RETINAL FEATURES PREDICTIVE OF PROGRESSIVE STAGE 4 RETINOPATHY OF PREMATURITY

To determine the retinal features predictive of progressive stage 4 retinopathy of prematurity (ROP) after laser treatment for threshold ROP

FIBROVASCULAR ORGANIZATION IN THE VITREOUS FOLLOWING LASER FOR ROP: Implications for Prognosis

To study associations between surgical outcome and mean postmenstrual age (PMA) when fibrovascular organization is detected between vascular and avascular retina following laser for acute retinopathy of prematurity (ROP)

VEGF isoforms and their expression after a single episode of hypoxia or repeated fluctuations between hyperoxia and hypoxia: Relevance to clinical ROP

Purpose—Fluctuations in oxygen are associated with the development of severe retinopathy of prematurity (ROP) in humans. However, the causal relationships between oxygen variability and severe ROP remain unknown. We investigated whether isoforms of vascular endothelial growth factor (VEGF) were differentially stimulated by hypoxia and by repeated fluctuations between hypoxia and hyperoxia, and whether isoforms were differentially expressed in association with intravitreous neovascularization. We also determined whether pigment epithelium-derived factor (PEDF) was dysregulated by oxygen fluctuations perhaps contributing to a delay in normal retinal vascular development. Methods—We used the 50/10 oxygen-induced retinopathy (50/10 OIR) model that exposes newborn rat pups to repeated cycles of 24 h of 50% oxygen alternating with 24 h of 10% oxygen to cause a condition similar to human ROP. Animals were euthanized at postnatal day 2 (P2; after one cycle of 50/10% oxygen), P7 (after 3.5 cycles of 50/10% oxygen), and P14 (after seven cycles of 50/10% oxygen). Room air raised control rat pups were also exposed to a single episode of 24 h of hypoxia at P7 and P14 and assayed immediately afterwards. Retinal VEGF isoforms and PEDF were measured by RT-PCR. Total VEGF protein was measured by ELISA. Results—We found that repeated cycles of hyperoxia and hypoxia caused greater expression of VEGF protein compared to control than did a single cycle of hyperoxia and hypoxia. VEGF164 mRNA had a greater fold change over control after repeated oxygen fluctuations than after a single episode of hypoxia. However, the other isoforms, VEGF188 and VEGF120, were expressed to a similar degree regardless of whether the stimulus was a single episode of hypoxia or repeated fluctuations in oxygen. VEGF164 was the predominant isoform expressed at the time of maximal intravitreous neovascularization. Retinal PEDF expression was elevated in pups in the 50/10 OIR model compared to control at P7, immediately after 50% oxygen. PEDF expression in the experimental group was similar to control at P18, when intravitreous neovascularization occurred. Conclusions—Repeated fluctuations in oxygen results in a greater expression of the pathologic isoform, VEGF164, than does hypoxia alone. However, the other isoforms were upregulated to an equivalent degree over control by repeated fluctuations in oxygen or a single episode of hypoxia. Total VEGF protein was increased to a greater degree by repeated fluctuations in oxygen compared to a single cycle of oxygen. PEDF was increased over control early in the 50/10 OIR model and may play a role in the observed delay in retinal vascularization. These findings provide insight into the effect of repeated oxygen fluctuations on the development of severe ROP in preterm infant

Exogenous leukemia inhibitory factor (LIF) attenuates retinal vascularization reducing cell proliferation not apoptosis

To study the effect of leukemia inhibitory factor (LIF) on rat retinal vascular development, Sprague–Dawley rats at postnatal age 3 days (p3) were given intraperitoneal (IP) LIF and analysis performed at p6 (p3/6). p7 rats were given intravitreous (IV) LIF and analysis performed at p9 (p7/9). Control animals were PBS injected. At the time of analysis retinal flatmounts were prepared and stained with Griffonia lectin and activated caspase-3. The retinal peripheral avascular area was measured and number of apoptotic cells counted. In vitro, human retinal microvascular endothelial cells (RMVECs) were cultured in media containing LIF, with and without neutralizing antibody to LIF. Cells were stained with activated caspase-3 and apoptotic cells counted. Proliferation was measured by counting cell numbers, and cell cycle stage was determined using propidium iodide staining and FACS analysis. LIF injected either IP or IV had no effect on body weight or total retina area, but significantly increased the peripheral retinal avascular area. In both IP and IV injected groups there was no difference in the number of apoptotic cells between PBS-or LIF-injected groups; although in the p7/9 retinas, both injected groups had significantly more apoptotic cells than the non-injected group. In vitro, there was no effect of LIF on RMVEC apoptosis; however, cell counts were significantly lower in the LIF-treated group. Antibody to LIF restored the cell counts to untreated levels. LIF reduced the number of cells in S phase. LIF attenuates retinal vascular development in vivo through growth arrest, and not apoptosis, of endothelial cells

Triamcinolone Reduces Neovascularization, Capillary Density and IGF-1 Receptor Phosphorylation in a Model of Oxygen-Induced Retinopathy

To study the effects of intravitreous triamcinolone acetonide (TA) on neovascularization (NV), capillary density, and retinal endothelial cell (REC) viability in a model of oxygen-induced retinopathy (OIR)

Connexin 30 expression and frequency of connexin heterogeneity in astrocyte gap junction plaques increase with age in the rat retina.

We investigated age-associated changes in retinal astrocyte connexins (Cx) by assaying Cx numbers, plaque sizes, protein expression levels and heterogeneity of gap junctions utilizing six-marker immunohistochemistry (IHC). We compared Wistar rat retinal wholemounts in animals aged 3 (young adult), 9 (middle-aged) and 22 months (aged). We determined that retinal astrocytes have gap junctions composed of Cx26, -30, -43 and -45. Cx30 was consistently elevated at 22 months compared to younger ages both when associated with parenchymal astrocytes and vascular-associated astrocytes. Not only was the absolute number of Cx30 plaques significantly higher (P<0.05) but the size of the plaques was significantly larger at 22 months compared to younger ages (p<0.05). With age, Cx26 increased significantly initially, but returned to basal levels; whereas Cx43 expression remained low and stable with age. Evidence that astrocytes alter connexin compositions of gap junctions was demonstrated by the significant increase in the number of Cx26/Cx45 gap junctions with age. We also found gap junctions comprised of 1, 2, 3 or 4 Cx proteins suggesting that retinal astrocytes use various connexin protein combinations in their gap junctions during development and aging. These data provides new insight into the dynamic and extensive Cx network utilized by retinal astrocytes for communication within both the parenchyma and vasculature for the maintenance of normal retinal physiology with age. This characterisation of the changes in astrocytic gap junctional communication with age in the CNS is crucial to the understanding of physiological aging and age-related neurodegenerative diseases