ELEVATED ERYTHROPOIETIN AND VASCULAR ENDOTHELIAL GROWTH FACTOR LEVELS IN AN ADOLESCENT WITH RETINAL NEOVASCULARIZATION FROM A CHRONIC RHEGMATOGENOUS RETINAL DETACHMENT Ben J. Kim, MD,* Nadia K. Waheed, MD,† Mario Romano, MD,† Fabrizio Scotti, MD,† Ali Hafezi-Moghadam, MD, PHD,‡ Donald J. D’Amico, MD†
Purpose: To evaluate the role of erythropoietin and vascular endothelial growth factor (VEGF) in a patient with retinal neovascularization from a rhegmatogenous retinal detachment of long duration. Methods: Fundus photography, fluorescein angiography, and vitreous analysis were performed. The vitreous concentrations of erythropoietin and VEGF were measured by enzyme-linked immunosorbent assays and compared with control levels. Results: An adolescent with a history of mild retinopathy of prematurity presented with a retinal detachment found by routine examination. The patient had a rhegmatogenous retinal detachment with signs of chronicity and extensive retinal neovascularization. The patient’s erythropoietin level was higher than those of patients with proliferative diabetic retinopathy. The patient’s VEGF level was not as high as those of patients with proliferative diabetic retinopathy but was elevated compared with those of patients without neovascularization. Conclusion: Vitreous concentrations of erythropoietin and VEGF can be elevated in patients with neovascularization secondary to a rhegmatogenous retinal detachment of long duration. RETINAL CASES & BRIEF REPORTS 2:117–120, 2008
D
From the *Department of Ophthalmology, the †Retina Service, and the ‡Angiogenesis Laboratory, Massachusetts Eye and Ear Infirmary, Harvard Medical School, Boston, Massachusetts.
uring retinal ischemia, there are likely multiple factors leading to retinal neovascularization. Vascular endothelial growth factor (VEGF) and, more recently, erythropoietin have been shown to play critical roles in retinal neovascularization during proliferative diabetic retinopathy.1–3 It has previously been reported that retinal neovascularization can also occur in the setting of a rhegmatogenous retinal detachment of long duration.4 Elevated VEGF levels have been found in the vitreous of some patients with chronic retinal detachments.1 We were interested in measuring the vitreous levels of erythropoietin and VEGF in a patient with retinal neovascularization secondary to a chronic rhegmatogenous retinal detachment. Similar
Supported in part by the American Diabetes Association and the Vitreoretinal Research Fund. Presented in part as a poster at the Association for Research in Vision and Ophthalmology Annual Meeting; Fort Lauderdale, FL; April 30 to May 4, 2006. The authors have no proprietary interest in the material presented. D. J. D. has been a consultant for Eyetech Pharmaceuticals and Alcon Laboratories. Reprint requests: Donald J. D’Amico, MD, Department of Ophthalmology, Weill Cornell Medical Center, 525 East 68th Street, Room F-835, New York, NY 10021; email: [email protected]
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Fig. 1. Fundus photographs and fluorescein angiogram of a 13-year-old girl diagnosed during a routine examination with a rhegmatogenous retinal detachment of long duration in the right eye. Left, Extensive retinal neovascularization was seen at the inferior equator of the right eye. Middle, Fluorescein angiogram of the right eye demonstrating fluorescein leakage from sites of retinal neovascularization. Right, Fundus photograph of the left eye showing peripheral retina changes consistent with mild retinopathy of prematurity.
to proliferative diabetic retinopathy, we found that VEGF and especially erythropoietin levels were elevated in this patient. Case Report A 13-year-old girl with a history of mild retinopathy of prematurity and high myopia was referred to the Retina Service at the Massachusetts Eye and Ear Infirmary (Boston) for evaluation of a retinal detachment found incidentally during an eye examination. Approximately 2.5 years earlier, the visual acuity was 20/30 in each eye with correction, and the refractive error was ⫺8.75 diopters for the right eye and ⫺12.25 diopters for the left eye. At that time, fundus examination did not reveal evidence of retinal detachment or neovascularization. She presented to the Retina Service with visual acuity of 20/200⫹2 in the right eye and 20/30 in the left eye. Examination of the right eye revealed a rhegmatogenous retinal detachment that extended from the 1:30-o’clock position through the 11-o’clock position. The detachment involved the macula and had signs of chronicity, including an inferior retina
that was extremely atrophic (Fig. 1, left). There was striking neovascularization at the inferior equator within the retina, which was confirmed by a fluorescein angiogram (Fig. 1, middle). Preretinal membranes and subretinal bands were also present. Examination of the left eye was notable for a posterior vitreous detachment and peripapillary atrophy. Both eyes had peripheral changes consistent with mild retinopathy of prematurity (Fig. 1, right). The patient underwent surgery to repair the detachment. Postoperatively, the retina remained reattached, although vision remained unchanged at 20/200.
Materials and Methods At the time of retinal surgery, undiluted vitreous specimens were acquired from this patient and five other patients (Table 1) who served as controls. All patients provided written informed consent. The study was conducted in accordance with HIPAA regulations and was approved by the Massachusetts Eye and Ear
Table 1. Patient Characteristics CRD Diagnosis
Age (y) Sex Diabetes history
PDR1
Proliferative Chronic diabetic retinal retinopathy detachment with vitreous with neovashemorrhage cularization
13 F —
PDR2
Proliferative diabetic retinopathy with clinically significant macular edema, vitreous hemorrhage, and tractional retinal detachment 58 56 M M 20-y history of 44-y history of diabetes type 1 currently diabetes managed with mellitus; insulin; previous previous panretinal panretinal photocoagulation photocoagulation of both eyes of both eyes
ARD Acute retinal detachment (⬍48 h) with multiple horseshoe tears superiorly
45 F —
ERM
FLO
Epiretinal Visually membrane significant floaters with a history of cataract surgery and YAG laser capsulotomy; mild epiretinal membrane 66 51 F M — —
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Fig. 2. Vitreous concentrations of erythropoietin and vascular endothelial growth factor (VEGF) as measured by enzyme linked immunosorbent assays (ELISAs). CRD, chronic retinal detachment; PDR, proliferative diabetic retinopathy; ARD, acute retinal detachment; ERM, epiretinal membrane; FLO, visually significant floaters. See Table 1 for further clinical data regarding these patients. Individual patient samples were measured in duplicates. Each of the data points (circles and squares) and the averages (horizontal line) are shown. Top Left, The patient with neovascularization from a chronic retinal detachment had the highest level of erythropoietin. Top Right, This patient’s VEGF level was not as high as those of the patients with proliferative diabetic retinopathy. Patient samples CRD, PDR1, and FLO were measured by ELISA 1 (triangles) and PDR2, ERM, and ARD were measured by ELISA 2 (diamonds) for the erythropoietin and VEGF assays. Bottom Left, Bottom Right, The standard curves for each of these ELISAs are shown.
Infirmary Institutional Review Board. The vitreous samples were immediately placed in heparinized tubes and stored at ⫺80°C. Enzyme linked immunosorbent assays (ELISAs) for erythropoietin and VEGF were performed according to the manufacturer’s instructions (Quantikine Human Epo Immunoassay and Human VEGF Immunoassay; R&D Systems, Inc., Minneapolis, MN). This VEGF ELISA measures the levels of the soluble isoforms (VEGF165 and VEGF121); however, it does not measure VEGF189 or VEGF206.5 For both erythropoietin and VEGF assays, three patient samples were run in parallel. Patient samples CRD, PDR1, and FLO were measured by ELISA 1 and PDR2, ERM, and ARD were measured by ELISA 2 for the erythropoietin and VEGF assays (see Table 1 for patient abbreviations). Individual patient samples were run in duplicate on each ELISA plate. Standard curves for each ELISA were constructed using the linear best line fit function. Results The vitreous of the patient with neovascularization secondary to the chronic retinal detachment had an erythropoietin level of 133 mIU/mL (Fig. 2, top left). Surprisingly, this level of erythropoietin was even higher than those of the two patients with proliferative diabetic ret-
inopathy. In contrast, the erythropoietin levels were low for the patients with no neovascularization. The patient with neovascularization secondary to the chronic retinal detachment also had a mildly elevated vitreous concentration of VEGF165 (77 pg/mL) compared with the patients with no neovascularization. However, this level of VEGF was not as high as the VEGF levels of the patients with proliferative diabetic retinopathy (Fig. 2, top right). Discussion We have shown that the vitreous concentrations of erythropoietin and VEGF are elevated in an adolescent patient with neovascularization secondary to a rhegmatogenous retinal detachment of long duration. Although elevated VEGF levels have been found in patients with a chronic retinal detachment,1 we are unaware of any previous report of an elevated erythropoietin level in such a case. The data demonstrate that the association of erythropoietin with retinal neovascularization is not limited to proliferative diabetic retinopathy. Surprisingly, the patient’s erythropoietin level in vitreous was even higher than those of the patients with proliferative diabetic retinopathy, while the level of VEGF was not as elevated compared with these patients. It has been previously reported that
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erythropoietin and VEGF are independent angiogenic factors in neovascularization.3 It is feasible that erythropoietin may be mechanistically more important than VEGF in the pathogenesis of some disorders that involve angiogenesis, such as chronic retinal detachment. If this were the case, then VEGF-blocking therapies alone would have limited success in preventing neovascularization in such a patient. Nevertheless, this conclusion cannot be made definitively from our data because it is not known how these varying concentrations of erythropoietin relate to stimulation of its receptor. One should also consider that erythropoietin is capable of playing several roles involving the retina. In addition to its angiogenic activity, erythropoietin is known to have protective effects for retinal cells by inhibiting apoptosis.6 In this case of a chronic retinal detachment, it is possible that erythropoietin may have roles in the prevention of apoptosis as well as neovascularization. Future studies will be necessary to elucidate the function of erythropoietin in retinal detachments and its relation to VEGF.
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Key words: erythropoietin, neovascularization, retinal detachment, vascular endothelial growth factor. References 1.
2.
3.
4.
5.
6.
Aiello LP, Avery RL, Arrigg PG, et al. Vascular endothelial growth factor in ocular fluid of patients with diabetic retinopathy and other retinal disorders. N Engl J Med 1994;331:1480– 1487. Aiello LP, Pierce EA, Foley ED, 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 1995;92:10457–10461. Watanabe D, Suzuma K, Matsui S, et al. Erythropoietin as a retinal angiogenic factor in proliferative diabetic retinopathy. N Engl J Med 2005;353:782–792. Bonnet M. Peripheral neovascularization complicating rhegmatogenous retinal detachments of long duration. Graefes Arch Clin Exp Ophthalmol 1987;225:59–62. Funatsu H, Yamashita H, Sakata K, et al. Vitreous levels of vascular endothelial growth factor and intracellular adhesion molecule 1 are related to macular edema. Ophthalmology 2005;112:806–816. Becerra SP, Amaral J. Erythropoietin—an endogenous retinal survival factor. N Engl J Med 2002;347:1968–1970.
Purpose: To evaluate the role of erythropoietin and vascular endothelial growth factor (VEGF) in a patient with retinal neovascularization from a rhegmatogenous retinal detachment of long duration. Methods: Fundus photography, fluorescein angiography, and vitreous analysis were performed. The vitreous concentrations of erythropoietin and VEGF were measured by enzyme-linked immunosorbent assays and compared with control levels. Results: An adolescent with a history of mild retinopathy of prematurity presented with a retinal detachment found by routine examination. The patient had a rhegmatogenous retinal detachment with signs of chronicity and extensive retinal neovascularization. The patient’s erythropoietin level was higher than those of patients with proliferative diabetic retinopathy. The patient’s VEGF level was not as high as those of patients with proliferative diabetic retinopathy but was elevated compared with those of patients without neovascularization. Conclusion: Vitreous concentrations of erythropoietin and VEGF can be elevated in patients with neovascularization secondary to a rhegmatogenous retinal detachment of long duration. RETINAL CASES & BRIEF REPORTS 2:117–120, 2008
D
From the *Department of Ophthalmology, the †Retina Service, and the ‡Angiogenesis Laboratory, Massachusetts Eye and Ear Infirmary, Harvard Medical School, Boston, Massachusetts.
uring retinal ischemia, there are likely multiple factors leading to retinal neovascularization. Vascular endothelial growth factor (VEGF) and, more recently, erythropoietin have been shown to play critical roles in retinal neovascularization during proliferative diabetic retinopathy.1–3 It has previously been reported that retinal neovascularization can also occur in the setting of a rhegmatogenous retinal detachment of long duration.4 Elevated VEGF levels have been found in the vitreous of some patients with chronic retinal detachments.1 We were interested in measuring the vitreous levels of erythropoietin and VEGF in a patient with retinal neovascularization secondary to a chronic rhegmatogenous retinal detachment. Similar
Supported in part by the American Diabetes Association and the Vitreoretinal Research Fund. Presented in part as a poster at the Association for Research in Vision and Ophthalmology Annual Meeting; Fort Lauderdale, FL; April 30 to May 4, 2006. The authors have no proprietary interest in the material presented. D. J. D. has been a consultant for Eyetech Pharmaceuticals and Alcon Laboratories. Reprint requests: Donald J. D’Amico, MD, Department of Ophthalmology, Weill Cornell Medical Center, 525 East 68th Street, Room F-835, New York, NY 10021; email: [email protected]
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Fig. 1. Fundus photographs and fluorescein angiogram of a 13-year-old girl diagnosed during a routine examination with a rhegmatogenous retinal detachment of long duration in the right eye. Left, Extensive retinal neovascularization was seen at the inferior equator of the right eye. Middle, Fluorescein angiogram of the right eye demonstrating fluorescein leakage from sites of retinal neovascularization. Right, Fundus photograph of the left eye showing peripheral retina changes consistent with mild retinopathy of prematurity.
to proliferative diabetic retinopathy, we found that VEGF and especially erythropoietin levels were elevated in this patient. Case Report A 13-year-old girl with a history of mild retinopathy of prematurity and high myopia was referred to the Retina Service at the Massachusetts Eye and Ear Infirmary (Boston) for evaluation of a retinal detachment found incidentally during an eye examination. Approximately 2.5 years earlier, the visual acuity was 20/30 in each eye with correction, and the refractive error was ⫺8.75 diopters for the right eye and ⫺12.25 diopters for the left eye. At that time, fundus examination did not reveal evidence of retinal detachment or neovascularization. She presented to the Retina Service with visual acuity of 20/200⫹2 in the right eye and 20/30 in the left eye. Examination of the right eye revealed a rhegmatogenous retinal detachment that extended from the 1:30-o’clock position through the 11-o’clock position. The detachment involved the macula and had signs of chronicity, including an inferior retina
that was extremely atrophic (Fig. 1, left). There was striking neovascularization at the inferior equator within the retina, which was confirmed by a fluorescein angiogram (Fig. 1, middle). Preretinal membranes and subretinal bands were also present. Examination of the left eye was notable for a posterior vitreous detachment and peripapillary atrophy. Both eyes had peripheral changes consistent with mild retinopathy of prematurity (Fig. 1, right). The patient underwent surgery to repair the detachment. Postoperatively, the retina remained reattached, although vision remained unchanged at 20/200.
Materials and Methods At the time of retinal surgery, undiluted vitreous specimens were acquired from this patient and five other patients (Table 1) who served as controls. All patients provided written informed consent. The study was conducted in accordance with HIPAA regulations and was approved by the Massachusetts Eye and Ear
Table 1. Patient Characteristics CRD Diagnosis
Age (y) Sex Diabetes history
PDR1
Proliferative Chronic diabetic retinal retinopathy detachment with vitreous with neovashemorrhage cularization
13 F —
PDR2
Proliferative diabetic retinopathy with clinically significant macular edema, vitreous hemorrhage, and tractional retinal detachment 58 56 M M 20-y history of 44-y history of diabetes type 1 currently diabetes managed with mellitus; insulin; previous previous panretinal panretinal photocoagulation photocoagulation of both eyes of both eyes
ARD Acute retinal detachment (⬍48 h) with multiple horseshoe tears superiorly
45 F —
ERM
FLO
Epiretinal Visually membrane significant floaters with a history of cataract surgery and YAG laser capsulotomy; mild epiretinal membrane 66 51 F M — —
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Fig. 2. Vitreous concentrations of erythropoietin and vascular endothelial growth factor (VEGF) as measured by enzyme linked immunosorbent assays (ELISAs). CRD, chronic retinal detachment; PDR, proliferative diabetic retinopathy; ARD, acute retinal detachment; ERM, epiretinal membrane; FLO, visually significant floaters. See Table 1 for further clinical data regarding these patients. Individual patient samples were measured in duplicates. Each of the data points (circles and squares) and the averages (horizontal line) are shown. Top Left, The patient with neovascularization from a chronic retinal detachment had the highest level of erythropoietin. Top Right, This patient’s VEGF level was not as high as those of the patients with proliferative diabetic retinopathy. Patient samples CRD, PDR1, and FLO were measured by ELISA 1 (triangles) and PDR2, ERM, and ARD were measured by ELISA 2 (diamonds) for the erythropoietin and VEGF assays. Bottom Left, Bottom Right, The standard curves for each of these ELISAs are shown.
Infirmary Institutional Review Board. The vitreous samples were immediately placed in heparinized tubes and stored at ⫺80°C. Enzyme linked immunosorbent assays (ELISAs) for erythropoietin and VEGF were performed according to the manufacturer’s instructions (Quantikine Human Epo Immunoassay and Human VEGF Immunoassay; R&D Systems, Inc., Minneapolis, MN). This VEGF ELISA measures the levels of the soluble isoforms (VEGF165 and VEGF121); however, it does not measure VEGF189 or VEGF206.5 For both erythropoietin and VEGF assays, three patient samples were run in parallel. Patient samples CRD, PDR1, and FLO were measured by ELISA 1 and PDR2, ERM, and ARD were measured by ELISA 2 for the erythropoietin and VEGF assays (see Table 1 for patient abbreviations). Individual patient samples were run in duplicate on each ELISA plate. Standard curves for each ELISA were constructed using the linear best line fit function. Results The vitreous of the patient with neovascularization secondary to the chronic retinal detachment had an erythropoietin level of 133 mIU/mL (Fig. 2, top left). Surprisingly, this level of erythropoietin was even higher than those of the two patients with proliferative diabetic ret-
inopathy. In contrast, the erythropoietin levels were low for the patients with no neovascularization. The patient with neovascularization secondary to the chronic retinal detachment also had a mildly elevated vitreous concentration of VEGF165 (77 pg/mL) compared with the patients with no neovascularization. However, this level of VEGF was not as high as the VEGF levels of the patients with proliferative diabetic retinopathy (Fig. 2, top right). Discussion We have shown that the vitreous concentrations of erythropoietin and VEGF are elevated in an adolescent patient with neovascularization secondary to a rhegmatogenous retinal detachment of long duration. Although elevated VEGF levels have been found in patients with a chronic retinal detachment,1 we are unaware of any previous report of an elevated erythropoietin level in such a case. The data demonstrate that the association of erythropoietin with retinal neovascularization is not limited to proliferative diabetic retinopathy. Surprisingly, the patient’s erythropoietin level in vitreous was even higher than those of the patients with proliferative diabetic retinopathy, while the level of VEGF was not as elevated compared with these patients. It has been previously reported that
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erythropoietin and VEGF are independent angiogenic factors in neovascularization.3 It is feasible that erythropoietin may be mechanistically more important than VEGF in the pathogenesis of some disorders that involve angiogenesis, such as chronic retinal detachment. If this were the case, then VEGF-blocking therapies alone would have limited success in preventing neovascularization in such a patient. Nevertheless, this conclusion cannot be made definitively from our data because it is not known how these varying concentrations of erythropoietin relate to stimulation of its receptor. One should also consider that erythropoietin is capable of playing several roles involving the retina. In addition to its angiogenic activity, erythropoietin is known to have protective effects for retinal cells by inhibiting apoptosis.6 In this case of a chronic retinal detachment, it is possible that erythropoietin may have roles in the prevention of apoptosis as well as neovascularization. Future studies will be necessary to elucidate the function of erythropoietin in retinal detachments and its relation to VEGF.
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2008
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VOLUME 2
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NUMBER 2
Key words: erythropoietin, neovascularization, retinal detachment, vascular endothelial growth factor. References 1.
2.
3.
4.
5.
6.
Aiello LP, Avery RL, Arrigg PG, et al. Vascular endothelial growth factor in ocular fluid of patients with diabetic retinopathy and other retinal disorders. N Engl J Med 1994;331:1480– 1487. Aiello LP, Pierce EA, Foley ED, 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 1995;92:10457–10461. Watanabe D, Suzuma K, Matsui S, et al. Erythropoietin as a retinal angiogenic factor in proliferative diabetic retinopathy. N Engl J Med 2005;353:782–792. Bonnet M. Peripheral neovascularization complicating rhegmatogenous retinal detachments of long duration. Graefes Arch Clin Exp Ophthalmol 1987;225:59–62. Funatsu H, Yamashita H, Sakata K, et al. Vitreous levels of vascular endothelial growth factor and intracellular adhesion molecule 1 are related to macular edema. Ophthalmology 2005;112:806–816. Becerra SP, Amaral J. Erythropoietin—an endogenous retinal survival factor. N Engl J Med 2002;347:1968–1970.
