Ординатура / Офтальмология / Английские материалы / Retinal and Choroidal Angiogenesis_Penn_2008

vasculature and may exacerbate the insult to the peripheral retina or to the leading edge of the vasculature, leading to preretinal neovascularization. Systemic T4 supplementation in human infants is controversial, and its potential role in reducing risk for ROP deserves further study.30,31

7.GENETIC FACTORS

Although genetic factors may not lead directly to ROP or preretinal neovascularization, they appear to be important in setting the stage for whether neovascularization will develop or not, given a specific set of stimuli.

In a study of OIR and AIR in Sprague-Dawley rats from two different vendors (Harlan [Indianapolis, IN] versus Charles River [Wilmington, MA]), we found more frequent and more severe neovascularization in Charles River rats in conditions of OIR but not AIR.32 Similarly, in a recent study comparing unpigmented Sprague-Dawley rats to pigmented Brown Norway rats, we found more frequent and severe neovascularization in Brown Norway rats, again in OIR but not in AIR.33 We speculate that there may be different responses of critical growth factors or enzymes, in the context of different genetic backgrounds, to similar angiogenic stimuli. Further work is needed to elucidate which critical factors differ between rats from different strains and sources, and the role of these factors in the development of preretinal neovascularization.

8.CONCLUSION

A variety of non-oxygen factors lead to preretinal neovascularization in immature retinae: carbon dioxide, systemic acidosis, systemic alkalosis, gastrointestinal infection, low serum IGF-1, and low serum T4. Our working hypothesis is that these factors may damage the developing vasculature in a way that is analogous to the damage by oxygen. Whether that “damage” leads down a common path to increased ischemia of the peripheral retina and subsequent neovascularization, or whether angiogenesis is a direct response to that damage, remains to be elucidated. Nevertheless, in any strategy of ROP prevention, and perhaps prevention of other diseases characterized by preretinal neovascularization, attention must also be given to factors other than oxygen.

REFERENCES

1.V.E. Kinsey, Retrolental fibroplasia: cooperative study of retrolental fibroplasia and the use of oxygen, Arch. Ophthalmol. 56, 481-529, (1956).

2.V.E. Kinsey, H.J. Arnold, R.E. Kalina, L. Stern, M. Stahlman, G. Odell, J.M. Driscoll Jr, J.H. Elliott, J. Payne and A. Patz, PaO2 levels and retrolental fibroplasia: a report of the cooperative study, Pediatrics, 60(5), 655-668, (1977).

3.D.L. Phelps, Retinopathy of prematurity: an estimate of vision loss in the United States-- 1979, Pediatrics, 67(6), 924-925, (1981).

4.E.A. Palmer, The continuing threat of retinopathy of prematurity, Am. J. Ophthalmol. 122(3), 420-423, (1996).

5.J.M. Holmes and L.A. Duffner, The effect of postnatal growth retardation on abnormal neovascularization in the oxygen exposed neonatal rat, Curr. Eye Res. 15(4), 403-409, (1996).

6.J.S. Penn, M.M. Henry and B.L. Tolman, Exposure to alternating hypoxia and hyperoxia causes severe proliferative retinopathy in the newborn rat.[erratum appears in Pediatr. Res. 1995 Mar;37(3):353], Pediatr. Res. 36(6), 724-731, (1994).

7.J.S. Penn, B.L. Tolman and L.A. Lowery, Variable oxygen exposure causes preretinal neovascularization in the newborn rat, Invest. Ophthalmol.Vis. Sci. 34(3), 576-585, (1993).

8.X. Reynaud and C.K. Dorey, Extraretinal neovascularization induced by hypoxic episodes in the neonatal rat, Invest. Ophthalmol. Vis. Sci. 35(8), 3169-3177, (1994).

9.C.K. Dorey, S. Aouididi, X. Reynaud, H.F. Dvorak and L.F. Brown, Correlation of vascular permeability factor/vascular endothelial growth factor with extraretinal neovascularization in the rat., Arch. Ophthalmol. 114(10), 1210-1217, (1996).

10.B.A. Berkowitz and W. Zhang, Significant reduction of the panretinal oxygenation response after 28% supplemental oxygen recovery in experimental ROP,

Invest.Ophthalmol. Vis. Sci. 41(7), 1925-1931, (2000).

11.G.A. Lutty and D.S. McLeod, A new technique for visualization of the human retinal vasculature, Arch. Ophthalmol. 110, 267-276, (1992).

12.J.M. Holmes and L.A. Duffner, The effect of litter size on normal retinal vascular development in the neonatal rat, Curr. Eye Res. 14(8), 737-740, (1995).

13.E.A. Pierce, R.L. Avery, E.D. Foley, L.P. Aiello and L.E. Smith, Vascular endothelial growth factor/vascular permeability factor expression in a mouse model of retinal neovascularization, Proc. Natl. Acad. Sci. USA, 92(3), 905-909, (1995).

14.E.A. Pierce, E.D. Foley and L.E. Smith, Regulation of vascular endothelial growth factor by oxygen in a model of retinopathy of prematurity, Arch. Ophthalmol. 114(10), 1219-1228, (1996).

15.K.J. Johns, J.A. Johns, S.S. Feman and D.A. Dodd, Retinopathy of prematurity in infants with cyanotic congenital heart disease, Am. J. Dis. Child. 145(2), 200-203, (1991).

16.J.F. Lucey and B. Dangman, A reexamination of the role of oxygen in retrolental fibroplasia, Pediatrics, 73(1), 82-96, (1984).

17.J.M. Holmes, S. Zhang, D.A. Leske and W.L. Lanier, The effect of carbon dioxide on oxygen-induced retinopathy in the neonatal rat, Curr. Eye Res. 16(7), 725-732, (1997).

18.J.M. Holmes, D.A. Leske and S. Zhang, The effect of raised inspired carbon dioxide on normal retinal vascular development in the neonatal rat, Curr. Eye Res. 16(1), 78-81, (1997).

19.J.M. Holmes, S. Zhang, D.A. Leske and W.L. Lanier, Carbon dioxide-induced retinopathy in the neonatal rat, Curr. Eye Res. 17(6), 608-616, (1998).

20.J.M. Holmes, S. Zhang, D.A. Leske and W.L. Lanier, Metabolic acidosis-induced retinopathy in the neonatal rat, Invest. Ophthalmol. Vis. Sci. 40(3), 804-809, (1999).

21.S. Zhang, D.A. Leske, W.L. Lanier, B.A. Berkowitz and J.M. Holmes, Preretinal neovascularization associated with acetazolamide-induced systemic acidosis in the neonatal rat, Invest. Ophthalmol. Vis .Sci. 42(5), 1066-1071, (2001).

22.Y. Chen, D.A. Leske, S. Zhang, R.A. Karger, W.L. Lanier and J.M. Holmes, Duration of acidosis and recovery determine preretinal neovascularization in the rat model of acidosis-induced retinopathy, Curr. Eye Res. 24(4), 281-288, (2002).

23.Y. Harel, V. Niranjan and B.J. Evans, The current practice patterns of mechanical ventilation for respiratory failure in pediatric patients, Heart Lung, 27(4), 238-244, (1998).

24.A.N. Ammari and K.F. Schulze, Uses and abuses of sodium bicarbonate in the neonatal intensive care unit, Curr. Opin. Pediatr. 14(2), 151-156, (2002).

25.J.P. Berdahl, D.A. Leske, M.P. Fautsch, W.L. Lanier and J.M. Holmes, Effect of bicarbonate on retinal vasculature and acidosis-induced retinopathy in the neonatal rat,

Graefes Arch. Clin. Exp. Ophthalmol. 243, 367-73, (2005).

26.S. Zhang, D.A. Leske, J.R. Uhl, F.R. Cockerill, 3rd, W.L. Lanier and J.M. Holmes, Retinopathy associated with enterococcus enteropathy in the neonatal rat, Invest. Ophthalmol. Vis. Sci. 40(6), 1305-1309, (1999).

27.S. Zhang, D.A. Leske, W.L. Lanier and J.M. Holmes, Postnatal growth retardation exacerbates acidosis-induced retinopathy in the neonatal rat, Curr. Eye Res. 22(2), 133-139, (2001).

28.A. Hellstrom, E. Engstrom, A.L. Hard, K. Albertsson-Wikland, B. Carlsson, A. Niklasson, C. Lofqvist, E. Svensson, S. Holm, U. Ewald, G. Holmstrom and L.E. Smith, Postnatal serum insulin-like growth factor I deficiency is associated with retinopathy of prematurity and other complications of premature birth, Pediatrics, 112(5), 1016-1020, (2003).

29.M. Mookadam, D.A. Leske, M.P. Fautsch, W.L. Lanier and J.M. Holmes, The antithyroid drug methimazole induces neovascularization in the neonatal rat analogous to ROP, Invest. Ophthalmol. Vis. Sci. 45, 4145-4150, (2004).

30.L. Mutapcic, S.M.E. Wren, D.A. Leske, M.P. Fautsch, J.M. Holmes. The effect of L-Thyroxine supplementation on retinal vascular development in neonatal rats. Curr Eye Res 30, 1035-40, (2005).

31.S.M.E. Wren, L. Mutapcic, D.A. Leske, M.P. Fautsch, J.M. Holmes. The effect of L-Thyroxine supplementation in a neonatal rat model of ROP, Curr. Eye Res. 31, 669-674, (2006).

32.A.S. Kitzmann, D.A. Leske, Y. Chen, A.M. Kendall, W.L. Lanier and J.M. Holmes, Incidence and severity of neovascularization in oxygenand metabolic acidosis-induced retinopathy depend on rat source, Curr. Eye Res. 25(4), 215-220, (2002).

33.B.N.I. Floyd, D.A. Leske, S.M.E. Wren, M. Mookadam, M.P. Fautsch and J.M. Holmes, Differences between rat strains in models of retinopathy of prematurity. Mol. Vis. 11, 524-530, (2005).

Chapter 16

GROWTH FACTOR SYNERGY

IN ANGIOGENESIS

Growth Factor Interactions

Alexander V. Ljubimov

Ophthalmology Research Laboratories, Cedars-Sinai Medical Center, and David Geffen School of Medicine, UCLA, Los Angeles, California

1.INTRODUCTION

Angiogenesis is a fundamental process of blood and lymphatic vessel growth during development and tissue repair. Angiogenesis also occurs in pathological conditions, such as myocardial infarction, proliferative retinopathies, wet form of macular degeneration, and cancer. This process involves a number of tightly controlled discrete steps.1,2 New vessel growth may occur from endothelial precursor or stem cells (vasculogenesis) or as budding, sprouting, and elongation of pre-existing vessels (angiogenesis). Both mechanisms have been demonstrated for developmental and pathological angiogenic processes.

Since the pioneering work of Folkman3 and others in the 1970’s, a variety of growth factors and cytokines have been described that can induce, enhance, attenuate, inhibit, or otherwise regulate normal and pathological angiogenesis. The best known angiogenesis regulators (for review, see 4) include basic fibroblast growth factor (FGF-2); vascular endothelial (VEGF),

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insulin-like (IGF-I), hepatocyte (HGF), platelet-derived (PDGF), placenta (PlGF), pigment epithelium-derived (PEDF), and transforming (TGF-E) growth factors; stromal-derived factor-1 (SDF-1); interleukin (IL)-6, -8, and –10; tumor necrosis factor (TNF)-D; and angiopoietins (Ang). Extracellular matrix proteins, such as tenascin-C, endostatin, and thrombospondins, have also been shown to either enhance or inhibit angiogenesis.

All these different molecular effectors interact with endothelial cells using various receptors, which transduce signals inside the cell. In recent years, significant progress has been made in unraveling major signaling pathways of angiogenic growth factors and cytokines. The emergence of reliable models of retinal neovascularization has helped tremendously to dissect important pathways leading to the angiogenic response.5 However, many aspects of receptor signaling that mediates angiogenic responses remain unclear.4 Understanding the mechanisms of action of angiogenic growth factors and cytokines would facilitate the development of rational approaches to inhibit the pathological angiogenesis, called neovascularization (NV), seen in cancer and various retinopathies. One of the most important aspects, which will be considered here, is how different angiogenic factors interact with each other at the molecular level to control the specificity and extent of the angiogenic response of the cell.

Earlier work using tumor models and later studies using retinopathy models established the role of specific growth factors in NV. At first, only a few factors, like endothelial cell growth factor (ECGF) and FGFs, were considered angiogenic. In the late 1980’s, VEGF was cloned by Ferrara’s group, which started an avalanche of studies of angiogenesis and the development of therapeutic modalities for its prevention in cancer, diabetic retinopathy, and macular degeneration. Direct expression experiments have identified a causative role for specific factors, e.g., VEGF, in NV in tumors and retinopathies.5-7

At the same time, after the properties of the first few angiogenic growth factors had been described, it became apparent that tumors secreted more than one such factor.8-14 In the eye, since the pioneering paper by Grant’s group15 documenting increased expression of IGF-I in the vitreous of patients with diabetic retinopathy (DR), elevated concentrations of a variety of angiogenic growth factors, including VEGF, PlGF, FGF-2, PDGF, HGF, TGF-E, etc., were found in the vitreous of patients with DR and proliferative DR.4,16-21 Increased expression of several growth factors was found in retinas and vitreous from patients with proliferative DR, sickle cell retinopathy, and vitreoretinopathy.19,22-25 It was thus suggested that endothelial cell proliferation in vivo with new vessel formation could probably be triggered only when more than one angiogenic factor was upregulated.26 In fact, in the earlier work of Montesano27 and Folkman,28 it was found that angiogenic

growth factors could act in concert with each other to enhance or, in other situations, inhibit the angiogenic response of endothelial cells. Below, the literature on the interactions of selected growth factors will be reviewed in more detail.

2.COMBINED EFFECTS OF ANGIOGENIC GROWTH FACTORS

2.1VEGFs

Vascular endothelial growth factor is a member of a family of at least six

angiogenic factors and a major stimulator of NV in tumors and retinopathies.9,14,29-31 It is also known as vascular permeability factor and a mediator of vascular leakage during NV.14,29,31 VEGF (the correct name is

VEGF-A) is currently considered to be the main mediator in hypoxiaand ischemia-induced tumor and ocular NV.32,33 Inhibitors of VEGF and its

signaling are currently being evaluated in several clinical trials for blocking cancer and ocular NV.8,29,31,34-36

In vitro, VEGF-A was shown to synergize with many growth factors in various angiogenic assays. In some reports, the effects of VEGF combinations with other factors exceeded those exerted by each factor alone

or a sum of individual factor effects (true synergy).37 In other instances, only additive effects could be demonstrated.26 The first reports27,28 have found

VEGF synergy with FGF-2 in increasing tube-like structure formation, cell proliferation, migration, and vascular sprouting in cultured microvascular and umbilical vein endothelial cells (Table 1). The combination was found to be more potent than the sum of the two factors. These data were corroborated by subsequent studies37,38 using Matrigel chamber and fibrin gel invasion assays (Table 1). Hata et al.39 have documented the induction by FGF-2 of VEGF receptor-2 (VEGFR2/KDR/flk-1) that is known to promote cell proliferation and migration. This induction occurs through stimulation of protein kinase C (PKC) and extracellular signal-regulated kinase (MAPK/ERK) pathways and may provide a mechanism for VEGF and FGF- 2 synergy.

At the same time, other researchers did not observe such a synergy in various in vitro26 and in vivo40 assays. There may be several reasons behind this discrepancy. If the in vitro assay is conducted without serum or in very low serum,26 direct interaction of VEGF and FGF-2 would be promoted. However, in the presence of serum that contains IGF-I, IGF-I receptor may be activated, in turn enhancing VEGF signaling through ERK.41 In these conditions, a synergy between VEGF and IGF-I26 (rather than between VEGF and FGF-2) would be taking place. Another explanation may be that the response of cells to growth factors may differ depending on the cell source (for example, different vascular beds may respond in a diverse manner) and on the species studied. In fact, synergy between VEGF-A and PlGF (a VEGF homolog) was observed in bovine aortic endothelial cells but not in human umbilical vein cells.42 In vivo, the combined action of exogenous angiogenic factors may be counteracted by the presence of antiangiogenic factors absent in vitro, e.g., nitric oxide, that may attenuate the effect.40 The controversy described demonstrates that the best way to test growth factor combinations is to use in vivo assays.

VEGF has also been shown to synergize in vitro (Table 1) with its homologs, VEGF-C43 and PlGF,37,44 as well as with FGF-1,45 HGF,46-49 IGF-

I,26 and estrogen.50 In these papers, tubulogenesis and cell proliferation were the most common parameters enhanced by growth factor combinations. It should be mentioned that VEGF synergy with PlGF in vitro was observed only when cells from PlGF knockout mice were used but not wild type cells.37 Therefore, these data should be interpreted with caution.

In different systems, possible mechanisms of synergy were examined. In a model of collagen gel invasion by endothelial cells, the combination of VEGF-A and VEGF-C potently induced plasminogen activator, which may have promoted matrix dissolution and cell invasion.43 In the case of PlGF, VEGF is able to directly stimulate its synthesis through PKC and MEK protein kinase.44 This, in turn, activates VEGFR-1, which may amplify the effects of VEGF on the endothelial cells as suggested by the authors.37 Synergy with HGF may be explained, at least in part, by the ability of HGF to induce VEGF expression and upregulate signaling cascades leading to an angiogenic response.46-49

Vascularization of ovarian cancer cell plugs in Matrigel implanted subcutaneously into mice was enhanced by a combination of VEGF and SDF-1 more than by either factor alone. VEGF was shown in this system to stimulate the synthesis of SDF-1 receptor.51 Together with VEGF, SDF-1 has been recently implicated in the development of NV in diabetic retinopathy and macular degeneration.52,53 Possibly, they synergize in these conditions as well, triggering abnormal angiogenesis in the retina and choroid, respectively.

Using an in vivo model of heart angiogenesis, VEGF was found to synergize with Ang-2 in stimulating NV.54,55 It also synergized with Ang-1

in a model of hind limb ischemic angiogenesis.56 However, in the heart model or in muscle angiogenesis,55,57 Ang-1 counteracted VEGF and its

combination with Ang-2 (Table 2), effectively reducing the angiogenic response. The mechanisms of these interactions remain to be elucidated.

An established VEGF antagonist is pigment epithelium-derived factor

(PEDF), which is considered to be the most potent endogenous antiangiogenic factor.58,59 PEDF inhibits NV in a mouse model of ischemic

retinopathy60 and increases apoptosis in tumor cells.59 Recent data indicate that PEDF can block VEGF expression at the mRNA and protein levels in osteosarcoma cells61 and inhibit VEGF-induced MAPK activation,58 which could be the mechanism of its antagonism with VEGF. However, when endothelial cells were cultured with VEGF for prolonged periods, PEDF became synergistic with VEGF (and also with FGF-2). This was accompanied by the stimulation of ERK phosphorylation by PEDF, which did not occur in cells cultured without VEGF.58 These data suggest that interpreting growth factor interactions should be done cautiously because these interactions may be influenced by a variety of variables, including cell type, assay system (in vitro vs. in vivo), assay duration, prior growth factor exposure, and even animal strain used.62

There are only limited data on the interactions of VEGF family members

beside VEGF-A with other growth factors. Both VEGF-C and PlGF were found to exert additive effects with FGF-2 using cultured cells.26,43 The

mechanisms of their combined action have not been investigated. It remains unknown whether FGF-2 can induce the expression of PlGF or VEGF-C.

2.2PDGFs

PDGF was one of the first isolated and cloned angiogenic growth factors.63 In the retina, it is generally considered as a pericyte recruitment and survival factor.2 However, it is also a potent endothelial mitogen and angiogenic

factor.63 Isoforms PDGF-BB and PDGF-CC are more potent in stimulating angiogenesis than PDGF-AA.64,65 In a rat model of myocardial infarction,

gene transfer of PDGF-BB and FGF-2 increased the number of both capillaries and arterioles more than each factor alone and gave rise to stable capillaries.66 However, in cultured arterial endothelial cells, and a chick chorioallantoic membrane model, PDGF inhibited FGF-2-induced tubulogenesis, cell migration, proliferation, and angiogenesis.67 This was attributed to a block of MAPK activation by PDGF-stimulated PDGF