Antiangiogenic and Neurogenic Activities ofSleeping Beauty-Mediated PEDF-Transfected RPE CellsIn VitroandIn Vivo

Abstract

Pigment epithelium-derived factor (PEDF) is a potent multifunctional protein that inhibits angiogenesis and has neurogenic and neuroprotective properties. Since the wet form of age-related macular degeneration is characterized by choroidal neovascularization (CNV), PEDF would be an ideal candidate to inhibit CNV and support retinal pigment epithelial (RPE) cells. However, its short half-life has precluded its clinical use. To deliver PEDF to the subretinal space, we transfected RPE cells with the PEDF gene using the Sleeping Beauty transposon system. Transfected cells expressed and secreted biologically active recombinant PEDF (rPEDF). In cultures of human umbilical vein endothelial cells, rPEDF reduced VEGF-induced cumulative sprouting by ≥47%, decreased migration by 77%, and increased rate of apoptosis at least 3.4 times. rPEDF induced neurite outgrowth in neuroblastoma cells and protected ganglion and photoreceptor cells in organotypic retinal cultures. In a rat model of CNV, subretinal transplantation of PEDF-transfected cells led to a reduction of the CNV area by 48% 14 days after transplantation and decreased clinical significant lesions by 55% and 40% after 7 and 14 days, respectively. We showed that transplantation of pigment epithelial cells overexpressing PEDF can restore a permissive subretinal environment for RPE and photoreceptor maintenance, while inhibiting choroidal blood vessel growth.

Engineering of PEDF-Expressing Primary Pigment Epithelial Cells by the SB Transposon System Delivered by pFAR4 Plasmids

“…Effective SB -mediated gene transfer has been reported for various in vitro and in vivo disease models,41, 42, 43 and it has been approved for clinical application of genetically modified T cells to treat B-lymphoid malignancies 44, 45. We have reported long-term and continuous PEDF secretion by RPE cells that were transfected with the PEDF transgene using the SB100X transposon system 25 and demonstrated that the subretinal transplantation of PEDF-transfected cells in a rat model of CNV reduces neovascularization and inhibits new vessel formation 46 …”

A Critical Analysis of the AvailableIn VitroandEx VivoMethods to Study Retinal Angiogenesis

“…However, we also found several studies using nonretinal specific ECs with the aim of studying retinal angiogenesis. In this group, the most used cells are human umbilical vein endothelial cells (HUVECs) [ 26 , 53 , 70 , 76 , 78 , 96 – 130 ]. Human microvascular endothelial cells (HMVECs) [ 131 , 132 ] and human dermal microvascular endothelial cells (HMVEC-D) [ 133 ] constitute other examples.…”

“…Only 9 [ 31 , 50 , 55 , 56 , 62 , 97 , 127 , 130 , 133 ] apply a 3D model to study tubulogenesis. In these assays, two different methods are reported.…”

PEDF Reduces the Severity of Herpetic Simplex Keratitis in Mice

“…[10][11][12] Previous reports have confirmed the effect of PEDF on the suppression of neovascularization via inhibiting the proliferation and migration of vascular endothelial cells in choroid, retina, and cornea. [13][14][15] In capsaicin-induced neurotrophic keratouveitis, PEDF can promote the recovery of tear secretion and attenuate the corneal ulceration, neovascularization and leukocyte infiltration. 16 Recently, PEDF has been reported to accelerate corneal epithelial wound healing, stimulate nerve regeneration, and inhibit pathologic neovascularization.…”