Curr Eye Res. 2017 Feb;42(2):237-243. doi: 10.1080/02713683.2016.1227448.

Subchoroidal Release of VEGF and bFGF Produces Choroidal Neovascularization in Rabbit.

Wong CG1, Taban M2, Osann K3, Ross-Cisneros FN4, Bruice TC5, Zahn G6, You T7.
1SCLERA LLC , Carlsbad , CA , USA.
2Department of Ophthalmology , College of Medicine, University of California Irvine , Irvine , CA , USA.
3Department of Medicine , College of Medicine, University of California Irvine , Irvine , CA , USA.
4Doheny Eye Institute , Los Angeles , CA , USA.
5University of California Los Angeles , Los Angeles , CA , USA.
6Consulting , Berlin , Germany.
7Orange County Retina Group , Santa Ana , CA , USA.

 

Abstract

Purpose: Intravitreal ascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) produced florid retinal neovascularization and hemorrhage in the rabbit. This study seeks to determine whether sustained subchoroidal release of both VEGF and bFGF can induce robust choroidal neovascularization (CNV) in the rabbit.

Methods: Subchoroidal implantation through the sclera of polymeric pellets containing both 15 ug VEGF and 15 ug bFGF was performed on adult pigmented male Dutch belted rabbits (N = 6) and NZW albinos (N = 8). As negative controls, blank pellets with no growth factors were implanted in both Dutch belted rabbits (N = 6) and NZW albino rabbits (N = 4). Development of CNV was documented weekly over a four-week period with indirect ophthalmoscopy, color fundus photography, and fluorescein angiography. Eyes were enucleated and prepared for histologic and immunohistochemical analyses at the end of the study. Amounts of VEGF and bFGF that were released in vitro from the pellets were measured by ELISA.

Results: In all eyes with subchoroidal implants containing both VEGF and bFGF, strong fluorescein leakage was observed at 2, 3, and 4 weeks (P < 0.005); no leakage was seen initially in week 1. Negative control groups with blank implants showed no fluorescein leakage throughout the four-week study period. Histologic analysis confirmed the presence of experimental CNV. New subretinal blood vessel growth occurred in all eyes with VEGF/bFGF implants. Negative control eyes with blank implants showed no vascular changes. In vitro sustained release of both VEGF and bFGF was confirmed by ELISA.

Conclusion: Sustained subchoroidal release of both VEGF and bFGF produced experimental CNV rapidly in the rabbit. Understanding how these growth factors induce CNV may suggest novel therapeutic strategies in the large rabbit eye.

Key words: Robust rabbit CNV model; basic fibroblast growth factor; vascular endothelial growth factor; subchoroidal sustained release; transscleral implantation

PMID: 27749103; DOI: 10.1080/02713683.2016.1227448

 

Supplement:

Both diabetic retinopathy (DR) and aging-related macular degeneration (AMD) are diseases of the retina. Both of these retinal diseases are due to abnormal leaky blood vessel formation, which is known as neovascularization (NV). With age and time, both diseases can lead to blindness. The two diseases differ only in the location of intraocular NV. The abnormal blood vessels may proliferate either along the surface of the retina leading to DR or underneath the retina leading to advanced wet AMD. There is also retinopathy of prematurity (ROP), which is the leading cause of visual loss in newborns.

 

DR is the major cause of new blindness in developed countries within the working age group. Moreover, AMD is the most common cause of severe visual loss in people over the age of 65. “Wet” or neovascular AMD occurs when neovascularization develops underneath the retina. This abnormal tissue causes physical separation of the retinal layers with destruction of the normal tissue. Almost all of the abnormal blood vessels leak and bleed at times. Loss of central vision occurs due to this rapid growth of abnormal leaking blood vessels. This wet neovascular form of AMD is characterized by choroidal neovascularization (CNV). With its abnormal growth of leaky blood vessels, CNV is the hallmark of wet AMD and is responsible for approximately 90% of cases with severe visual loss.

 

To determine the pathogenesis of wet AMD to facilitate early diagnostic detection and to test potential pharmacologic treatments, a pre-clinical experimental model of wet AMD with CNV must be developed in a large animal eye that is similar in size to the human eye so that potential drugs can be administered and the effects observed over a longer period of time. We have developed experimental rabbit models of both retinal NV and CNV for testing potential retinal therapeutics (1,2). The patented technology involves placing an intraocular implant that releases human growth factors in a sustained manner either intravitreally over the retina or subchoroidally beneath the retina. These implants induce the abnormal growth of leaky blood vessels in the rabbit eye over a short period of time to produce the experimental retinal diseases. Currently, we are determining whether biodegradable sustained release drug delivery implants can be developed for long-term delivery of drugs against retinal diseases.

 

In the rabbit eye, the two photographs (left and middle) below demonstrate intravitreal placement of our non-biodegradable implant containing both human growth factors VEGF and bFGF and a second implant that contains only saline. The third photo (far right) demonstrates a VEGF/bFGF-containing non-biodegradable implant that is producing retinal NV in the rabbit eye. Utilizing biodegradable implants, current studies with our experimental rabbit models of retinal NV and CNV are defining potential therapeutics that can stop the progression of these blinding retinal diseases.

 

 

References

  1. Grit Zahn, PhD; Dörte Vossmeyer, PhD; Roland Stragies, PhD; Margaret Wills, MS; Corinne G. Wong, PhD; Karin U. Löffler, MD; Anthony P. Adamis, MD; Jochen Knolle, PhD. Preclinical Evaluation of the Novel Small-Molecule Integrin α5β1 Inhibitor JSM6427 in Monkey and Rabbit Models of Choroidal Neovascularization. Arch Ophthal. 127(10):1329-35 (2009). Available online:

http://www.tandfonline.com/eprint/n9dasEpcwg7xvZzqqb9S/full

  1. Rabin D, Bruice TC, You TT, Wong CG. Both edema and leakage are expressed and are separable in long-term experimental CNV via sustained simultaneous re-lease of VEGF and bFGF within the suprachoroidal space through transscleral implantation of a delivery device [ARVO abstract 1374]. Invest Ophthalmol VisSci. 2008;49:e-abstract 1374.