title: RPE cells modulate subretinal neovascularization, but do not cause regression in mice with sustained expression of VEGF.
abstract: OBJECTIVE
  Previous studies using models of choroidal neovascularization (CNV) in which the angiogenic stimulus is not sustained, have concluded that the retinal pigmented epithelium (RPE) causes regression of neovascularization (NV). However, the withdrawal of angiogenic stimuli may actually be the major modulator of NV, and RPE cells may simply be responding to withdrawal of the angiogenic stimuli or something released by NV because of the withdrawal. In this study, the long-term course of NV and the behavior of the RPE in rhodopsin/VEGF transgenic mice, in which there is a sustained angiogenic stimulus, was investigated.
  
  
  METHODS
  Hemizygous mice from the V-6 line were killed at 0.75, 1, 3, 6, and 12 months after birth, and at each time point mRNA for VEGF, VEGF-R1, and VEGF-R2 was measured by RT-PCR. Some mice were perfused with fluorescein-labeled dextran and retinal flatmounts were examined by fluorescence microscopy. Light and electron microscopy was performed on Epon-embedded eyes.
  
  
  RESULTS
  The mRNA levels for VEGF, VEGF-R1, and VEGF-R2 remained constant from the earliest to the latest time point. Retinal flatmounts showed numerous small areas of subretinal NV at 3 weeks and at 1 month, and there were a similar number of larger lesions. By 6 months, many of the individual NV lesions had grown together to form large networks of new vessels. At 12 months, NV networks were similar to those at 6 months, but some of the vessels were not perfused. Light microscopy showed serous retinal detachments overlying NV lesions in mice up to 3 months of age, but at 6 and 12 months, the RPE completely surrounded new vessels and formed tight junctions to reestablish the outer blood-retinal barrier, and there were no serous detachments. Electron microscopy showed that compared with more acute NV lesions, chronic lesions contained thinner endothelial cells, similar to those of the choriocapillaris in that they had scant cytoplasm and numerous fenestrations, or pinocytotic vesicles with thick basement membrane surrounded by extracellular matrix (ECM). Bruch's membrane remained intact.
  
  
  CONCLUSIONS
  Despite persistent high expression of VEGF and its receptors, NV stopped growing and reached a plateau in older V-6 mice. RPE cells modulated the NV by surrounding it and reestablishing the blood-retinal barrier, but did not cause regression, although some vessels in chronic lesions were not perfused. These data do not support the conclusion of several previously reported studies, that RPE cells cause regression of CNV.
authors.affiliation: Department of Ophthalmology, Kansai Medical University, Moriguchi, Osaka, Japan.
authors.forename: Hisashi
authors.initials: H
authors.lastname: Ida
authors.forename: Takao
authors.initials: T
authors.lastname: Tobe
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Nambu
authors.forename: Miyo
authors.initials: M
authors.lastname: Matsumura
authors.forename: Masanobu
authors.initials: M
authors.lastname: Uyama
authors.forename: Peter A
authors.initials: PA
authors.lastname: Campochiaro
mesh_terms: D000818:Animals; D001813:Blood-Retinal Barrier; D003911:Dextrans; D016650:Fluorescein-5-isothiocyanate / Q000031:analogs & derivatives*; D005786:Gene Expression Regulation / Q000502:physiology*; D051379:Mice; D008822:Mice, Transgenic; D008856:Microscopy, Fluorescence; D010857:Pigment Epithelium of Eye / Q000502:physiology* / Q000648:ultrastructure; D012333:RNA, Messenger / Q000378:metabolism; D015861:Retinal Neovascularization / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D012171:Retinal Vessels / Q000378:metabolism* / Q000648:ultrastructure; D020133:Reverse Transcriptase Polymerase Chain Reaction; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics*; D040281:Vascular Endothelial Growth Factor Receptor-1 / Q000235:genetics; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000235:genetics
chemical_list: D003911:Dextrans; D012333:RNA, Messenger; D042461:Vascular Endothelial Growth Factor A; C015219:fluorescein isothiocyanate dextran; D040281:Vascular Endothelial Growth Factor Receptor-1; D040301:Vascular Endothelial Growth Factor Receptor-2; D016650:Fluorescein-5-isothiocyanate