The effect of intense visible light (light damage) on the expression of heme oxygenase 1 (HO-1), a protein induced by oxidative stress, was investigated in the rat retina. A sensitive reverse transcription-PCR assay demonstrated the expression of mRNA for HO-1 as well as HO-2, the noninducible HO form, in the normal retina. As analyzed by Northern blotting, however, HO-1 mRNA was barely detectable under normal circumstances. After exposure to intense visible light, retinas had markedly higher HO-1 mRNA levels than unexposed controls, with increases up to 52- and 98-fold at 12 and 24 hr of exposure, respectively. Intense light exposure also resulted in an increase in HO-1 protein. In contrast, no appreciable change in HO-2 mRNA or protein was observed. The increase in HO-1 message was more pronounced in rats previously reared in the dark than in those reared in a weak cyclic-light environment. A marked decrease from the high level of HO-1 mRNA induced by light insult was observed when the animals were allowed to recover in the dark for 24 hr after light exposure. Most important, treatment of animals with 1,3-dimethylthiourea, a synthetic antioxidant, prior to light exposure effectively blocked the increase in HO-1 mRNA. Thus, HO-1 is a sensitive marker for assessing light-induced insult in the retina. Since increased expression of HO-1 is thought to be a cellular defense against oxidative damage, its expression may play an important role in protecting the retina against light damage.

Surgically excised retinoblastomas from 14 patients (age range nine months to two years) were assessed by immunocytochemistry for the expression of photoreceptor-specific proteins and neuronal and glial cell markers. Adjacent tissues were examined for messenger RNA expression of interphotoreceptor retinoid-binding protein (IRBP) using Northern blots. For immunocytochemical stains (ABC method), monoclonal and polyclonal antibodies included S-Ag, rhodopsin, neuron specific enolase (NSE), glial fibrillary acidic protein (GFAP), IRBP, neural adhesion molecule (N-CAM), and rod and cone specific transducin (TR alpha and TC alpha). Histopathology revealed mostly poorly differentiated tumors with necrosis and lack of Flexner-Wintersteiner rosettes. Immunocytochemical staining showed focal IRBP expression in one of the tumors and S-antigen in two cases. Immunoreactivity with rhodopsin was negative. N-CAM, a neural adhesive protein which appears to be involved in the regulation of adhesive interaction during neuronal differentiation, was positive except in two cases. All tumors showed immunoreactivity with NSE, whereas GFAP staining was limited to the perivascular glial tissue confirming the essential neuronal nature of retinoblastoma cells. TC alpha was detected in all tumors and TR alpha in one case. Messenger RNA for IRBP was detected in tumors in which IRBP immunoreactivity could not be detected.

Interphotoreceptor retinoid-binding protein (IRBP) is synthesized and secreted by photoreceptor cells and is thought to facilitate the transport of retinoids during the visual cycle as well as fatty acids essential to the maintenance of normal outer segment membranes. Proteins such as IRBP, which are unique to the photoreceptor cells in the retina, are prime candidates in the consideration of biochemical defects which could contribute to photoreceptor cell degeneration in man and animals. In this study, the association between IRBP and retinal degeneration was examined using the progressive rod-cone degeneration (prcd) mutant retina in dogs as an animal model. This study shows that loss of IRBP is not an early occurrence in prcd. IRBP is present in relatively normal amounts and distribution even at 1.7 years of age, a time when there is extensive visual cell disease and degeneration. By 2.7-3.0 years of age, IRBP loss correlates with the severity of the disease and concomitant loss of photoreceptor cells. IRBP immunoreactivity was present in the interphotoreceptor matrix (IPM) as long as inner segments were present to a significant degree. The late loss of IRBP immunoreactivity seems to be, therefore, the result of advanced degeneration and end-stage atrophy of the retina. In addition, immunological studies were carried out in order to examine the possible role of an autoimmune response against IRBP in the disease cascade. Normal, heterozygote and prcd-affected dogs had measurable antibody titers to IRBP, but there was no correlation between disease state and antibody levels.

The postnatal development of immunoreactivity for photoreceptor-specific markers was studied in mice carrying the genes rd (retinal degeneration) and rds (retinal degeneration slow) in different combinations. Antibodies raised against three specific photoreceptor proteins (opsin, alpha-transducin and S-antigen) were applied on retinae from mice with the following allelic combinations at the rd and rds loci: +/+, +/+ (control); rd/rd, +/+; rds/rds, +/+; rds/+, +/+; and rd/rd, rds/rds. Immunoreactivity for each antibody appeared simultaneously in normal and mutants. Thereafter, the immunoreactivity patterns in the mutants diverged from the normal phenotype. Except for a dramatic loss of photoreceptor cells in the mutants, the main divergence from the normal development consisted of a progressive loss of the intracellular immunoreactivity compartmentalization for each protein. As degeneration progressed, the remaining photoreceptors became homogeneously labelled; one this labelling pattern was acquired, it was maintained during subsequent stages of development. It is proposed that this pattern, common for all phenotypes studied, may be due to the loss of structural and biochemical polarity of the photoreceptor cells undergoing degeneration, and that this may be an important primary or secondary aspect of the disease process.

Using immunocytochemical techniques, development of opsin, transducin alpha and S-antigen in photoreceptor cells of the mice homozygous or heterozygous for the rd or rds genes has been found to be similar to that of control animals during the first postnatal week. Even though the absolute amounts of these proteins are low (eg. opsin in the rds retina) or decrease in the postnatal period (as in the rd retina), we can demonstrate their persistence during the entire degeneration process. In fact, the content of the proteins in the photoreceptor perikarya actually appear to be higher after postnatal day 11 in all mutants studied. Thus, one of the major manifestations of the mutant retinae is a loss of polarity of the photoreceptor cells at the time of ROS degeneration without a loss in the ability to synthesize these important proteins of the visual cycle. This correlates well with the apparent defect in IRBP secretion and its intracellular accumulation in mutant photoreceptor cells as previously observed (van Veen et al. 1986). In the Abyssinian cat model for progressive retinal atrophy, the development an cellular distribution of all the proteins studied are similar in affected and control retinae until the beginning of stage 2 of the disease. At this time, outer segments begin to degenerate and immunoreactivity increases in the photoreceptor perikarya. The IRBP content of the retina declines markedly at stage 2, preceding extensive loss of photoreceptors.

After visual-pigment bleaching, single isolated rod photoreceptors of Ambystoma tigrinum recover their sensitivity to light when supplied with 11-cis-retinal from liposomes or with 11-cis-retinal bound to interphotoreceptor retinoid-binding protein. Bleached rods do not recover sensitivity, or do so only very slowly, after exposure to 11-cis-retinol. The latter retinoid is "toxic" in that rods actually lose sensitivity in its presence. In contrast, bleached isolated cone cells recover sensitivity when either retinoid is supplied. It is suggested that the major pathway for rhodopsin regeneration during dark adaptation in the intact eye is transport of 11-cis-retinal from the pigment epithelium to the retina. The results also suggest that there may be separate pathways for visual-pigment regeneration in rods and cones during dark adaptation.

An asymptomatic 19-year-old male with choroideremia had diffuse loss of retinal pigment epithelium (RPE) and choroid except for the periphery and macula. Fluorescein angiography of the arteriovenous phase showed absence of retinal pigment epithelium and exaggerated visualization of choroidal vessels in involved areas. The mother was a typical carrier with pigment stippling of the midperipheral retina. Histopathologic examination of affected areas of one eye showed marked degeneration of the outer and midretina with loss of retinal pigment epithelium and Bruch's membrane, absence of choriocapillaris, chorioretinal adhesions and gliosis. Atrophy of inner and mid-choroid was also observed. Pigmented macrophage-like cells had migrated into the outer and midretinal layers. Electron microscopy disclosed macrophage-like cells with trilaminar structures and photoreceptor phagosomes in the RPE and outer retina. Remnants of photoreceptor outer segments were adherent to the plasma membranes of the macrophage-like cells. Biochemical analysis of retinal tissue samples for interphotoreceptor retinoid-binding protein (IRBP) showed marked reduction in the 146K bands in the equator and posterior pole in the patient compared to controls. Cyclic nucleotide content was altered in the retinal equator. Cyclic AMP was several-fold higher in the RPE-choroid complex of the affected eye than in the control.

Abstract— As assessed by sucrose density gradient ultracentrifugation, bovine retinal cytosol exhibits 2S and 7S vitamin A binding species (‘receptors’). Upon fractionation of the retina, outer segment photoreceptor units are enriched in 7S receptor whereas the outer segment poor layers of the retina have a decreased amount of 7S receptor. The 2S vitamin A receptor is found both in the photoreceptor fraction and in the rod‐poor layers of the retina. The supernatant fraction of fetal retina demonstrates 2S binding but no 7S binding; a small 7S peak observed in adult pigment epithelial supernatant preparations is also not seen in the supernatant fraction of fetal pigment epithelial cells. The binding pattern in the newborn retina is similar to that in adult retina, i.e. extensive 7S as well as 2S binding. Adult bovine brain exhibits a large 7S receptor peak which is missing in fetal brain supernatant and virtually absent in newborn brain. The 7S receptor may thus be compartmentalized in retinal photoreceptors but is not unique to the retina since it is also observed in brain. The ontogenic patterns in the two tissues are different however.