Photoreceptor rescue by an abbreviated human RPGR gene in a murine model of X-linked retinitis pigmentosa.

The X-linked RP3 gene codes for the ciliary protein RPGR and accounts for over 10% of inherited retinal degenerations. The critical RPGR-ORF15 splice variant contains a highly repetitive purine-rich linker region that renders it unstable and difficult to adapt for gene therapy. To test the hypothesis that the precise length of the linker region is not critical for function, we evaluated whether adeno-associated virus-mediated replacement gene therapy with a human ORF15 variant containing in-frame shortening of the linker region could reconstitute RPGR function in vivo. We delivered human RPGR-ORF15 replacement genes with deletion of most (314 codons, 'short form') or 1/3 (126 codons, 'long form') of the linker region to Rpgr null mice. Human RPGR-ORF15 expression was detected post treatment with both forms of ORF15 transgenes. However, only the long form correctly localized to the connecting cilia and led to significant functional and morphological rescue of rods and cones. Thus the highly repetitive region of RPGR is functionally important but that moderate shortening of its length, which confers the advantage of added stability, preserves its function. These findings provide a theoretical basis for optimizing replacement gene design in clinical trials for X-linked RP3.

Gene therapy with a promoter targeting both rods and cones rescues retinal degeneration caused by AIPL1 mutations.

Aryl hydrocarbon receptor-interacting protein-like 1 (AIPL1) is required for the biosynthesis of photoreceptor phosphodiesterase (PDE). Gene defects in AIPL1 cause a heterogeneous set of conditions ranging from Leber's congenital amaurosis (LCA), the severest form of early-onset retinal degeneration, to milder forms such as retinitis pigmentosa (RP) and cone-rod dystrophy. In mice, null and hypomorphic alleles cause retinal degeneration similar to human LCA and RP, respectively. Thus these mouse models represent two ends of the disease spectrum associated with AIPL1 gene defects in humans. We evaluated whether adeno-associated virus (AAV)-mediated gene replacement therapy in these models could restore PDE biosynthesis in rods and cones and thereby improve photoreceptor survival. We validated the efficacy of human AIPL1 (isoform 1) replacement gene controlled by a promoter derived from the human rhodopsin kinase (RK) gene, which is active in both rods and cones. We found substantial and long-term rescue of the disease phenotype as a result of transgene expression. This is the first gene therapy study in which both rods and cones were targeted successfully with a single photoreceptor-specific promoter. We propose that the vector and construct design used in this study could serve as a prototype for a human clinical trial.

Clinical features and mutations in patients with dominant retinitis pigmentosa-1 (RP1).

PURPOSE: To survey patients with dominant retinitis pigmentosa (RP) for mutations in the RP1 gene to determine the spectrum of dominant mutations in this gene, to estimate the proportion of dominant RP caused by this gene, and to determine whether the clinical features of patients with RP1 mutations differ from features of those with rhodopsin mutations. METHODS: A set of 241 patients who did not have mutations in the rhodopsin gene (based on previous work) formed the basis for the study. Of these patients, 117 had also been previously evaluated and were found not to carry mutations in the RDS gene. The single-strand conformation polymorphism (SSCP) method was used to search for sequence variants, which were then directly sequenced. The relatives of selected patients were recruited for segregation analyses. Clinical evaluations of patients included a measurement of Snellen visual acuity, final dark adaptation thresholds, visual fields, and ERGs. Clinical data were compared with those obtained earlier from a study of 128 patients with dominant rhodopsin mutations. RESULTS: Of the 241 patients, all were screened for the most common RP1 mutation (Arg677Ter), and 10 patients were found to have this mutation. In addition, an evaluation of a subset of 189 patients in whom the entire coding sequence was evaluated revealed the following mutations: Gln679Ter (1 case), Gly723Ter (2 cases), Glu729(1-bp del) (1 case), Leu762(5-bp del) (2 cases), and Asn763(4-bp del) (1 case). All of these mutations cosegregated with RP in the families of the index patients. Nine missense mutations that were each found in six or fewer patients were encountered. The segregation of eight of these was evaluated in the respective patients' families, and only one segregated with dominant RP. This cosegregating missense change was in cis with the nonsense mutation Gln679Ter. Although patients with RP1 mutations had, on average, slightly better visual acuity than patients with rhodopsin mutations, there was no statistically significant difference in final dark-adaptation thresholds, visual field diameters, or cone electroretinogram (ERG) amplitudes. Comparably aged patients with RP1 mutations had visual function that varied by approximately two orders of magnitude, based on visual fields and ERG amplitudes. CONCLUSIONS: Dominant RP1 alleles typically have premature nonsense codons occurring in the last exon of the gene and would be expected to encode mutant proteins that are only approximately one third the size of the wild-type protein, suggesting that a dominant negative effect rather than haploinsufficiency is the mechanism leading to RP caused by RP1 mutations. On average, patients with RP1 mutations have slightly better visual acuity than patients with dominant rhodopsin mutations; otherwise, they have similarly severe disease. The wide range in severity among patients with RP1 mutations indicates that other genetic or environmental factors modulate the effect of the primary mutation.

Endothelin-1-mediated retinal artery vasospasm and the rabbit electroretinogram.

Electroretinogram (ERG) changes invariably accompany the selective interruption of the retinal circulation that occurs in human central retinal artery occlusion. Since arteriolar ligation or ocular hypertension in the rabbit eye is occasionally used to model human central retinal artery occlusion, we conducted the present study to determine whether selective interruption of the retinal circulation of the rabbit eye alters retinal function as measured by the ERG. The vasoconstrictor, endothelin-1, was injected into the vitreous of rabbits' eyes to induce complete vasospasm and selective interruption of the retinal circulation. This procedure was compared to vascular ligation of the ophthalmic and ciliary arteries in which both the retinal and choroidal circulations were interrupted. A total of 8 rabbits was studied. Circulation was monitored angiographically in half of the eyes, and retinal function was monitored by the ERG in the remaining eyes. Endothelin-1 obliterated retinal arteriolar blood flow without affecting choroidal blood flow for at least 1 hr. Although ERG a-wave amplitude showed a small decline over 2 hr, b-wave and oscillatory potential amplitudes (measures of inner retinal function) showed no loss over this period. In contrast, ligation of the ophthalmic and ciliary arteries produced complete obliteration of both retinal arteriolar and choroidal blood flow and complete loss of the ERG after 2 min. Endothelin-1 induces acute, selective interruption of retinal arteriolar blood flow which has no significant physiologic effect on inner retinal function of the rabbit as monitored by the ERG. The avascular rabbit retina appears to be a poor choice for modeling human retinal artery occlusion.

X-linked retinitis pigmentosa: mutation spectrum of the RPGR and RP2 genes and correlation with visual function.

PURPOSE: To assess the frequency of RPGR and RP2 mutations in a set of 85 patients with X-linked retinitis pigmentosa (XLRP) and to compare the visual function of patients with mutations in RPGR versus RP2. METHODS: Eighty-five unrelated patients with XLRP were ascertained, mainly from North America. The single-strand conformation polymorphism (SSCP) and a direct sequencing technique were used to screen their DNA for mutations in the coding region and splice sites of RPGR and RP2. The Snellen visual acuities, visual field areas, and 0.5-Hz and 30-Hz electroretinograms (ERGs) were measured in male patients. The visual function parameters were compared using multiple regression analysis. RESULTS: A wide spectrum of mutations was found in both genes, including missense, nonsense, splice-site, and frameshift mutations. Twenty putative pathogenic mutations in RPGR, 15 of which were novel, were found in 22 patients (26%), whereas 6 mutations in RP2, 4 of which were novel, were found in 6 patients (7%). A high fraction of the mutations in both genes affected amino acid residues within or adjacent to presumed functional domains. Comparison of visual function between comparably aged patients with mutations in RPGR versus RP2 showed that, on average, patients with RPGR mutations have lower ERG amplitudes and smaller visual field areas. CONCLUSIONS: Mutations in RPGR and RP2 genes together account for approximately 33% of cases of XLRP in North America. Patients with RPGR mutations have less overall retinal function on average than those with RP2 mutations, on the basis of measurements of visual field areas and full-field ERG amplitudes.

A retinitis pigmentosa GTPase regulator (RPGR)-deficient mouse model for X-linked retinitis pigmentosa (RP3).

The X-linked RP3 locus codes for retinitis pigmentosa GTPase regulator (RPGR), a protein of unknown function with sequence homology to the guanine nucleotide exchange factor for Ran GTPase. We created an RPGR-deficient murine model by gene knockout. In the mutant mice, cone photoreceptors exhibit ectopic localization of cone opsins in the cell body and synapses and rod photoreceptors have a reduced level of rhodopsin. Subsequently, both cone and rod photoreceptors degenerate. RPGR was found normally localized to the connecting cilia of rod and cone photoreceptors. These data point to a role for RPGR in maintaining the polarized protein distribution across the connecting cilium by facilitating directional transport or restricting redistribution. The function of RPGR is essential for the long-term maintenance of photoreceptor viability.

Acuity recovery and cone pigment regeneration after a bleach in patients with retinitis pigmentosa and rhodopsin mutations.

PURPOSE: To assess visual acuity recovery times and cone photopigment regeneration kinetics after a bleach in the fovea of patients with dominant retinitis pigmentosa due to rhodopsin mutations. METHODS: The authors measured acuity recovery times by computerized photostress testing in 13 patients with dominant retinitis pigmentosa and one of eight rhodopsin mutations. The authors also measured their time constants of cone photopigment regeneration with a video imaging fundus reflectometer to determine whether acuity recovery time depended on pigment regeneration kinetics. These values were compared with those of normal subjects, by the Mann-Whitney U test. The relationship between acuity recovery time and the time constant of cone photopigment regeneration among the patients was quantified by the Spearman rank correlation. RESULTS: The visual acuity recovery times, which averaged 22.0 seconds for the patients with retinitis pigmentosa and 11.2 seconds for the normal subjects, were significantly slower for the patient group (P < 0.001). The time constants of cone pigment regeneration, which averaged 172 seconds for the patients with retinitis pigmentosa and 118 seconds for the normal subjects, also were significantly slower for the patient group (P = 0.043). The authors also found a significant, positive correlation between the visual acuity recovery time and the time constant of pigment regeneration for the patients with retinitis pigmentosa (r = 0.65, P = 0.017). CONCLUSIONS: A slowing of foveal visual acuity recovery and cone pigment regeneration, which are related to each other, can occur in patients with retinitis pigmentosa, due to a rod-specific gene defect.

The effect of a lower blood pressure on choroidal filling in age-related macular degeneration.

PURPOSE: To determine whether the occurrence of delayed choroidal filling or a slower foveal electroretinogram (ERG) implicit time is related to systemic blood pressure in patients with age-related macular degeneration. METHODS: We assessed delayed choroidal filling by fluorescein angiography and foveal ERG implicit time with a hand-held stimulator-ophthalmoscope for the fellow eye of 67 patients with unilateral neovascular age-related macular degeneration. Blood pressure, measured by ascultation, was converted to mean arterial pressure. The relations of delayed choroidal filling and foveal ERG implicit time to the mean arterial pressure were assessed by logistic and linear regression, respectively, controlling for age, gender, and intake of hypertensive medication. RESULTS: The probability of having delayed choroidal filling decreased with increasing mean arterial pressure (P = 0.01). The foveal ERG implicit time was also inversely related to the mean arterial pressure (P = 0.009). CONCLUSION: Delayed choroidal filling and foveal ERG implicit time are related to blood pressure in patients with age-related macular degeneration.

Effect of vitamin A supplementation on rhodopsin mutants threonine-17 --> methionine and proline-347 --> serine in transgenic mice and in cell cultures.

A therapeutic effect of vitamin A supplementation on the course of photoreceptor degeneration, previously reported for patients with retinitis pigmentosa, was tested in two transgenic mouse models of this disease, each carrying a dominant rhodopsin mutation. The threonine-17 --> methionine (T17M) mutation is a class II rhodopsin mutation, characterized by a thermal instability/folding defect and minimal regeneration with the chromophore. The proline-347 --> serine (P347S) mutation belongs to class I, comprised of a smaller number of mutations that exhibit no recognized biochemical abnormality in vitro. In the present study, each of the two mouse models was fed a diet containing 2.5 mg of vitamin A palmitate (control) or 102.5 mg of vitamin A palmitate (high vitamin A) per kilogram of diet. Dark-adapted, full-field electroretinograms showed that the high vitamin A diet significantly reduced the rate of decline of a-wave and b-wave amplitudes in the T17M mice but had no significant effect on the decline of electroretinogram amplitude in the P347S mice. Correspondingly, histologic evaluation revealed that the treatment was associated with significantly longer photoreceptor inner and outer segments and a thicker outer nuclear layer in the T17M mice but had no effect on photoreceptor morphology in the P347S mice. In a separate series of experiments, the instability defect of the T17M mutant opsin expressed in vitro was partially alleviated by inclusion of 11-cis-retinal in the culture media. These results show that vitamin A supplementation slows the rate of photoreceptor degeneration caused by a class II rhodopsin mutation. Vitamin A supplementation may confer therapeutic benefit by stabilizing mutant opsins through increased availability of the chromophore.

Rod and cone function in the Nougaret form of stationary night blindness.

BACKGROUND: Recently, a mutation (Gly38Asp) was identified in the alpha subunit of rod transducin in members of the Nougaret pedigree affected with dominantly inherited stationary night blindness. OBJECTIVE: To evaluate retinal function in patients with the Gly38Asp gene defect. DESIGN: Ocular examinations, including specialized measures of rod and cone function. SETTING: A clinical research facility in Boston, Mass. PATIENTS: A father (aged 48 years) and son (aged 25 years) with the Gly38Asp mutation. MAIN OUTCOME MEASURES: Psychophysical thresholds to white and narrowband lights and full-field electroretinographic (ERG) responses. RESULTS: Both patients showed dark-adapted thresholds to white light that were elevated approximately 2 log-units across the retina. Spectral sensitivity testing revealed thresholds that seemed to be governed mostly by rods. Although both patients' dark-adapted ERG responses to a dim blue flash were nondetectable, their dark-adapted ERGs to a white flash showed an a-wave with cone and rod components and a b-wave amplitude larger than what could have been generated by cone function alone. Rod ERGs to bright blue flashes had subnormal, but detectable, amplitudes that seemed to result from a profound reduction in sensitivity. The patients also showed loss of a cone subcomponent in the dark-adapted response to a red flash. The abnormal dark-adapted ERG responses of the patients could be simulated in the ERG responses of normal subjects tested with blue, white, and red flashes presented in the presence of a mesopic background. CONCLUSIONS: Although the Nougaret form of stationary night blindness has been cited as a prototype of absent rod function with normal cone function, our findings, based on the genealogically and genotypically documented descendants of Jean Nougaret, show that rod function is present, although subnormal, and that there is slight impairment of cone function. The data also suggest that these abnormalities can be simulated by light-adapting the normal retina, compatible with the proposal that the rod transducin encoded by the mutant gene is constitutively active and that the night blindness results from partial desensitization of rods caused by the constitutive activity.

High-risk characteristics of fellow eyes of patients with unilateral neovascular age-related macular degeneration.

OBJECTIVE: This study aimed to determine whether clinical tests of ocular function and macular appearance independently can help to predict which patients with unilateral neovascular age-related macular degeneration (AMD) will have a choroidal neovascular membrane (CNVM) develop in their fellow eye. DESIGN: The study design was a prospective cohort study. PARTICIPANTS: One hundred twenty-seven patients with unilateral neovascular AMD observed for up to 4.5 years participated. INTERVENTION: Functional measurements included visual acuity, macular visual field, glare recovery time, and foveal electroretinogram amplitude and implicit time. MAIN OUTCOME MEASURE: The age-adjusted proportion of patients having a CNVM develop over follow-up assessed by the Cox proportional hazards model with stepwise selection was measured. RESULTS: On average, 8.8% of patients had a CNVM develop each year. Independent risk factors for the fellow eye were its glare recovery time in minutes (relative risk = 1.30, confidence interval = 1.10-1.54, P = 0.003) and its extent of visible macular abnormalities on a four-point scale (relative risk = 1.62, confidence interval = 1.06-2.59, P = 0.03). Of the fellow eyes that converted, the interval to have a CNVM develop was inversely related to the foveal electroretinogram implicit time. CONCLUSIONS: A slower recovery from glare and more extensive funduscopic changes appear to be independent risk factors for the development of a CNVM in the fellow eyes of patients with unilateral neovascular AMD. A slower foveal electroretinogram implicit time may be a sign of early stage CNVM development, perhaps because of outer retinal ischemia. These results have clinical management implications, particularly for those patients at high risk of having a potentially treatable form of AMD develop.

13-cis-retinoic acid in silicone-fluorosilicone copolymer oil in a rabbit model of proliferative vitreoretinopathy.

The purpose of this study was to evaluate the effect of 13-cis-Retinoic Acid (RA) in Silicone-Fluorosilicone Copolymer Oil (SiFO) in a rabbit model of proliferative vitreoretinopathy (PVR). Rabbits underwent gas-compression vitrectomy. During gas-SiFO exchange, group 1 was injected with 1 ml (10 microg ml-1) 13-cis-RA in SiFO, group 2 with 1.5 ml (9 microg 1.5 ml-1) all-trans-RA in SiFO, group 3 with 1 ml SiFO alone, and group 4 with balanced salt solution (BSS). Groups 1-4 were also injected with 0.1 ml suspension of fibroblasts (75,000 0.1 ml-1) and 0.05 ml platelet rich plasma (70,000 0.1 ml-1), and were observed for 4 weeks. Group 5 was injected with SiFO alone, group 6 with 1 ml (10 microg ml-1) 13-cis-RA in SiFO, group 7 with 1.5 ml (9 microg 1.5 ml-1) all-trans-RA in SiFO, and group 8 with BSS. After 4 weeks, groups 5-7 underwent SiFO-BSS exchange. ERG and histopathology were performed to test for retinal toxicity in groups 5-8. The incidence of traction retinal detachment at 4 weeks was: group 1, 42.9%; group 2, 36.4%; group 3, 87.5%; and group 4, 88.9%. A significant difference in the incidence of PVR was noted between treated eyes (groups 1 and 2) and control eyes (groups 3 and 4) at 2, 3, and 4 weeks (P<0.05). No significant difference in the incidence of PVR was found between groups 1 and 2 during the same observation periods. ERG and histopathological studies showed no differences between the treated and the control fellow eyes (group 5-7) after 4 weeks. 13-cis-RA in SiFO (10 microg ml-1) is as effective as all-trans-RA in SiFO (9 microg 1.5 ml-1) in controlling the incidence of PVR when used for short term retinal tamponade and does not appear to be associated with retinal toxicity.

The relationship between visual field size and electroretinogram amplitude in retinitis pigmentosa.

PURPOSE: To determine to what degree visual field size is correlated with electroretinogram (ERG) amplitude among patients with the common forms of retinitis pigmentosa (RP). METHODS: Visual field equivalent diameter to the V4e white test light of the Goldmann perimeter was correlated with log ERG amplitude elicited by 0.5 Hz or 30 Hz full-field flashes of white light. Primary analyses were conducted on data from 583 patients with the common forms of RP. Subset analyses were performed on data from patients with ERG responses with different ranges of amplitude to assess to what extent the correlation depends on ERG amplitude, as well as on data from patients of a given genetic type to determine whether the correlation depends on the mode of transmission. Data from patients with the rhodopsin, Pro23His mutation (n = 38) or with the rhodopsin, Pro347Leu mutation (n = 24) were analyzed to determine the correlation between visual field size and ERG amplitude for patients with the same mutation. RESULTS: Visual field size was significantly correlated with ERG amplitude for every comparison (P < or = 0.0003). Correlations generally were higher for ERGs elicited by 30 Hz flashes (r = 0.62 for the entire sample) than they were for those elicited by 0.5 Hz flashes (r = 0.53 for the entire sample). They were lower for truncated ranges of ERG amplitude, higher for patients with dominant or recessive disease than for patients with x-linked disease or for patients of all genetic types combined, and strong for patients with the same rhodopsin mutation (reaching a value of 0.87). CONCLUSIONS: Visual field size is significantly correlated with ERG amplitude for patients with RP. Correlation depends on the range of ERG amplitudes, the inheritance type, and, particularly, on whether the analysis is confined to a single gene mutation.

Evaluation of patients with retinitis pigmentosa receiving electric stimulation, ozonated blood, and ocular surgery in Cuba.

OBJECTIVE: To evaluate the effect of intervention with electric stimulation, autotransfused ozonated blood, and ocular surgery, performed in Cuba, on the course of the common forms of retinitis pigmentosa. DESIGN: Ocular evaluations over 6 to 8 months before and after intervention in Cuba. SETTING: Evaluations performed at a US clinical research facility. PATIENTS: Ten adult patients aged 25 to 67 years with retinitis pigmentosa. MAIN OUTCOME MEASURES: Visual acuity, visual field area, and electroretinogram (ERG) amplitude. RESULTS: No significant change in visual acuity or visual field area was observed on average between preintervention and postintervention values over a 6- to 8-month interval. Mean 30-Hz cone ERG amplitude declined by 15.5% between preintervention and postintervention values (P = .006). When data on change in visual field area from 1 statistically significant outlier were excluded from the analysis, a significant decline of 12.9% in mean visual field area was observed (P = .025). CONCLUSIONS: These data support the conclusion that the intervention offered in Cuba provides no benefit to patients with retinitis pigmentosa as measured by visual acuity, visual field area, and ERG. The magnitudes of the mean declines observed in ERG amplitude and visual field area over a 6- to 8-month interval, relative to those reported in previous studies, raise the possibility that this intervention may worsen the course of the disease.

Isolation of focal rod electroretinograms from the dark-adapted human eye.

PURPOSE: To isolate focal rod electroretinograms (ERGs) from the dark-adapted human eye. METHODS: In two normal volunteers, dark-adapted focal rod ERGs were recorded from the peripheral retina in response to 30 degree diameter blue flashes of varying retinal illuminance and from different retinal regions in response to 10 degree diameter bright blue flashes. Dark-adapted focal rod ERGs also were recorded from a patient with the multiple evanescent white dot syndrome (MEWDS) and an enlarged blind spot in response to 30 degree diameter blue flashes presented within and outside the scotoma. The slower and larger stray light rod component elicited by these flashes was removed by subtracting the matching rod response to a dimmer, full-field flash, or was ignored when it did not overlap the faster and smaller focal rod component. RESULTS: The focal rod ERG had a waveform and sensitivity similar to those of the full-field rod ERG, was approximately proportional in amplitude to the density of rods directly illuminated, and was nondetectable within the retinal area corresponding to the enlarged blind spot of the patient with MEWDS. CONCLUSIONS: Focal rod ERG a- and b-waves, in response to stimuli as small as 10 degrees, can be recorded from different regions of the dark-adapted human retina to evaluate localized rod function.

Submicrovolt full-field cone electroretinograms: artifacts and reproducibility.

The clinical utility of submicrovolt full-field 30-Hz (cone) electroretinograms was assessed by quantifying their contamination by electrical and photoelectric artifacts from xenon-flash stimulators and their test-retest variation in patients with retinitis pigmentosa. Artifacts obtained in saline with four commonly used electrodes varied with electrode type and consisted of an early, brief electrical component and a superimposed, extended photoelectric component. Techniques for minimizing these artifacts are described. Electroretinogram recordings from patients with advanced retinitis pigmentosa or congenital rod monochromatism indicate that these artifacts can be virtually eliminated with bipolar lenses. To assess test-retest variation, narrow-band-filtered responses were obtained twice during 6 weeks from patients with amplitudes less than 1 microV; threshold criteria for significant (p < 0.05) change in amplitude with this technique were approximately 0.25 log unit for each of two different systems.

Low incidence of retinitis pigmentosa among heterozygous carriers of a specific rhodopsin splice site mutation.

PURPOSE: To determine whether a rhodopsin splice donor site mutation at the 5' end of intron 4 is a cause of autosomal dominant retinitis pigmentosa. METHODS: Heterozygous carriers of the same rhodopsin splice site mutation in two pedigrees were identified using single-strand conformation polymorphism analysis. Twelve heterozygous carriers were evaluated by ophthalmoscopy. Goldmann kinetic visual fields, dark adaptation thresholds, and full-field electroretinograms including rod intensity-response functions. Clinical findings from the heterozygous carriers of the splice site mutation were compared with those from heterozygous carriers from a separate family with a known recessive rhodopsin null mutation, Glu249X. RESULTS: Analysis of DNA from 48 members of two pedigrees revealed 25 heterozygous carriers of the splice site mutation, ranging in age from 14 to 82 years. There were no homozygotes with the rhodopsin splice site mutation. Of the 25 heterozygous carriers, 24 were asymptomatic. Eleven asymptomatic heterozygotes were examined, including four older than 65 years of age. They were found to have normal fundi, full visual fields, and slightly elevated final rod dark adaptation thresholds. Their rod electroretinographic b-wave amplitudes were slightly diminished over the full range of blue light intensities. Rod a-wave implicit times were slightly but significantly prolonged in response to the brightest blue flash of light. These subtle abnormalities in rod function were similar to those found in asymptomatic heterozygous carriers of the recessive Glu249X mutation. Only one of the 25 heterozygous carriers of the splice site mutation had symptoms and signs of retinitis pigmentosa. CONCLUSIONS: Because 96% of these heterozygous carriers do not have retinitis pigmentosa, it is unlikely that this mutation in intron 4 is a dominant allele. The subtle abnormalities of rod function found in asymptomatic carriers are similar to those found in heterozygous carriers of a recessive rhodopsin allele. The one heterozygous carrier with retinitis pigmentosa probably has a second mutation in the rhodopsin gene or has a defect or defects in another gene that causes his disease.

Clinical expression correlates with location of rhodopsin mutation in dominant retinitis pigmentosa.

PURPOSE: To determine whether severity of retinitis pigmentosa caused by dominant rhodopsin mutations depends on the location altered by the mutation. METHODS: Data from 128 patients (age range, 7 to 73 years), each with 1 to 27 rhodopsin mutations, were analyzed. To approximate normal distributions, visual acuities were converted to ranks and then to the normal form, kinetic visual fields to a V4e test light were converted to equivalent diameters, and dark-adapted sensitivities to an 11 degrees diameter stimulus and electroretinogram (ERG) amplitudes to full-field 0.5-Hz and 30-Hz flashes were converted to common logarithms. Each of these measures was then regressed on age, refractive error (for the ERG), and domain (intradiscal, transmembrane, or cytoplasmic) or codon number of the opsin molecule altered by the mutation. RESULTS: All five measures of function varied significantly with the domain (P < or = 0.0007) or codon number (P < 0.0001) altered by a mutation; visual acuity, visual field diameter, dark-adapted sensitivity, and ERG amplitudes were highest for mutations altering the intradiscal domain or low-numbered codons and lowest for mutations altering the cytoplasmic domain or high-numbered codons. CONCLUSIONS: These data indicate that severity of disease correlates with the location of the amino acid residue altered by a rhodopsin mutation in dominant retinitis pigmentosa.

Red blood cell membrane phosphatidylethanolamine fatty acid content in various forms of retinitis pigmentosa.

In order to test the hypothesis that retinitis pigmentosa (RP) is associated with fatty acid abnormalities within cell membrane phospholipids, red blood cell membrane (RBC) phosphatidylethanolamine (PE) fatty acid content (% of total fatty acids) was measured using high performance liquid chromatography and capillary column gas chromatography in 155 patients from separate families with different genetic types of RP and 101 normal subjects. After controlling for the effects of age and sex, patients with all genetic forms of RP had significantly (P < 0.001) reduced mean RBC PE 22:6 omega 3 (n-3) (docosahexaenoic acid, DHA) content, and significantly (P < 0.001) elevated mean RBC PE dimethyl acetal (DMA) forms of 16:0, 18:0, and 18:1 omega 9 (n-9) as compared with normal subjects. RBC PE content of 22:5 omega 3 (n-3) (a precursor to DHA) and 18:2 omega 6 (n-6) (the major dietary essential fatty acid) were not significantly different in RP than in controls. Analysis by genetic types of RP showed that the mean RBC PE DHA percentages were significantly reduced by 24%, 14%, 30%, and 17%, respectively, in dominant, recessive, X-linked, and isolate forms of RP. The relative amounts of plasmalogens as indicated by DMA forms of 16:0 and 18:0 were significantly (P < 0.01) increased in dominant (by 33% and 25%), recessive (by 36% and 25%), and isolate cases (by 32% and 26%) of RP as compared with normal subjects. No such differences were seen in X-linked cases versus controls. Our data indicate that RBC PE DHA content is decreased in all genetic types of RP patients as compared to control subjects, and that RBC PE plasmalogens are increased in dominant, recessive, and isolate forms of RP. These data raise the possibility that membrane phospholipid fatty acid abnormalities may contribute to the pathogenesis of RP.

Foveal electroretinograms and choroidal perfusion characteristics in fellow eyes of patients with unilateral neovascular age-related macular degeneration.

AIMS/BACKGROUND: A prolonged choroidal filling phase on fluorescein angiography has been reported to be a common finding and associated with visual function abnormalities in patients with age-related macular degeneration (AMD). This investigation was carried out to determine whether this perfusion defect was related to the slowing of foveal cone electroretinogram (ERG) implicit time seen in patients with AMD. METHODS: Fluorescein angiograms and foveal cone ERGs were evaluated in the fellow eyes of 67 patients with unilateral neovascular AMD. RESULTS: Twenty eight (42%) of the eyes had a choroidal perfusion defect. ERG implicit times averaged 1 ms slower (p = 0.0167) and were more likely to be delayed (p = 0.0078) in eyes with abnormal choroidal perfusion than in eyes with normal choroidal filling; significant relations were found also after controlling for age. ERG implicit time was also inversely related to ETDRS visual acuity and positively related to the extent of macular drusen; and the latter showed a borderline significant tendency to be more prevalent in eyes with prolonged choroidal perfusion. However, an association of a delayed ERG implicit time with prolonged choroidal filling remained after controlling for age, acuity, and the extent of drusen. CONCLUSION: These findings further establish prolonged choroidal perfusion as a common finding in AMD and link it to retinal malfunction.