Ординатура / Офтальмология / Английские материалы / Retinal and Choroidal Angiogenesis_Penn_2008

precise A class receptor(s) that function in the tumor endothelium remains undefined. Since EphA2 is a major class A Eph receptor expressed in the adult endothelium (Bowen and Chen, unpublished results), EphA2-deficient

mice were studied further. Gene disruption of EphA2 did not affect survival or major embryonic developmental events,51,62,63 reflecting the fact that the

EphA2 receptor is not expressed in the embryonic vasculature.42 However, EphA2 receptors are expressed in adult endothelium and tumor neovasculature. Accordingly, EphA2-null endothelial cells failed to undergo cell migration and vascular assembly both in vitro and in vivo.51 In addition, tumor growth, angiogenesis, and metastasis were significantly inhibited in EphA2-deficient receipt mice.64 These data suggest that host EphA2 RTK function is required in the tumor microenvironment for tumor angiogenesis and metastatic progression.

In addition to the function of EphA receptors in tumor neovascularization, ephrin-B2 expression has been observed in tumor arterioles infiltrating transplanted Lewis lung carcinomas and B16 melanomas in mice,33 suggesting a role in tumor angiogenesis. However, modulation of ephrin-B2 or EphB4 signaling separately has pleiotropic effects on tumor progression. For example, overexpression of ephrin-B2 in human colorectal cell increases tumor vessel density but suppresses tumor growth.65 Also, expression of a cytoplasmic truncation mutant of EphB4 in breast cancer cells promotes tumor angiogenesis and tumor growth.66 Nevertheless, blocking of bidirectional signaling between ephrin-B2 and EphB4 by a soluble EphB4 monomer effectively inhibited A375 melanoma growth and tumor angiogenesis.67 Taken together, these data suggest that ephrin-B2 or EphB4 could be targets for inhibition of tumor neovascularization, but blocking of bidirectional signaling would be a prerequisite of developing these targets for therapeutic intervention.

The precise mechanism of how Eph/ephrin signaling regulates tumor angiogenesis is not known. However, from the available data it is conceivable that Eph/ephrin-dependent tumor neovascularization is mediated by the interplay of Eph receptors and ephrin ligands expressed by tumor cells and endothelial cells. During the initial phase of the angiogenic response, activation of EphA2 receptors on vascular smooth muscle induces retraction of perivascular supporting cells via inhibition of the Rac1 GTPase/PAK pathway,68 allowing endothelial cells to respond to angiogenic cues. In contrast, the signaling of EphA2 receptors on endothelial cells stimulates PI3 kinase-dependent Rac1 GTPase activation, promoting endothelial cell migration and vessel assembly.51 Ephrins expressed on the tumor cells may function as contact-dependent organizing molecules to guide incoming vessels that express EphA2 receptor. Alternatively, angiogenic factors such as VEGF or TNF-α in the tumor microenvironment may induce the expression

and/or activation of ephrins in endothelial cells, as suggested in studies from cultured endothelial cells.40 The ephrins may then interact with Eph receptors on adjacent endothelial cells to promote endothelial cell sprouting, migration, and capillary tube formation through bidirectional signaling. Furthermore, the interactions between tumor cells and host blood vessels may provide a mechanism for the intravasation of tumor cells into the blood stream, facilitating tumor metastasis. Regardless of the mechanism, Eph receptors and ephrin ligands are attractive candidates for tumor prognostic markers and potential targets for therapeutic intervention in cancer.

6.ROLE OF EPH/EPHRIN IN RETINAL ANGIOGENESIS

Retinopathy of prematurity, as well as diabetic retinopathy, neovascular glaucoma, and age-related macular degeneration, involves abnormal ocular angiogenesis. Large numbers of new blood vessels lead to blindness by disturbing the refractive surface of the ocular epithelium.69 Expression of ephrin-B2 and relatively lower levels of EphB4 has been observed in human retinal endothelial cells.70 In addition, reverse signaling through B class ephrins can affect retinal endothelial cells in culture, a fact made evident when treatment with the soluble EphB4-Fc receptor ectodomain induced retinal endothelial proliferation and migration via activation of PI3K, nitric oxide synthase, and ERK1/2.70 These in vitro cell culture data are consistent with recent findings in human retina with pathological angiogenesis.71 Ephrin-B2 was expressed in a significant percentage of the endothelial cells of fibroproliferative membranes that were obtained from patients with proliferative diabetic retinopathy (65%) and retinopathy of prematurity (25%). While the expression of EphB4 receptor was not detected in these clinical samples, EphB2 and EphB3 receptors were synthesized in these proliferative membranes. Given their functions in developmental angiogenesis, these data suggest that B class Eph RTKs and ephrins could contribute to pathogenic neovascularization in the retina.

Recently, new functional data demonstrate that A class Eph RTKs and ephrins participate in retinopathy of prematurity.72 EphA2 is expressed in human retinal endothelial cells, and treatment with soluble EphA2-Fc, which inhibits activation of endogenous EphA RTKs in response to ephrin-A stimulation, reduces retinal neovascularization in a rat retinopathy of prematurity model. Inhibition of disease severity likely targets EphA2expressing retinal endothelium, since soluble EphA-Fc receptors have been

shown to inhibit ephrin-induced migration and sprouting of endothelial cells in culture.39,40 In addition, soluble EphA receptors may also block the effects

fellowship 0120147B and Department of Defense postdoctoral fellowship DAMD17-03-1-0379 to D. B.-S.; and NIH grant EY07533 and the Lew R. Wasserman Merit Award for Research to Prevent Blindness to J. P.

REFERENCES

1.P. Lee, C. C. Wang, and A. P. Adamis, Ocular neovascularization: An epidemiological review, Surv. Ophthalmol. 43, 245-269 (1998).

2.M. A. Speicher et al., Pharmacologic therapy for diabetic retinopathy, Expert Opin. Emerg. Drugs 8, 239-250 (2003).

3.G. D. Yancopoulos et al., Vascular-specific growth factors and blood vessel formation, Nature 407, 242-248 (2000).

4.A. W. Griffioen and G. Molema, Angiogenesis: potentials for pharmacologic intervention in the treatment of cancer, cardiovascular diseases, and chronic inflammation, Pharmacol. Rev. 52 (2), 237-268 (2000).

5.G. Bergers and L. E. Benjamin, Tumorigenesis and the angiogenic switch, Nat. Rev. Cancer 3 (6), 401-410 (2003).

6.R. Kerbel and J. Folkman, Clinical translation of angiogenesis inhibitors, Nature Review/Cancer 2, 727-739 (2002).

7.N. W. Gale and G. D. Yancopoulos, Growth factors acting via endothelial cell-specific receptor tyrosine kinases: VEGFs, angiopoietins, and ephrins in vascular development,

Genes & Development 13, 1055-1066 (1999).

8.M. Boulton et al., VEGF localisation in diabetic retinopathy, Br. J. Ophthalmol. 82, 561-568 (1998).

9.E. A. Pierce et al., Vascular endothelial growth factor/vascular permeability factor expression in a mouse model of retinal neovascularization, Proc. Natl. Acad. Sci. USA 92, 905-909 (1995).

10.S. G. Robbins et al., Detection of vascular endothelial growth factor (VEGF) protein in vascular and non-vascular cells of the normal and oxygen-injured rat retina, Growth Factor 14, 279-295 (1997).

11.S. G. Robbins, V. S. Rajaratnam, and J. S. Penn, Evidence for upregulation and redistribution of vascular endothelial growth factor receptors flt-1 and flk-1 in the oxygen-injured rat retina, Growth Factors 16, 1-9 (1998).

12.L. P. Aiello et al., Suppression of retinal neovascularization in vivo by inhibition of vascular endothelial growth factor (VEGF) using soluble VEGF-receptor chimeric proteins, Proc. Natl. Acad. Sci. USA 92, 10457-10461 (1995).

13.G. S. Robinson et al., Oligodeoxynucleotides inhibit retinal neovascularization in a murine model of proliferative retinopathy, Proc. Natl. Acad. Sci. USA 93, 4851-4856 (1996).

14.M. Hangai et al., Systemically expressed soluble Tie2 inhibits intraocular neovascularization, Hum. Gene Ther. 12, 1311-1321 (2001).

15.H. Takagi et al., Potential role of the angiopoietin/tie2 system in ischemia-induced retinal neovascularization, Invest. Ophthalmol. Vis. Sci. 44, 393-402 (2003).

16.E. N. Committee, Unified nomenclature for Eph family receptors and their ligands, the Ephrins, Cell 90, 403-404 (1997).

17.N. W. Gale et al., Eph receptors and ligands comprise two major specificity subclasses and are reciprocally compartmentalized during embryogenesis, Neuron 17, 9-19 (1996).

18.J. P. Himanen et al., Repelling class discrimination: ephrin-A5 binds to and activates EphB2 receptor signaling, Nat. Neurosci. 7, 501-509 (2004).

19.K. Bruckner et al., EphrinB ligands recruit GRIP family PDZ adaptor proteins into raft membrane microdomains, Neuron 22, 511-524 (1999).

20.R. Torres, B. L. Firestein, H. Dong, J. Staudinger, E. N. Olson, R. L. Huganir, and G. D. Yancopoulos, PDZ proteins bind, cluster, and synaptically colocalize with Eph receptors and their ephrin ligands, Cell 21, 1453-1463 (1998).

21.J. P. Himanen et al., Crystal structure of an Eph receptor-ephrin complex, Nature 414, 933-938 (2001).

22.K. Kullander and R. Klein, Mechanisms and functions of Eph and ephrin signaling, Nat. Rev. Mol. Cell Biol. 3, 475 (2002).

23.N. K. Noren and E. B. Pasquale, Eph receptor-ephrin bidirectional signals that target Ras and Rho proteins, Cell Signal. 16, 655-666 (2004).

24.R. H. Adams et al., Roles of ephrinB ligands and EphB receptors in cardiovascular development: demarcation of arterial/venous domains, vascular morphogenesis, and sprouting angiogenesis, Genes & Development 3, 295-306 (1999).

25.R. K. Baker and P. B. Antin, Ephs and ephrins during early stages of chick embryogenesis, Dev. Dyn. 228 (1), 128-142 (2003).

26.P. M. Helbling, D. M. Saulnier, and A. W. Brandli, The receptor tyrosine kinase EphB4 and ephrin-B ligands restrict angiogenic growth of embryonic veins in Xenopus laevis, Development 127 (2), 269-278 (2000).

27.K. Othman-Hassan et al., Arterial identity of endothelial cells is controlled by local cues, Dev. Biol. 237 (2), 398-409 (2001).

28.H. U. Wang, Z. F. Chen, and D. J. Anderson, Molecular distinction and angiogenic interaction between embryonic arteries and veins revealed by ephrin-B2 and its receptor Eph-B4, Cell 93, 741-753 (1998).

29.S. S. Gerety, et al., Symmetrical mutant phenotypes of the receptor EphB4 and its specific transmembrane ligand ephrin-B2 in cardiovascular development, Molecular Cell 4, 403-414 (1999).

30.R. H. Adams et al., The cytoplasmic domain of the ligand ephrinB2 is required for vascular morphogenesis but not cranial neural crest migration, Cell 104, 57-69 (2001).

31.Z. Wang et al., Ephrin receptor, EphB4, regulates ES cell differentiation of primitive mammalian hemangioblasts, blood, cardiomyocytes, and blood vessels, Blood 4, 4 (2003).

32.N. W. Gale, et al., Ephrin-B2 selectively marks arterial vessels and neovascularization sites in the adult, with expression in both endothelial and smooth-muscle cells, Dev. Biol. 230, 151-160 (2001).

33.D. Shin et al., Expression of ephrinB2 identifies a stable genetic difference between arterial and venous vascular smooth muscle as well as endothelial cells, and marks subsets of microvessels at sites of adult neovascularization, Dev. Biol. 230 (2), 139-150 (2001).

34.Y. Oike et al., Regulation of vasculogenesis and angiogenesis by EphB/ephrin-B2 signaling between endothelial cells and surrounding mesenchymal cells, Blood 100 (4), 1326-1333 (2002).

35.S. S. Gerety and D. J. Anderson, Cardiovascular ephrinB2 function is essential for embryonic angiogenesis, Development 129 (6), 1397-1410 (2002).

36.X. Q. Zhang et al., Stromal cells expressing ephrin-B2 promote the growth and sprouting of ephrin-B2(+) endothelial cells, Blood 98 (4), 1028-1037 (2001).

37.T. Fuller et al., Forward EphB4 signaling in endothelial cells controls cellular repulsion and segregation from ephrinB2 positive cells, J. Cell Sci. 116 (Pt 12), 2461-2470 (2003).

38.K. Hamada et al., Distinct roles of ephrin-B2 forward and EphB4 reverse signaling in endothelial cells, Arterioscler. Thromb. Vasc. Biol. 23 (2), 190-197 (2003).

39.D. M. Brantley et al., Soluble EphA receptors inhibit tumor angiogenesis and progression in vivo, Oncogene 21, 7011-7026 (2002).

40.N. Cheng et al., Blockade of EphA receptor tyrosine kinase activation inhibits VEGFinduced angiogenesis, Mol. Cancer Res. (formerly Cell Growth and Differentiation) 1, 2-11 (2002).

41.T. O. Daniel et al., Elk and LERK-2 in developing kidney and microvascular endothelial assembly, Kidney Int. Suppl. 57, S73-S81 (1996).

42.J. L. McBride and J. C. Ruiz, ephrinA1 is expressed at sites of vascular development in the mouse, Mech. Dev. 77, 201-204 (1998).

43.A. Pandey et al., Role of B61, the ligand for the Eck receptor tyrosine kinase, in TNF- alpha-induced angiogenesis, Science 268 (5210), 567-569 (1995).

44.H. Takahashi and T. Ikeda, Molecular cloning and expression of rat and mouse B61 gene: implications on organogenesis, Oncogene 11 (5), 879-883 (1995).

45.K. Nagashima et al., Adaptor protein Crk is required for ephrin-B1-induced membrane ruffling and focal complex assembly of human aortic endothelial cells, Mol. Biol. Cell 13 (12), 4231-4242 (2002).

46.Y. Sawai et al., Expression of ephrin-B1 in hepatocellular carcinoma: possible involvement in neovascularization, J. Hepatol. 39 (6), 991-996 (2003).

47.E. Stein et al., Eph receptors discriminate specific ligand oligomers to determine alternative signaling complexes, attachment, and assembly responses, Genes Dev. 12 (5), 667-678 (1998).

48.U. Huynh-Do et al., Ephrin-B1 transduces signals to activate integrin-mediated migration, attachment and angiogenesis, J. Cell Sci. 115, 3073-3081 (2002).

49.C. Vindis et al., EphB1 recruits c-Src and p52Shc to activate MAPK/ERK and promote chemotaxis, J. Cell Biol. 162 (4), 661-671 (2003).

50.H. Maekawa et al., Ephrin-B2 induces migration of endothelial cells through the phosphatidylinositol-3 kinase pathway and promotes angiogenesis in adult vasculature,

Arterioscler. Thromb. Vasc. Biol. 23 (11), 2008-2014 (2003).

51.D. Brantley-Sieders et al., EphA2 receptor tyrosine kinase regulates endothelial cell migration and assembly through phosphoinositide 3-kinase-mediated Rac1 GTPase activation, J. Cell Sci. 117, 2037-2049 (2004).

52.H. Fujiwara et al., Human endometrial epithelial cells express ephrin A1: possible interaction between human blastocysts and endometrium via Eph-ephrin system, J. Clin. Endocrinol. Metab. 87 (12), 5801-5807 (2002).

53.L. C. Kao et al., Expression profiling of endometrium from women with endometriosis reveals candidate genes for disease-based implantation failure and infertility, Endocrinology 144 (7), 2870-2881 (2003).

54.G. Berclaz et al., Activation of the receptor protein tyrosine kinase EphB4 in endometrial hyperplasia and endometrial carcinoma, Ann. Oncol. 14, 220-226 (2003).

55.N. Takai et al., Expression of receptor tyrosine kinase EphB4 and its ligand ephrin-B2 is associated with malignant potential in endometrial cancer, Oncol. Rep. 8, 567-573 (2001).

56.S. Ozuysal et al., Angiogenesis in endometrial carcinoma: correlation with survival and clinicopathologic risk factors, Gynecol. Obstet. Invest. 55 (3), 173-177 (2003).

57.J. S. Davis, B. R. Rueda, and K. Spanel-Borowski, Microvascular endothelial cells of the corpus luteum, Reprod. Biol. Endocrinol. 1 (1), 89 (2003).

58.M. Egawa et al., Ephrin B1 is expressed on human luteinizing granulosa cells in corpora lutea of the early luteal phase: the possible involvement of the B class Eph-ephrin system during corpus luteum formation, J. Clin. Endocrinol. Metab. 88 (9), 4384-4392 (2003).

59.M. E. Schaner et al., Gene expression patterns in ovarian carcinomas, Mol. Biol. Cell 14 (11), 4376-4386 (2003).

60.K. Ogawa et al., The ephrin-A1 ligand and its receptor, EphA2, are expressed during tumor neovascularization, Oncogene 19, 6043-6052 (2000).

61.N. Cheng et al., Inhibition of VEGF-dependent multi-stage carcinogenesis by soluble EphA receptors, Neoplasia 5, 445-456 (2003).

62.J. Chen et al., Germline inactivation of the murine Eck receptor tyrosine kinase by retroviral insertion, Oncogene 12, 979-988 (1996).

63.C. M. Naruse-Nakajima, M. Asano, and Y. Iwakura, Involvement of EphA2 in the formation of the tail notochord via interaction with ephrinA1, Mech. Dev. 102, 95-105 (2001).

64.D. M. Brantley-Sieders et al., Impaired tumor microenvironment in EphA2-deficient mice inhibits tumor angiogenesis and metastatic progression, FASEB J. 19, 1884-1886 (2005).

65.W. Liu et al., Effects of overexpression of ephrin-B2 on tumour growth in human colorectal cancer, Br. J. Cancer 90, 1620-1626 (2004).

66.N. K. Noren et al., Interplay between EphB4 on tumor cells and vascular ephrin-B2 regulates tumor growth, Proc. Natl. Acad. Sci. U S A. 101, 5583-5588 (2004).

67.G. Martiny-Baron et al., Inhibition of tumor growth and angiogenesis by soluble EphB4, Neoplasia 6, 248-257 (2004).

68.C. Deroanne et al., EphrinA1 inactivates integrin-mediated vascular smooth muscle cell spreading via the Rac/PAK pathway, J. Cell Sci. 116, 1367-1376 (2003).

69.A. P. Adamis, L. P. Aiello, and R. A. D’Amato, Angiogenesis and ophthalmic disease, Angiogenesis 3 (1), 9-14 (1999).

70.J. J. Steinle et al., Role of ephrin B2 in human retinal endothelial cell proliferation and migration, Cell. Signal. 15 (11), 1011-1017 (2003).

71.N. Umeda et al., Expression of ephrinB2 and its receptors on fibroproliferative membranes in ocular angiogenic diseases, Am. J. Ophthalmol. 138, 270-279 (2004).

72.J. Chen et al., Inhibition of retinal neovascularization by soluble EphA2 receptor, Exp. Eye Res. 82 (4), 664-673 (2006).

73.D. Orioli et al., Sek4 and Nuk receptors cooperate in guidance of commissural axons and in palate formation, EMBO J. 15, 6035-6049 (1996).

74.M. Henkemeyer et al., Nuk controls pathfinding of commissural axons in the mammalian central nervous system, Cell 86, 35-46 (1996).

Chapter 12

ADENOSINE IN RETINAL VASCULOGENESIS AND ANGIOGENESIS IN OXYGEN-INDUCED RETINOPATHY

Gerard A. Lutty, PhD, and D. Scott McLeod

Wilmer Ophthalmological Institute, Johns Hopkins Hospital, Baltimore, Maryland

221

J.S. Penn (ed.), Retinal and Choroidal Angiogenesis, 221–240.

© Springer Science+Business Media B.V. 2008

A2B receptor significantly reduced retinal neovascularization in mice,1 as did cleavage of A2B by a ribozyme.2 These studies suggest that adenosine and its receptors are important in retinal vascular development and may be a therapeutic target in OIR.

1.INTRODUCTION

Adenosine is a ubiquitous molecule that is produced predominantly by catabolism of adenosine triphosphate. Few signaling molecules have the ability to influence development like the nucleoside adenosine. Linden called adenosine a “primordial signaling molecule” that is present in and modulates physiological responses in all mammals.3 Adenosine levels in tissues change with increased tissue activity, hypoxia, or stress. Adenosine levels in normal tissues range from 1-50 nM and can rapidly climb to 1000 nM with ischemia and increased tissue activity.4 The action of adenosine is most prominent in tissues where oxygen demand is high, like retina.

There are two major sources of adenosine: S-adenylsylhomocysteine and adenosine monophosphate (AMP). S-adenylsylhomocysteine is hydrolyzed to adenosine by S-adenylsylhomocysteine hydrolase, which is prominent in myocardium, but little is known about this source of adenosine in retina. AMP, however, is prominent in the eye, and it is processed, as in other tissues, primarily by 5’ nucleotidase (5’N, also known as CD73). 5’N (E.C. 3.1.3.5.) is an ectoenzyme that catalyzes the hydrolysis of purine, but not pyrimidine, nucleotide monophosphates to their corresponding nucleosides. Although 5’N can metabolize all purine monophosphates, the major product in ischemic dog hearts is adenosine.5 Braun et al. have demonstrated that 5’N expression is elevated during cerebral ischemia.6 In heart, 5’N is upregulated during hypoxia, and subsequently, adenosine levels increase 50fold.7 Synnestradt and associates recently demonstrated that 5’N expression is regulated by hypoxia-inducible factor-1 (HIF-1), explaining at least in part the increased production of adenosine in hypoxia.8 Adenosine is degraded to inosine by adenosine deaminase, which has been found in endothelial and smooth muscle cells, or adenosine kinase, which makes AMP from it.

2.ADENOSINE RECEPTORS

The four recognized adenosine receptor subtypes are A1, A2A, A2B, and A3, and all are coupled to G-proteins. The A1 receptor has the highest affinity for adenosine and acts through Gi- and Go-proteins;9 therefore, binding the A1 receptor inhibits adenylate cyclase,10 activates phospholipase C,11 and opens

K+ATP channels.12 The A2 receptors are coupled to the Gs-protein,13 and their transduction systems involve stimulation of adenylate cyclase14 and

activation of N-type Ca2+ channels (Figure 1).15 A3 receptors are coupled to Gi- and Gq-proteins,16 and their transduction system includes the activation of phospholipase C/D17 and inhibition of adenylate cyclase.18,19

3.EFFECTS OF ADENOSINE ON BLOOD VESSELS

Adenosine is a local regulator of blood flow in several organs.20-22 It may either contract or relax blood vessels, depending on the organ and which receptors are present.23 Vasodilation in response to adenosine may be modulated through nitric oxide (NO) production.24

Dusseau and Hutchins demonstrated that hypoxia-induced angiogenesis on the chorioallantoic membrane (CAM) in chicken was due to adenosine

production and uptake.25 In vitro, adenosine is chemotactic and mitogenic for endothelial cells from large blood vessels.26,27 We have determined that

adenosine does not stimulate proliferation of dog retinal microvascular endothelial cells but does stimulate endothelial cell migration and tube formation, two events that are critical in the development of the primary retinal vasculature in dog.28 Olanrewaju et al. found that human and porcine coronary artery endothelial cells have both A2A and A2B receptors.29 Feoktistov and associates have recently demonstrated a differential expression of adenosine receptors by large vessel versus microvascular endothelial cells: dermal microvascular endothelial cells expressed A2B receptors, and human umbilical vein endothelial cells (HUVECs) expressed A2A receptors.30 Dubey et al. observed that A2B agonists, and not A2A agonists, stimulated proliferation of porcine and rat arterial endothelial cells in culture.31 Grant and associates, using human retinal microvascular endothelial cells, found that A2B receptor stimulation caused increased proliferation, migration, tube formation, and extracellular signal-related kinase (ERK) activation (Figure 1).32