The role of leptin in choroidal neovascularization

sutures were cut. After intravitreal injection of 0.4 ml of pure sulfur hexafluoride, the patient was instructed to maintain the position of turning onto her left side for 1 day. Twenty-four hours after gas injection, her macular fold had nearly resolved with adequate foveal displacement, and laser photocoagulation was immediately applied for the choroidal neovascular membrane. The fovea remained displaced at 1100 ␮m, and her visual acuity had improved to 20/40 3 months after the initial surgery (Figure 2). Retinal folds cannot be avoided in macular translocation achieved by scleral imbrication. In addition, retinal folds involving the macula may impair visual acuity because the macular fold can prevent the fovea from being reattached over new retinal pigment epithelium and can cause disturbance or loss of photoreceptors.4 Although, for these reasons, repeat surgery is necessary in eyes with severe postoperative macular folds, repeat vitrectomy may be complicated and invasive, not contributing to a better visual outcome.2,3 Refined treatment of this severe postoperative complication is required, therefore, to improve surgical results with limited macular translocation. This report demonstrates the feasibility of a simplified procedure without repeat vitrectomy for managing eyes with postoperative macular folds. Presumably, the mechanism by which the relocated fovea does not return to its original position after scleral shortening release is based on the retinal redundancy achieved by stretching of the neurosensory retina. de Juan and Vander reported a case of limited macular translocation without scleral shortening, which supports the possible stretching and flexibility of the detached retina.5 Scleral shortening release and intravitreal gas injection may be considered for the management of severe macular folds caused by limited macular translocation, because these procedures are simple and can achieve a good visual outcome. Additional studies are needed to further define the effectiveness of our novel treatment strategy.

The Role of Leptin in Choroidal Neovascularization Jing Z. Cui, MD, Dan Hornan, MB, BS, Michael J. Potter, MD, Mark D. J. Greve, MD, Brad J. Hinz, MD, Arif Samad, MD, and Joanne A. Matsubara, PhD PURPOSE: The purpose of this study was to investigate the

role of leptin in choroidal neovascularization. We examined the localization of leptin by immunohistochemistry in nine choroidal neovascular membranes surgically excised from patients with agerelated macular degeneration, idiopathic choroidal neovascularization, and ocular histoplasmosis. Controls included omission of primary antibody, use of an irrelevant primary antibody and leptin staining of posterior segment of four normal donor eyes. RESULTS: Leptin was present in eight membranes and appeared vesicular, within the cytoplasm. The more vascular membranes and those consisting of a larger number of retinal pigment epithelium cells were associated with greater leptin staining. Leptin was not seen in the posterior segment of the four normal eyes. CONCLUSION: We suggest that leptin plays an active role in choroidal neovascularization, although further experiments are necessary to establish a causal relationship. (Am J Ophthalmol 2001;132:792–794. © 2001 by Elsevier Science Inc. All rights reserved.) METHODS:

C

REFERENCES

1. de Juan E Jr, Loewenstem A, Bressler NM, Alexander J. Translocation of the retina for management of subfoveal choroidal neovascularization, II: a preliminary report in humans, Am J Ophthalmol 1998;125:635– 646. 2. Lewis H, Kaiser PK, Lewis S, Estatanous M. Macular translocation for subfoveal choroidal neovascularization in agerelated macular degeneration: a prospective study. Am J Ophthalmol 1999;128:135–146. 3. Fujii GY, Pieramici DJ, Humayun MS, et al. Complications associated with limited macular translocation. Am J Ophthalmol 2000;130:751–762. 4. Hayashi A, Usui S, Kawaguchi K, et al. Retinal changes after retinal translocation surgery with scleral imbrication in dog eyes. Invest Ophthalmol Vis Sci 2000;41:4288 – 4292. 5. de Juan E Jr, Vander JF. Effective macular translocation without scleral imbrication. Am J Ophthalmol 1999;128:380 – 382.

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HOROIDAL NEOVASCULARIZATION IS THE MOST COM-

mon cause of legal blindness in older adults in the United States,1 the most frequent cause being age-related macular degeneration. Indeed, choroidal neovascularization membranes are responsible for most cases of severe visual loss in age-related macular degeneration. These membranes extend from the choroid, perforate Bruch’s, and proliferate in the subpigment epithelial space. Previous work suggests that abnormalities of the extracellular matrix of retinal pigment epithelial cells promote an angiogenic phenotype that contributes to the development of choroidal neovascularization.2 Neovascularization is a serious consequence of several eye diseases. Recently, the anti-obesity hormone leptin has also been found to be an angiogenic factor,3 and it is capable of inducing angiogenesis in vivo.4 In this experimental study, we examined leptin localizaAccepted for publication Jun 1, 2001. From the Department of Ophthalmology (J.Z.C., D.H., M.J.P., J.A.M.), University of British Columbia, Vancouver, British Columbia, Canada, the Department of Ophthalmology (M.D.J.G., B.J.H.), University of Alberta, Alberta, Ontario, Canada, and the Department of Ophthalmology (A.S.), Dalhousie University, Halifax, Canada. Inquiries to Joanne A. Matsubara, PhD, Department of Ophthalmology, University of British Columbia, 2550 Willow St, Vancouver, British Columbia, V5Z 3N9, Canada; fax: 604-875-4663; e-mail: jms@interchange. ubc.ca

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FIGURE 1. Light microscopic photograph of a choroidal neovascular membrane section surgically excised from a 60-year-old man with age-related macular degeneration. (A) Leptin staining is seen in red, some of which is indicated by arrows. (B) Cytokeratin staining for retinal pigment epithelial cells is seen in red, some of which is shown by arrows. (C) CD31 staining for vascular endothelial cells is seen in red, some of which is indicated by arrows.

tion by immunohistochemistry in nine surgically excised choroidal neovascularization membranes: four age-related macular degeneration, four idiopathic choroidal neovascularization, and one ocular histoplasmosis. The membranes were placed in optimal cutting temperature medium within 30 minutes of excision, shipped on dry ice, and frozen in liquid nitrogen on arrival. Multiple 8-␮m cryosections from each membrane were air-dried, fixed in acetone for 5 minutes, washed with phosphate buffered saline, treated with 0.3% hydrogen peroxide, and blocked for 15 minutes with 1% bovine serum in phosphate buffered saline. They were then incubated for 30 minutes with primary antibody and washed three times for 5 minutes with phosphate buffered saline. Bound antibody was detected with avidinbiotin-conjugated secondary antibody and developed with aminoethyl-carbazole as a red chromogen. Five sections of VOL. 132, NO. 5

each membrane were mounted with Mayer’s. Monoclonal antibodies against cytokeratin were used to identify retinal pigment epithelial cells (Dako Corp., California), monoclonal antibodies against CD31 were used to identify endothelial cells (Dako), and a polyclonal antibody was used to stain for leptin (Biotechnology, Inc., Santa Cruz, California). Immunohistochemical staining was repeated on cryosections of the nine choroidal neovascularization membranes, omitting the anti-leptin primary antibody, and using an irrelevant primary antibody as controls. Additional controls included immunohistochemical staining of the posterior sclera, choroid, and retina of four normal donor eyes for leptin. The nine choroidal neovascular membranes ranged from moderately cellular with prominent neovascularization to paucicellular and fibrotic with no vascular channels. Lep-

BRIEF REPORTS

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4. Sierra-Honigmann MR, Nath AK, Murakami C, et al. Biological action of leptin as an angiogenic factor. Science 1998;281:1683–1686. 5. Uotani S, Bjorbaek C, Tornoe J, Flier JS. Functional properties of leptin receptor isoforms: internalization and degradation of leptin and ligand-induced receptor downregulation. Diabetes 1999;48(2):279 –286.

TABLE 1. Clinical and Histological Characteristics Eye

1 2 3 4 5 6 7 8 9

r r l l r l r l r

Diagnosis

Age

Sex

Leptin

CD31

Cytokeratin

AMD AMD Idiopathic Idiopathic AMD Idiopathic Idiopathic Histoplas. AMD

67 89 41 29 60 42 43 15 81

M F M F F F M F F

⫹ ⫹ ⫹ ⫹⫹ ⫹⫹ ⫹ ⫺ ⫹ ⫹⫹

⫹⫹ ⫹⫹⫹ ⫹⫹ ⫹⫹⫹ ⫹⫹⫹ ⫹⫹ ⫺ ⫹⫹ ⫹⫹⫹

⫹⫹ ⫹⫹⫹ ⫹⫹ ⫹⫹ ⫹⫹⫹ ⫹⫹ ⫹ ⫺ ⫹⫹⫹

Aqueous Humor and Plasma Vascular Endothelial Growth Factor in UveitisAssociated Cystoid Macular Edema Howard F. Fine, MD, MHSc, Judit Baffi, MD, George F. Reed, PhD, Karl G. Csaky, MD, PhD, and Robert B. Nussenblatt, MD

Quantified by counting the number of leptin positive cells and vascular channels in multiple 40⫻ objective fields: ⫺ represents ⬍1; ⫹ represents 1–9; ⫹⫹ represents 10 –20; ⫹⫹⫹ represents ⬎20.

PURPOSE:

To determine the association between cystoid macular edema and vascular endothelial growth factor concentration in the aqueous humor and plasma of uveitis patients. METHODS: Cross-sectional study. Vascular endothelial growth factor concentrations were measured by enzymelinked immunosorbent assay in the aqueous humor of 20 uveitis patients (9 with and 11 without cystoid macular edema), and in the plasma of 40 uveitis patients (20 with and 20 without cystoid macular edema) and 20 healthy volunteers. RESULTS: Mean aqueous humor vascular endothelial growth factor concentrations for uveitis patients with and without cystoid macular edema were 152.3 and 109.5 pg/ml, respectively, P ⴝ .044. Mean plasma vascular endothelial growth factor concentrations in uveitis patients with and without cystoid macular edema and in healthy volunteers were 32.2, 29.6, and 55.0 pg/ml, respectively. Uveitis patients had lower plasma vascular endothelial growth factor levels than did healthy volunteers, P ⴝ .0002 CONCLUSION: In uveitis patients, vascular endothelial growth factor concentration is increased in the aqueous humor of eyes with cystoid macular edema. It may be useful to investigate vascular endothelial growth factor antagonists as a treatment for uveitis-associated cystoid macular edema. (Am J Ophthalmol 2001;132: 794 –796. © 2001 by Elsevier Science Inc. All rights reserved.)

tin was revealed in all but one of the membranes (Figure 1A). Most of the leptin was localized to the cytoplasmic compartment of cells and was punctate. Many retinal pigment epithelium cells were observed in eight out of nine membranes (Figure 1B). Staining for endothelial cells was also seen in the leptin-positive membranes (Figure 1C). The results are summarized in Table 1. Leptin was not found within the posterior segment of the four normal eyes examined. The punctate appearance of leptin in the choroidal neovascular membranes implies an intravesicular location. Leptin internalization by the cell occurs by receptormediated phagocytosis, whereas degradation takes place in lysosomes.3 In previous studies of leptin as an angiogenic factor, the submembrane portion of the active leptin receptor (OB-Rb) was found to be intravesicular in endothelial cells.5 Our finding of leptin inside vesicles may represent either leptin–OB-Rb complexes or intralysosomal leptin. Greater leptin staining was found in the more vascular membranes; leptin has previously been shown to promote endothelial cell migration in vitro.3 Membranes consisting of a larger number of retinal pigment epithelium cells, thought to promote angiogenesis in choroidal neovascularization,2 were also associated with greater leptin staining. These findings suggest that leptin plays an active role in choroidal neovascularization. However, further experiments are needed to establish a causal relationship. REFERENCES

Accepted for publication Jun 4, 2001. From the Harvard Medical School, Boston, Massachusetts (H.F.F.), the Laboratory of Immunology, National Eye Institute, National Institutes of Health, Bethesda, Maryland (H.F.F., J.B., K.G.C., R.B.N.), and the Biometry Branch, National Eye Institute, National Institutes of Health, Bethesda, Maryland (G.F.R.). Inquiries to Howard F. Fine, MD, MHSc, National Eye Institute, National Institutes of Health, 10/10N202, 10 Center Dr, MSC 1863, Bethesda, MD 20892-1863; fax: (561) 828-0784; e-mail: [email protected]

1. Spaide RF. Choroidal neovascularization in younger patients. Curr Opin Ophthalmol 1999;10(3):177–181. 2. Campochiaro PA, Soloway P, Ryan SJ, Miller JW. The pathogenesis of choroidal neovascularization in patients with age-related macular degeneration. Mol Vis 1999;5:34. 3. Bouloumie´ A, Drexler HCA, Lafontan M, Busse R. Leptin, the product of Ob gene, promotes angiogenesis. Circ Res 1998;83:1059 –1066.

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