00006982-200110000-00009

7/27/2019 00006982-200110000-00009

http://slidepdf.com/reader/full/00006982-200110000-00009 1/9

RELATIONSHIP BETWEEN VASCULAR ENDOTHELIAL GROWTH FACTOR ANDINTERLEUKIN–6 IN DIABETICRETINOPATHY

HIDEHARU FUNATSU, MD,* HIDETOSHI YAMASHITA, MD,† ERIKA SHIMIZU, MD,*

RIE KOJIMA, MD,* SADAO HORI, MD‡

Purpose: To determine the relationship between the severity of diabetic retinopathy and

the levels of interleukin-6 (IL-6) and vascular endothelial growth factor (VEGF) in aqueous

humor and plasma.

Methods: Forty-four eyes of 34 diabetic patients were studied. The concentrations of

VEGF and IL-6 in plasma samples and in aqueous specimens obtained from the eyes

during cataract surgery were measured by enzyme-linked immunosorbent assay.

Results: Aqueous levels of VEGF and IL-6 were significantly correlated with the severity

of diabetic retinopathy ( ϭ 0.793 and ϭ 0.744, respectively). Vascular endothelial growth

factor and IL-6 levels in aqueous humor were significantly correlated with the aqueous

protein concentration ( ϭ 0.641 and ϭ 0.646, respectively). The aqueous level of VEGF

was significantly correlated with that of IL-6 ( ϭ 0.627). Aqueous levels of VEGF and IL-6

were also significantly correlated with the grade of fundus findings. Vascular endothelialgrowth factor and IL-6 concentrations were higher in the aqueous than in the plasma.

Conclusion: The results of the current study suggest that there is a relationship

between VEGF and IL-6 but the role of IL-6 in diabetic retinopathy is unclear and may

warrant further investigation.

RETINA 21:469 –477, 2001

In the pathogenesis of diabetic retinopathy, chronic

hyperglycemia on various metabolic pathwayscauses breakdown of the vascular barrier and occlu-

sion of retinal vessels.1 Retinal edema, hard exudates,

and hemorrhages all develop after breakdown of the

vascular barrier. In addition, retinal endothelial dam-

age, blood coagulation abnormalities, and severe ret-

inal edema cause retinal capillary obstruction and

thereafter cause retinal ischemia.2 Clinical observa-tions have revealed that vitreoretinal neovasculariza-

tion is always followed by severe retinal ischemia. In

proliferative diabetic retinopathy, proliferative mem-

branes are formed by extracellular matrix and new

vessels, leading to tractional retinal detachment and

vitreous hemorrhage.

Many cytokines and growth factors, including basic

fibroblast growth factor (bFGF),3 insulin-like growth

factor-1,4 vascular endothelial growth factor (VEGF),5–7

and interleukin-6 (IL-6),8,9 are involved in the pathogen-

From the *Department of Ophthalmology, Diabetes Center, To-kyo Women’s Medical University; †Department of Ophthalmol-ogy, School of Medicine, Yamagata University; and ‡Departmentof Ophthalmology, Tokyo Women’s Medical University, Japan.

This study was supported by Health Science Research grants(#10060101, Drs. Funatsu, Hori, and Yamashita) from the Ministryof Health and Welfare, Research on Eye and Ear Sciences, Immu-nology, Allergy and Organ Transplantation, Japan.

This paper was presented at annual meeting of the Associationfor Research in Vision and Ophthalmology (ARVO); Fort Lauder-dale, Florida; May 3, 2000.

Reprint requests: Hideharu Funatsu, MD, 8-1 Kawada-cho,Shinjuku-ku, Tokyo 162-8666, Japan; e-mail: [email protected]

469

7/27/2019 00006982-200110000-00009


esis of diabetic retinopathy. Vascular endothelial growthfactor is an endothelial cell mitogen that induces an

increase of vascular permeability and angiogenesis.10–13

Vascular endothelial growth factor was localized in nu-

merous retinal cells including retinal pigment epithelialcells, pericytes, endothelial cells, Müller cells, and astro-

cytes.14–19 Vascular endothelial growth factor has beenrevealed to play a central role in the progression of

diabetic retinopathy.9,20,21 However, the pathogenesis of diabetic retinopathy cannot be explained by the actions

of VEGF alone. We previously reported that transform-ing growth factor-␤ (TGF-␤) and IL-6 are also involved

in the pathogenesis of diabetic retinopathy.22 These cy-tokines and growth factors form a network that influ-

ences the progression of diabetic retinopathy. Analysis of this network is important to clarify the pathogenesis of

diabetic retinopathy and to develop valuable therapies.Interleukin-6 is a multifunctional cytokine and may

be a major mediator of anterior uveitis23–27

and pro-liferative vitreoretinopathy.28–30 Interleukin-6 is syn-

thesized by a variety of cells, including fibroblasts,macrophages, epidermal cells, synovial cells, vascular

smooth muscle, and vascular endothelium.31,32 Withinthe eye, the sources of IL-6 include the retinal pigment

epithelial cells, corneal epithelial cells, keratocytes,iris, and ciliary body.33–37 Interleukin-6 was previ-

ously reported to be related to hyperglycemia anddiabetic nephropathy.38 In addition, IL-6 is considered

to be an indirect inducer of angiogenesis that exerts itsactivity through the induction of VEGF.39

In this study, to investigate the relationship betweenVEGF and IL-6 in diabetic retinopathy, we measured

the concentrations of VEGF and IL-6 in the samespecimens simultaneously. The current study revealed

that the aqueous levels of VEGF and IL-6 are signif-

icantly correlated with the severity of diabetic retinop-athy and suggested that both VEGF and IL-6 acted in

the pathogenesis of diabetic retinopathy.

Subjects and Methods

Subjects

Paired samples of undiluted aqueous humor andplasma were obtained from 34 diabetic patients and 16

nondiabetic patients who underwent cataract surgery(Table 1). The samples were collected from 44 eyes of

diabetic patients and 16 eyes of nondiabetic patients.The mean age of the patients with diabetes mellitus

was 67.4 years (range, 36–77 years) and that of thosewithout diabetes mellitus was 72.8 years (range,

54–81 years). The mean duration of diabetes mellituswas 17.9 years (range, 1–32 years). The protocol for

sample collection was approved by the institutional

review board and all patients gave their informedconsent. Exclusion criteria included prior ocular sur-

gery, a history of intraocular inflammation, and ahistory of intraocular ischemia due to causes other

than diabetic retinopathy.

Fundus Findings

The preoperative and operative fundus findings

were recorded. Severity of diabetic retinopathy wasconfirmed by standardized fundus color photography

and fluorescein angiography (FA) with a TopconTRC-50IA fundus camera and an image-net system

(Tokyo Optical Co Ltd, Japan). Ten overlapping, non-stereoscopic 50° photographs were taken of each eye.

Eyes were examined immediately after surgery and upto 48 hours after surgery until it was possible to

determine the severity of retinopathy. The severity of diabetic retinopathy was graded according to the mod-

ified Early Treatment Diabetic Retinopathy Study(ETDRS) retinopathy severity scale.40,41 For the grad-

ing of fundus findings, retinal hemorrhages, hard ex-

Table 1. Clinical and Laboratory Characteristicsof the Subjects

CharacteristicNo. (%) or (Mean Ϯ SD)

(Total n ϭ 34)

GenderFemale 17 (50.0)

Male 17 (50.0) Age, yr 34 (57.4 Ϯ 9.8)Duration of DM, yr 34 (13.9 Ϯ 9.4)Treatment

Diet 3 (8.8)OHA 10 (29.4)Insulin 21 (61.8)

HbA1c (%) 34 (7.3 Ϯ 1.5)Smoking history

Absent 16 (47.1)Present 18 (52.9)

Alcohol consumption Absent 21 (61.8)Present 13 (38.2)

Hypertension Absent 17 (50.0)Present 17 (50.0)

Systolic blood pressure, mmHg 34 (132.5 Ϯ 18.0)Diastolic blood pressure, mmHg 34 (68.8Ϯ 8.4)Body mass index, kg/m2* 34 (23.1 Ϯ 3.2)Severity of nephropathy

Normal 2 (5.9)Mild (albuminuria) 12 (35.3)Moderate (proteinuria) 20 (58.8)

* The body mass index is expressed as the weight in kilogramsdivided by the square of the height in meters.

SD, standard deviation; DM, diabetes mellitus; OHA, oral hy-poglycemic agents.

470 RETINA, THE JOURNAL OF RETINAL AND VITREOUS DISEASES ● 2001 ● VOLUME 21 ● NUMBER 5

7/27/2019 00006982-200110000-00009


udates, soft exudates, intraretinal microvascular ab-normalities (IRMA), venous beading, venous loops,

new vessels elsewhere (NVE), new vessels on orwithin 1 disk diameter of the disk (NVD), fibrous

proliferation elsewhere (FPE), and vitreous hemor-rhage were graded according to the ETDRS retinop-

athy grading system.40,41 Each lesion’s severity wasgraded according to the ETDRS severity scale40 and

the graded method was modified so that the averagelesion severity scale of each photograph was used as

the lesion severity and the retinopathy severity levelwas graded according to the ETDRS severity scale.41

Two graders independently assessed the diabetic ret-inopathy from photographs.

Sample Collection

At the beginning of eye surgery, a sample of undi-

luted aqueous humor (0.1–0.2 mL) was manuallyaspirated into a disposable tuberculin syringe and

transferred immediately to a sterile tube, before turn-ing on the infusion and completing the surgical pro-

cedure. The samples were stored in a deep freezer atϪ80 °C until assay.

Plasma samples were also collected from the 34patients. Blood samples were immediately placed on

ice, clarified by centrifugation at 3,000 ϫ g for 5minutes at 4 °C, and rapidly frozen at Ϫ80 °C until

assay. The institutional review board approved theprotocol for sample collection.

Analysis of VEGF and IL-6

The concentrations of VEGF and IL-6 were mea-sured by enzyme-linked immunosorbent assay

(ELISA) using human VEGF and IL-6 immunoassays(R&D Systems, Minneapolis, MN).42,43 For ELISA,

standard solution (100 L for VEGF, 200 L forIL-6) or sample (10 or 100 L for VEGF, 20 or 200

L for IL-6) were added to a 96-well plate filled withsolidified monoclonal antibody. After incubation, the

plate was washed, and enzyme-labeled antibody wasadded. After incubation, the plate was washed and

substrate was added. The reaction was stopped uponcolor development by adding stop solution. Optical

density was determined at 450 and 620 nm using anabsorption spectrophotometer (Titertek Multiscan

MCC/340; ICN, Tokyo, Japan). The standard curvewas plotted from the measurements made with the

standard solution (from 15.6–1000 pg/mL for VEGF,from 0.156–10 pg/mL for IL-6), and the concentration

of VEGF or IL-6 in the sample was determined. Wemeasured only two types of cytokines because of the

small sample volume of aqueous. The VEGF kit used

could detect two of the four VEGF isoforms, VEGF121

and VEGF165, and the assay was performed according

to the manufacturer’s instructions. The levels of thesefactors in aqueous humor and plasma were within the

detection range of the assays, with the minimum de-tectable concentration being 15.6 pg/mL for VEGF

and 0.156 pg/mL for IL-6.

Measurements of Aqueous Protein Concentration

Thirty minutes after dilation of the pupils of both eyes

with 1% tropicamide, the flare value was measured witha laser flare-cell meter (FC1000, Kowa Co Ltd, Tokyo,

Japan) within 48 hours before surgery. Aqueous flare

(photon counts) correlated with albumin concentrationusing the regression correlation between photon counts

and bovine albumin concentration.44,45

Statisitical Analysis

Analyses were performed with SAS software (SAS

Institute Inc, Cary, NC).46 Results were presented asthe mean Ϯ SD or geometric mean Ϯ SD on a loga-

rithmic scale. To assess the relationship between eachcytokine and the ETDRS scale, Spearman’s rank-

order correlation coefficients with 95% confidenceintervals were calculated. The significance of differ-

ences between groups was evaluated by the Kruskal-Wallis test. Two-tailed P values of less than 0.05 were

considered to indicate statistical significance.

Results

Severity of Retinopathy and Aqueous Cytokines

The VEGF level in aqueous humor ranged from26.5 to 1,177.0 pg/mL (mean, 215.0 Ϯ 246.8) and the

aqueous level of VEGF was significantly correlatedwith the severity of diabetic retinopathy ( ϭ 0.793,

PϽ 0.0001, Figure 1A). The IL-6 level in aqueoushumor ranged from 3.67 to 370.0 pg/mL (mean,

69.3Ϯ 85.1) and the aqueous level of IL-6 was alsosignificantly correlated with the severity of diabetic

retinopathy ( ϭ 0.744, P Ͻ 0.0001, Figure 1B). The

VEGF and IL-6 levels in aqueous humor of diabeticpatients were significantly higher than those of non-diabetic patients (P Ͻ 0.01, P Ͻ 0.01, respectively)

(Table 2). Aqueous levels of VEGF were significantlycorrelated with those of IL-6 ( ϭ 0.627, PϽ 0.0001,

Figure 2).

Fundus Findings and Aqueous Cytokines

Among the fundus findings, the aqueous levels of VEGF and IL-6 were significantly correlated with the

grade of retinal hemorrhages, hard exudates, soft ex-

471VEGF AND IL-6 IN DIABETIC RETINOPATHY • FUNATSU ET AL

7/27/2019 00006982-200110000-00009


7/27/2019 00006982-200110000-00009


7/27/2019 00006982-200110000-00009


the blood–aqueous barrier. Diabetes has been reported

to affect the iridial blood–aqueous barrier in rats.60 Ithas been reported that the aqueous protein concentra-

tion, measured by the flare intensity, increases withthe severity of diabetic retinopathy.61,62 Vascular en-

dothelial growth factor is known to induce hyperper-meability of microvessels and breakdown of the BRB

in nonproliferative diabetic retinopathy.49,50,63 Inter-leukin-6 is produced by retinal pigment epithelial

cells, corneal epithelial cells, keratocytes, iris, and the

ciliary body in the eye.33–37 Aqueous IL-6 levels arereported to be elevated in patients with uveitis, endo-

toxin-induced uveitis, and PVR,23–30 suggesting thatthe elevation of IL-6 in aqueous humor is due to local

production. Our results and these previous reportssuggest that the increase of IL-6 levels in aqueous

humor may be related to the breakdown of the BRBand/or the production of IL-6 in ocular tissues such as

retinal pigment epithelial cells, the iris, and the ciliarybody.

Aqueous levels of VEGF and IL-6 were also sig-nificantly correlated with fundus findings such as ret-

inal hemorrhages, hard and soft exudates, IRMA, ve-nous abnormalities, NVE, and vitreous hemorrhage.

Tolentino and associates found that bioactive VEGF-injected eyes developed dilated and tortuous vessels,

venous beading, retinal edema, microaneurysms, in-traretinal hemorrhages, and capillary closure with

ischemia.61

Interleukin-6 induces disruption of theblood– brain barrier.64,65 Although VEGF appears to

be able to produce intraretinal neovascularization, itmight not be sufficient to produce extraretinal neovas-

cularization across the internal limiting membrane.61

In the current study, the VEGF and IL-6 levels in

aqueous humor were correlated with the severity of retinal hemorrhage and hard exudates. Therefore, both

VEGF and IL-6 are implicated in the breakdown of the BRB in nonproliferative diabetic retinopathy. Fur-

thermore, the VEGF and IL-6 levels in aqueous humorwere correlated with the severity of NVE, so both

cytokines are also implicated in the pathogenesis of neovascularization in the proliferative stage. A recent

study indicated that there was a causal relationship

between IL-6 and VEGF in neovacularization.22,39

Even though VEGF alone is sufficient to produce iris

neovascularization in nonhuman primates after intra-vitreal injection, it does not rule out the possibility that

other growth factors may also be involved in intraoc-ular angiogenesis.22,53,59,66– 68 Cohen and associates

suggested that IL-6 may induce angiogenesis indi-rectly by stimulating VEGF expression, a hypothesis

consistent with the documented induction of VEGFmRNA by hypoxia.39 These reports and our results

suggest that VEGF is directly involved in the patho-genesis of both nonproliferative and proliferative di-

abetic retinopathy. Conversely, IL-6 may have a directrole in nonproliferative retinopathy and an indirect

role in the proliferative stage via VEGF. The relation-ship between VEGF and IL-6 may also be important

in the pathogenesis of diabetic retinopathy. Furtherinvestigations are required to clarify the clinical sig-

nificance of synergistic correlation between VEGFand IL-6 in the progression of diabetic retinopathy

according to follow-up diabetic patients.

Fig. 3. A, The correlation between the aqueous level of vascular

endothelial growth factor (VEGF) and the protein concentration. Theaqueous level of VEGF was significantly correlated with the proteinconcentration ( ϭ 0.64, P Ͻ 0.0001). B, The correlation between the

aqueous level of interleukin (IL)-6 and the protein concentration. Theaqueous level of IL-6 was significantly correlated with the protein

concentration ( ϭ 0.65, P Ͻ 0.0001).


7/27/2019 00006982-200110000-00009


7/27/2019 00006982-200110000-00009


growth factor by human retinal pigment epithelial cells. Bio-

chem Biophys Res Commun 1993;193:631–638.

15. Pe’er J, Shweiki D, Itin A, et al. Hypoxia-induced expression

of vascular endothelial growth factor by retinal cells is a

common factor in neovascularizing ocular diseases. Lab In-

vest 1995;72:638–645.

16. Pierce EA, Avery RL, Foley ED, et al. Vascular endothelial

growth factor/vascular permeability factor expression in amouse model of retinal neovascularization. Proc Natl Acad

Sci USA 1995;92:905–909.

17. Lutty GA, McLeod DS, Merges C, et al. Localization of

vascular endothelial growth factor in human retina and cho-

roid. Arch Ophthalmol 1996;114:971–979.

18. Dorey CK, Aouididi S, Reynaud X, et al. Correlation of

vascular permeability factor/vascular endothelial growth fac-

tor with extraretinal neovascularization in the rat. Arch Oph-

thalmol 1996;114:1210–1217.

19. Gerhardinger C, Brown LF, Roy S, et al. Expression of

vascular endothelial growth factor in the human retina and in

nonproliferative diabetic retinopathy. Am J Pathol 1998;152:

1453–1462.

20. Shweiki D, Itin A, Soffer D, Keshet E. Vascular endothelialgrowth factor induced by hypoxia may mediate hypoxia-

initiated angiogenesis. Nature 1992;359:843–845.

21. Aiello LP, Northrup JM, Keyt BA, et al. Hypoxic regulation

of vascular endothelial growth factor in retinal cells. Arch

Ophthalmol 1995;113:1538–1544.

22. Ideta R, Yamashita H, Tanaka Y, et al. Roles of cytokines in

diabetic retinopathy. Arch Ophthalmol 1999;117:700–701.

23. Murray PI, Hoekzema R, van Haren MAC, et al. Aqueous

humor interleukin-6 levels in uveitis. Invest Ophthalmol Vis

Sci 1990;31:917–920.

24. Hoekzema R, Murray PI, van Haren MAC, et al. Analysis of

interleukin-6 in endotoxin-induced uveitis. Invest Ophthal-

mol Vis Sci 1991;32:88–95.

25. Hoekzema R, Verhagen C, van Haren M, Kijlstra A. Endo-

toxin-induced uveitis in the rat. The significance of intraoc-

ular interleukin-6. Invest Ophthalmol Vis Sci 1992;33:532–

539.

26. Planck SR, Huang XN, Robertson JE, Rosenbaum JT. Cyto-

kine mRNA levels in rat ocular tissues after systemic endo-

toxin treatment. Invest Ophthalmol Vis Sci 1994;35:924 –

930.

27. DeVos AF, Klaren VNA, Kijlstra A. Expression of multiple

cytokines and IL-1RA in the uvea and retina during endo-

toxin-induced uveitis in the rat. Invest Ophthalmol Vis Sci

1994;35:3873–3883.

28. Limb GA, Little BC, Meager A, et al. Cytokines in prolifer-

ative vitreoretinopathy. Eye 1991;5:686– 693.

29. Kauffmann DJH, vanMeurs JC, Mertens DAE, et al. Cyto-

kines in vitreous humor: interleukin-6 is elevated in prolifer-

ative vitreoretinopathy. Invest Ophthalmol Vis Sci 1994;35:

900–906.

30. El-Asrar AMA, Van Damme J, Put W, et al. Monocyte

chemotactic protein-1 in proliferative vitreoretinal disorders.

Am J Ophthalmol 1997;123:599–606.

31. Hirano T, Akira S, Taga T, Kishimoto T. Biological and

clinical aspects of interleukin 6. Immunol Today 1990;11:

443–449.

32. Lotz M. Interleukin-6. A comprehensive review. Cancer

Treat Res 1995;80:209 –233.

33. Elner VM, Scales W, Elner SG, et al. Interleukin-6 (IL-6)

gene expression and secretion by cytokine-stimulated human

retinal pigment epithelium cells. Exp Eye Res 1992;54:361–

368.

34. Planck SR, Dang TT, Graves D, et al. Retinal pigment

epithelium cells secrete interleukin-6 in response to interleu-

kin-1. Invest Ophthalmol Vis Sci 1992;33:78–82.

35. Benson MT, Shepherd L, Rees RC, Rennie IG. Production of

interleukin-6 by human retinal pigment epithelium in vitro

and its regulation by other cytokines. Curr Eye Res 1992;11:173–179.

36. Cubitt CL, Lausch RN, Oakes JE. Differences in interleu-

kin-6 gene expression between cultured human corneal epi-

thelial cells and keratocytes. Invest Ophthalmol Vis Sci 1995;

36:330–336.

37. Knisely TL, Grabbe S, Nazareno R, Granstein RD. Produc-

tion of interleukin-6 and granulocyte-macrophage colony-

stimulating factor by murine iris and ciliary body explants.

Invest Ophthalmol Vis Sci 1994;35:4015–4022.

38. Morohoshi M, Fujisawa K, Uchimura I, Numano F. Glucose-

dependent interleukin 6 and tumor necrosis factor production

by human peripheral blood monocyte in vitro. Diabetes 1996;

45:954–959.

39. Cohen T, Nahari D, Cerem LW, et al. Interleukin 6 induces

the expression of vascular endothelial growth factor. J Biol

Chem 1996;271:736–741.

40. Early Treatment Diabetic Retinopathy Study Research

Group. Grading diabetic retinopathy from stereoscopic color

fundus photographs. An extension of the modified Airlie

House classification. ETDRS Report Number 10. Ophthal-

mology 1991;98:786 – 806.

41. Early Treatment Diabetic Retinopathy Study Research

Group. Fundus photographic risk factors for progression of

diabetic retinopathy. ETDRS Report Number 12. Ophthal-

mology 1991;98:823–833.

42. Hyodo I, Doi T, Endo H, et al. Clinical significance plasma

vascular endothelial growth factor in gastrointestinal cancer.

Eur J Cancer 1998;34:2041–2045.

43. Grey A, Mitnick MA, Shapses S, et al. Circulating levels of interleukin-6 and tumor necrosis factor are elevated in pri-

mary hyperparathyroidism and correlate with markers of

bone resorption. A clinical research center study. J Clin

Endocrinol Metab 1996;81:3450–3454.

44. Sawa M. Clinical application of laser flare-cell meter. Jpn J

Ophthalmol 1990;34:346–363.

45. Oshika T, Araie M. Time course of changes in aqueous

protein concentration and flow rate after oral acetazolamide.

Invest Ophthalmol Vis Sci 1990;31:527–534.

46. SAS Institute Inc. SAS/STAT software. Release 6.12. Cary,

NC: SAS Institute Inc, 1990.

47. Bresnick GH. Nonproliferative diabetic retinopathy. In: Ryan

SJ, ed. Retina. St. Louis: Mosby, 1994;1277–1318.

48. Kohner EM. Retinal ischemia. In: Ryan SJ, ed. Retina. St.

Louis: Mosby, 1994;1009–1018.

49. Cunha-Vaz J, de Abreu JRF, Campos AJ. Early breakdown of

the blood-retinal barrier in diabetes. Br J Ophthalmol 1975;

59:649–656.

50. Cunha-Vaz JG, Fonseca JR, Abreu JF, Ruas MA. A fol-

low-up study by vitreous fluorophotometry of early retinal

involvement in diabetes. Am J Ophthalmol 1978;86:467–

473.

51. Murata T, Ishibashi T, Khalil A, et al. Vascular endothelial

growth factor plays a role in hyperpermeability of diabetic

retinal vessels. Ophthalmic Res 1995;27:48–52.

52. Murata T, Nakagawa K, Khalil A, et al. The relation between

expression of vascular endothelial growth factor and break-


7/27/2019 00006982-200110000-00009


down of the blood-retinal barrier in diabetic rat retinas. Lab

Invest 1996;74:819 – 825.

53. Amin EH, Frank RN, Kennedy A, et al. Vascular endothelial

growth factor is present in glial cells of the retina and optic

nerve of human subjects with nonproliferative diabetic reti-

nopathy. Invest Ophthalmol Vis Sci 1997;38:36–47.

54. Mathews MK, Merges C, McLeod DS, Lutty GA. Vascular

endothelial growth factor and vascular permeability changesin human diabetic retinopathy. Invest Ophthalmol Vis Sci

1997;38:2729–2741.

55. Clermont AC, Aiello LP, Mori F, et al. Vascular endothelial

growth factor and severity of nonproliferative diabetic reti-

nopathy mediate retinal hemodynamics in vivo: a potential

role for vascular endothelial growth factor in the progression

of nonproliferative diabetic retinopathy. Am J Ophthalmol

1997;124:433–446.

56. Ambati J, Chalam KV, Chawla DK, et al. Elevated ␥ -ami-

nobutyric acid, glutamate, and vascular endothelial growth

factor levels in the vitreous of patients with proliferative

diabetic retinopathy. Arch Ophthalmol 1997;115:1161–1166.

57. Katsura Y, Okano T, Noritake M, et al. Hepatocyte growth

factor in vitreous fluid of patients with proliferative diabetic

retinopathy and other retinal disorders. Diabetes Care 1998;21:1759–1763.

58. D’Amore PA. Mechanisms of retinal and choroidal neovas-

cularization. Invest Ophthalmol Vis Sci 1994;35:3974 –3979.

59. Folkman J. Angiogenesis in cancer, vascular, rheumatoid and

other disease. Nature Med 1995;1:27–31.

60. Casey R, Li WW. Factors controlling ocular angiogenesis.

Am J Ophthalmol 1997;124:521–529.

61. Tolentino MJ, Miller JW, Gragoudas ES, et al. Intravitreous

injections of vascular endothelial growth factor produce ret-

inal ischemia and microangiopathy in an adult primate. Oph-

thalmology 1996;103:1820 –1828.

62. Ishibashi T, Tanaka K, Tanigchi Y. Disruption of iridial

blood-aqueous barrier in experimental diabetic rats. Graefes

Arch Clin Exp Ophthalmol 1982;219:159–164.

63. Oshika T, Kato S, Funatsu H. Quantitative assessment of

aqueous flare intensity in diabetes. Graefes Arch Clin ExpOphthalmol 1989;227:518–520.

64. Tolentino MJ, Miller JW, Gragoudas ES, et al. Vascular

endothelial growth factor is sufficient to produce iris neovas-

cularization and neovascular glaucoma in a nonhuman pri-

mate. Arch Opthalmol 1996;114:964–970.

65. Chen KH, Wu CC, Roy S, et al. Increased interleukin-6 in

aqueous humor of neovascular glaucoma. Invest Ophthalmol

Vis Sci 1999;40:2627–2632.

66. Moriarty AP, Spalton DJ, Moriarty BJ, et al. Studies of the

blood-aqueous barrier in diabetes mellitus. Am J Ophthalmol

1994;117:768–771.

67. Vinores SA, Gadegbeku C, Campochiaro PA, Green WR.

Immunohistochemical localization of blood-retinal barrier

breakdown in human diabetics. Am J Pathol 1989;134:231–235.

68. Miller JW, Adamis AP, Shima DT, et al. Vascular endothelial

growth factor/vascular permeability factor is temporally and

spatially correlated with ocular angiogenesis in a primate

model. Am J Pathol 1994;145:574–584.

69. de Vries HE, Blom-Roosemalen MCM, van Oosten M, et al.

The influence of cytokines on the integrity of the blood-brain

barrier in vitro. J Neuroimmunol 1996;64:37– 43.

477VEGF AND IL-6 IN DIABETIC RETINOPATHY • FUNATSU ET AL

Documents

00006982-200110000-00009