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Purpose: It has been reported that the platelet-derived growth factor (PDGF)-alpha receptor (alphaPDGFR) is required for experimental proliferative vitreoretinopathy (PVR) in rabbits. This study investigated which of the signaling enzymes downstream of the alphaPDGFR participate in PVR., Methods: A panel of cell lines that expressed alphaPDGFR signaling mutants were made and characterized. These cell lines were used in a rabbit model of PVR and in an in vitro collagen type I contraction assay., Results: Phosphoinositide 3-kinase (PI3K) and, to a lesser extent, phospholipase C (PLC)-gamma were the signaling enzymes required for the alphaPDGFR to mediate PVR. Furthermore, the cells lines that were the most effective at inducing PVR displayed the most potent activity in the in vitro contraction assay., Conclusions: PI3K and PLCgamma are necessary downstream effectors of the alphaPDGFR in experimental PVR. Consequently, these two signaling enzymes are required for one or more of the cellular responses (chemotaxis, proliferation, extracellular matrix production, contraction) that contribute to PVR.

Purpose: The work from numerous laboratories has led to the idea that the growth factors such as platelet-derived growth factor (PDGF) contribute to proliferative vitreoretinopathy (PVR) in experimental models of the disease, as well as in humans. In support of this idea, the authors have previously reported that cells unable to respond to PDGF had a greatly reduced PVR potential, compared with PDGF-responsive versions of the same cells. The goal of this study was to test the effect of blocking the output of the PDGF receptor in an experimental model of PVR., Methods: Polymerase chain reaction-based site-directed mutagenesis was used to generate point mutations in the human PDGF alpha receptor (alphaPDGFR) cDNA, which resulted in single amino acid substitutions. These changes were based on naturally occurring point mutations in the c-kit receptor tyrosine kinase, which suppresses the function of wild-type c-kit. A truncated alphaPDGFR was also made, in which the receptor ended just after the juxtamembrane domain. As with the point mutants, truncated receptors have been shown to block the action of wild-type receptors. All the alphaPDGFR mutants were introduced into cells that naturally express the wild-type receptor, and the PDGF-dependent output of the resultant cell lines was determined. In addition, the PVR potential of cell lines expressing the mutant receptors was tested in a PVR rabbit model., Results: Although the mutants differed in their ability to suppress PDGF-dependent signaling of the wild-type receptor, each mutant effectively blocked cell cycle progression. When expressed in rabbit conjunctival fibroblasts, a cell line that effectively induces PVR, the mutant receptors blocked PVR to various degrees. The most effective receptor was the truncated mutant., Conclusions: These data suggest that the alphaPDGFR plays an important role in PVR. In addition, these mutant receptors appear to have therapeutic potential for prevention of this blinding disease.

We tested the hypothesis that Src family kinases (SFK) contribute to c-Cbl-mediated degradation of the platelet-derived growth factor (PDGF) alpha receptor (alphaPDGFR). Using either a receptor mutant that does not engage SFKs (F72/74), or cells that that lack SFKs, we found that SFKs contributed to degradation of the alphaPDGFR. Overexpression of c-Cbl also reduced the receptor half-life, but only if the receptor was able to engage SFKs. In cultured cells, prolonging the half-life of the receptor correlated with enhanced signaling and more efficient S phase entry, whereas accelerating receptor degradation had the opposite effect. Consistent with these tissue culture findings, there was a statistically significant increase in the onset of a proliferative retinal disease when animals were injected with cells expressing the F72/74 receptor, as compared with cells expressing the WT receptor. Our findings suggest that SFKs cooperate with c-Cbl to negatively regulate the alphaPDGFR, and that the SFK/c-Cbl suppression of alphaPDGFR output is relevant to the onset and progression of a proliferative disease.

Purpose: The action of growth factors is thought to make a substantial contribution to the events leading to proliferative vitreoretinopathy (PVR). In this study, the importance of platelet-derived growth factor (PDGF) was tested in a rabbit model of PVR., Methods: The approach was to compare the extent of PVR induced by cells that do or do not express the receptors for PDGF and therefore differ in their ability to respond to PDGF., Results: Mouse embryo fibroblasts derived from PDGF receptor knock-out embryos that do not express either of the two PDGF receptors induced PVR poorly when injected into the eyes of rabbits that had previously undergone gas vitrectomy. Re-expression of the PDGF beta receptor in these cells did not improve the ability of the cells to cause PVR. In contrast, injection of cells expressing the PDGF alpha receptor resulted in stage 3 or higher PVR in 8 of 10 animals., Conclusions: These findings show that PDGF makes an important contribution to the development of PVR in this animal model. Furthermore, there is a marked difference between the two receptors for PDGF, and it is the PDGF alpha receptor that is capable of driving events that lead to PVR.

Purpose: To evaluate the antiproliferative properties of alpha-tocopherol and alpha-tocopheryl-acid-succinate in a rabbit model of proliferative vitreoretinopathy., Methods: Fifty-seven rabbits underwent gas-compression vitrectomy and gas/fluid exchange. Group 1 (n = 8): 50 micrograms alpha-tocopherol in 1 ml of 0.5% ethanol in balanced salt solution; Group 2 (n = 8): 50 micrograms alpha-tocopheryl-acid-succinate in 1 ml of 0.5% ethanol in balanced salt solution; Groups 3 (n = 4) and 4 (n = 2): 1 ml 0.5% ethanol in balanced salt solution; Groups 5 (n = 12) and 8 (n = 9): alpha-tocopherol in 1 ml silicone oil (12 mg/ml); Group 6 (n = 9): 1 ml silicone oil; Group 7 (n = 5): 1 ml balanced salt solution. Groups 1-3, and 5-7 also received fibroblasts and platelet-rich plasma injection. Fundus evaluation was performed during a four-week period. The eyes were enucleated for gross examination on day 28. Histopathology was performed on Group 4., Results: Alpha-tocopherol and alpha-tocopheryl-acid-succinate in saline solution delayed development of proliferative vitreoretinopathy, compared to the control group (statistically significant during the first week, Mann Whitney, p < 0.05). The alpha-tocopherol in the silicone-oil group delayed development of proliferative vitreoretinopathy, compared to the silicone-oil group during the second to fourth weeks (no statistically significant difference, p > 0.05)., Conclusions: Alpha-tocopherol and alpha-tocopheryl-acid-succinate in saline solution showed retardation of proliferative vitreoretinopathy traction retinal detachments. In silicone oil, alpha-tocopherol is slowly released and decreases the severity of proliferative vitreoretinopathy, especially during the second week of follow-up.

Purpose: To determine the antiproliferative activity in intravitreous retinoic acid (RA) dispersed in 1% sodium hyaluronate (HA)., Methods: Six groups of pigmented rabbits underwent gas-compression vitrectomy. Four days later, gas/HA or gas/balanced salt solution (BSS) exchange (1.0 m1) was performed in all rabbits. Groups A (n = 10) and B (n = 5) received intravitreous RA dissolved in 0.01 m1 of ethanol and dispersed in 1% HA (10 and 15 micrograms RA/m1, respectively). Group C (n = 10) received intravitreous RA dissolved in ethanol and dispersed in BSS (10 micrograms RA/m1). Groups D (n = 5) and F (n = 4) received 1 m1 of HA with ethanol; group E (n = 5) received 1 m1 of HA without ethanol. All groups except group F also received homologous fibroblasts and autologous, platelet-rich plasma intravitreously. The eyes were examined ophthalmoscopically for 1 month. Proliferative vitreoretinopathy (PVR) findings were graded according to the classification of Fastenberg et al. all group F eyes also were examined by light and electron microscopy., Results: RA in HA lessened PVR progression within 1 month when compared with HA injection controls and within 2 weeks when compared with the RA in BSS treatment group (both, p < 0.05). NO specific change attributable to ethanol was observed histopathologically., Conclusion: RA dissolved in ethanol and dispersed in HA could be useful to treat PVR when silicone oil is unnecessary.