG, McIntire LV, Dieffenbach CW. Fluid flow decreases preproendothelin mRNA levels and suppresses endothelin-1 peptide release in cultured human endothelial cells. J Vasc Surg. 1991;14:1-9.
75. Noris M, Morigi M, Donadelli R, Aiello S, Foppolo M, Todeschini M, Orisio S, Remuzzi G, Remuzzi A. Nitric oxide synthesis by cultured endothelial cells is modulated by flow conditions. Circ Res. 1995;76:536-543. 76. Masatsugu K, Itoh H, Chun TH, Ogawa Y, Tamura N, Yamashita J, Doi K, Inoue M, Fukunaga Y, Sawada N,
Saito T, Korenaga R, Ando J, Nakao K. Physiologic shear stress suppresses endothelin-converting enzyme-1 expression in vascular endothelial cells. J Cardiovasc Pharmacol. 1998;31(Suppl 1):S42-S45.
77. Malek AM, Izumo S. Physiological fluid shear stress causes downregulation of endothelin-1 mRNA in bovine aortic endothelium. Am J Physiol. 1992;263 (2Pt1):C389-C396.
79. Philipsen S, Suske G. A tale of three fingers: the family of mammalian Sp/XKLF transcription factors. Nucleic Acids Res. 1999;27:2991-3000.
80. Bieker JJ. Kruppel-like factors: three fingers in many pies. J Biol Chem. 2001;276:34355-34358.
81. Kuo CT, Veselits ML, Barton KP, Lu MM, Clendenin C, Leiden JM. The LKLF transcription factor is required for normal tunica media formation and blood vessel stabilization during murine embryogenesis. Genes Dev. 1997;11:2996-3006.
82. Wani MA, Means RT, Jr., Lingrel JB. Loss of LKLF function results in embryonic lethality in mice. Transgenic Res. 1998;7:229-238.
83. Basu P, Morris P, Wani MA, Lingrel JB, Lloyd JA. KLF2 is required for normal epsilon- and beta hl-globin expression. Blood Cells Molecules and Diseases. 2005;34:76.
84. Huddleson JP, Ahmad N, Srinivasan S, Lingrel JB. Induction of KLF2 by fluid shear stress requires a novel promoter element activated by a phosphatidylinositol 3-kinase-dependent chromatin-remodeling pathway. J Biol Chem. 2005;280:23371-23379.
85. SenBanerjee S, Lin Z, Atkins GB, Greif DM, Rao RM, Kumar A, Feinberg MW, Chen Z, Simon DI, Luscinskas FW, Michel TM, Gimbrone MA, Jr., Garcia-Cardena G, Jain MK. KLF2 Is a novel transcriptional regulator of endothelial proinflammatory activation. J Exp Med. 2004;199:1305-1315.
86. Dekker RJ, van Thienen JV, Elderkamp YW, Seppen J, de Vries CJM, Biessen EA, van Berkel TJ, Pannekoek H, Horrevoets AJG. Endothelial KLF2 links local arterial shear stress levels to the expression of vascular-tone regulating genes. Am J Pathol. 2005;167:609-618.
87. Marsden PA, Schappert KT, Chen HS, Flowers M, Sundell CL, Wilcox JN, Lamas S, Michel T. Molecular cloning and characterization of human endothelial nitric oxide synthase. FEBS Lett. 1992;307:287-293.
88. Lavallée M, Takamura M, Parent R, Thorin E. Crosstalk between endothelin and nitric oxide in the control of vascular tone. Heart Fail Rev. 2001;6:265-276.
89. Huang PL, Huang Z, Mashimo H, Bloch KD, Moskowitz MA, Bevan JA, Fishman MC. Hypertension in mice lacking the gene for endothelial nitric oxide synthase. Nature. 1995;377:239-242.
90. Shesely EG, Maeda N, Kim HS, Desai KM, Krege JH, Laubach VE, Sherman PA, Sessa WC, Smithies O. Elevated blood pressures in mice lacking endothelial nitric oxide synthase. Proc Natl Acad Sci U S A. 1996;93:13176-13181. 91. Gödecke A, Decking UK, Ding Z, Hirchenhain J, Bidmon HJ, Godecke S, Schrader J. Coronary hemodynamics in
endothelial NO synthase knockout mice. Circ Res. 1998;82:186-194.
92. Moroi M, Zhang L, Yasuda T, Virmani R, Gold HK, Fishman MC. Interaction of genetic deficiency of endothelial nitric oxide, gender, and pregnancy in vascular response to injury in mice. J Clin Invest. 1998;101:1225-1232. 93. Rudic RD, Shesely EG, Maeda N, Smithies O, Segal SS, Sessa WC. Direct evidence for the importance of
endothelium-derived nitric oxide in vascular remodeling. J Clin Invest. 1998;101:731-736.
94. Gödecke A, Heinicke T, Kamkin A, Kiseleva I, Strasser RH, Decking UK, Stumpe T, Isenberg G, Schrader J. Inotropic response to beta-adrenergic receptor stimulation and anti-adrenergic effect of ACh in endothelial NO synthase-deficient mouse hearts. J Physiol. 2001;532:195-204.
95. Fagan KA, Fouty BW, Tyler RC, Morris KG, Jr., Hepler LK, Sato K, LeCras TD, Abman SH, Weinberger HD, Huang PL, McMurtry IF, Rodman DM. The pulmonary circulation of homozygous or heterozygous eNOS-null mice is hyperresponsive to mild hypoxia. J Clin Invest. 1999;103:291-299.
96. Steudel W, Ichinose F, Huang PL, Hurford WE, Jones RC, Bevan JA, Fishman MC, Zapol WM. Pulmonary vasoconstriction and hypertension in mice with targeted disruption of the endothelial nitric oxide synthase (NOS 3) gene. Circ Res. 1997;81:34-41.
98. Feng Q, Song W, Lu X, Hamilton JA, Lei M, Peng T, Yee SP. Development of heart failure and congenital septal defects in mice lacking endothelial nitric oxide synthase. Circulation. 2002;106:873-879.
99. Ohashi Y, Kawashima S, Hirata K, Yamashita T, Ishida T, Inoue N, Sakoda T, Kurihara H, Yazaki Y, Yokoyama M. Hypotension and reduced nitric oxide-elicited vasorelaxation in transgenic mice overexpressing endothelial nitric oxide synthase. J Clin Invest. 1998;102:2061-2071.
100. Marsden PA, Heng HH, Scherer SW, Stewart RJ, Hall AV, Shi XM, Tsui LC, Schappert KT. Structure and chromosomal localization of the human constitutive endothelial nitric oxide synthase gene. J Biol Chem. 1993;268:17478-17488.
101. Venema Rc, Nishida K, Alexander RW, Harrison DG, Murphy TJ. Organization of the bovine gene encoding the endothelial nitric oxide synthase. Biochim Biophys Acta. 1994;1218:413-420.
102. Nishida K, Harrison DG, Navas JP, Fisher AA, Dockery SP, Uematsu M, Nerem RM, Alexander RW, Murphy TJ. Molecular cloning and characterization of the constitutive bovine aortic endothelial cell nitric oxide synthase. J Clin Invest. 1992;90:2092-2096.
103. Ziegler T, Silacci P, Harrison VJ, Hayoz D. Nitric oxide synthase expression in endothelial cells exposed to mechanical forces. Hypertension. 1998;32:351-355.
104. Poelmann RE, Gittenberger-de Groot AC, Groenendijk BCW, Hierck BP, Hogers B, Nieuwstadt FTM, Pourquie MJBM, Steendijk P, Stekelenburg-de Vos S, Ursem NTC, Wladimiroff JW. Shear stress in the developing cardiovascular system. Chapter 42: 169-173. In: Artman M, Benson DW, Srivastava D, Nakazawa M, eds. Cardiovascular development and congenital malformations: molecular and genetic mechanisms. Blackwell Futura, Massachusetts, 2005.
105. Stekelenburg-de Vos S, Ursem NTC, Hop WCJ, Wladimiroff JW, Gittenberger-de Groot AC, Poelmann RE. Acutely altered hemodynamics following venous obstruction in the early chick embryo. J Exp Biol. 2003;206:1051-1057.
106. Ursem NTC, Stekelenburg-de Vos S, Wladimiroff JW, Poelmann RE, Gittenberger-de Groot AC, Hu N, Clark EB. Ventricular diastolic filling characteristics in stage-24 chick embryos after extra-embryonic venous obstruction. J Exp Biol. 2004;207:1487-1490.
107. Stekelenburg-de Vos S, Steendijk P, Ursem NT, Wladimiroff JW, Delfos R, Poelmann RE. Systolic and Diastolic Ventricular Function Assessed by Pressure-Volume Loops in the Stage 21 Venous Clipped Chick Embryo. Pediatr Res. 2005;57:16-21.
108. Groenendijk BCW, Hierck BP, Vrolijk J, Baiker M, Pourquie MJBM, Gittenberger-de-Groot AC, Poelmann RE. Changes in shear stress-related gene expression after experimentally altered venous return in the chicken embryo. Circ Res. 2005;96:1291-1298.
