Ocular infection with a murine neurovirulent retrovirus does not cause retinal degeneration.

The developing eyes of CFW/D mice inoculated at birth with a neurovirulent mutant (ts1) of Moloney murine leukemia virus (MoMuLV), nonneurovirulent wild type (wt) MoMuLV, and conditioned virus-free medium were studied comparatively by immunohistochemistry, lectin histochemistry and light microscopy. Cellular targets for viral antigen expression in the eye were identical in both ts1 and wt MoMuLV-infected mice. Viral antigen first was observed in endothelial cells of the retina and subsequently spread in a spatial and temporal pattern consistent with normal vascularization of the developing retina. The virus also was observed in (1) epithelial cells of the bulbar and palpebral conjunctiva, ora ciliaris retinae, and lacrimal gland; (2) endothelial cells of the ciliary body, iris, choroid, and sclera; (3) amacrine cells of the retina; and (4) smooth muscle cells and endothelia of the periocular muscle. Although ts1 MoMuLV induced a spongiform encephalopathy in the brain and spinal cord, structural lesions were not observed in the retina or other ts1 MoMuLV-infected ocular structures; differentiation of the retina was normal. The lectin Ricinus communis agglutinin-I (RCA-I) labeled (1) endothelial cells of the hyaloid vessels, tunica vasculosa lentis, retina, ciliary body, iris, choroid, and sclera; (2) epithelial cells of the cornea, bulbar and palpebral conjunctiva, ora ciliaris retinae, and lacrimal gland; (3) smooth muscle cells and endothelia of the periocular muscle; (4) inner segments of the photoreceptor layer; and (5) amacrine cells of the retina.(ABSTRACT TRUNCATED AT 250 WORDS)     

Elovl4 haploinsufficiency does not induce early onset retinal degeneration in mice

ELOVL4 was first identified as a disease-causing gene in Stargardt macular dystrophy (STGD3, MIM 600110.) To date, three ELOVL4 mutations have been identified, all of which result in truncated proteins which induce autosomal dominant juvenile macular degenerations. Based on sequence homology, ELOVL4 is thought to be another member within a family of proteins functioning in the elongation of long chain fatty acids. However, the normal function of ELOVL4 is unclear. We generated Elovl4 knockout mice to determine if Elovl4 loss affects retinal development or function. Here we show that Elovl4 knockout mice, while perinatal lethal, exhibit normal retinal development prior to death at day of birth. Further, postnatal retinal development in Elovl4 heterozygous mice appears normal. Therefore haploinsufficiency for wildtype ELOVL4 in autosomal dominant macular degeneration likely does not contribute to juvenile macular degeneration in STGD3 patients. We found, however, that Elovl4+/- mice exhibit enhanced ERG scotopic and photopic a and b waves relative to wildtype Elovl4+/+ mice suggesting that reduced Elovl4 levels may impact retinal electrophysiological responses.     

Cdh23 mutations in the mouse are associated with retinal dysfunction but not retinal degeneration

Mutations in the cadherin 23 gene (CDH23) cause Usher syndrome type 1D in humans, a disease that results in retinitis pigmentosa and deafness. Cdh23 is also mutated in the waltzer mouse. In order to determine if the retina of the waltzer mouse undergoes retinal degeneration and to gain insight into the function of cadherin 23 in the retina, we have characterized the anatomy and physiology of retinas of waltzer mouse mutants. Three mutant alleles of Cdh23 were examined by histology and electroretinography (ERG). ERGs of the three Cdh23 mutant groups revealed two of them to have abnormal retinal function. One allele had a- and b-waves that were only approximately 80% of Cdh23 heterozygotes. Another allele had a significantly faster implicit time for both the a- and b-waves of the ERG. No anatomical abnormality was detected in any of the Cdh23 mutants by light microscopy. Because the mutant Cdh23 phenotype was found to be similar to the previously reported retinal phenotype of Myo7a mutant mice, the orthologue of another Usher syndrome (type 1B) gene, we generated mice that carried mutations in both genes to test for genetic interaction in the retina. No functional interaction between cadherin 23 and myosin VIIa was detected by either microscopy or ERG.     

Rod photoreceptor maturation does not vary with retinal eccentricity in mammalian retina.

PURPOSE: Test the hypothesis that the development of mammalian rod outer segments (ROS) varies with retinal eccentricity. METHODS: During the period of photoreceptor cell development, ROS lengths, opsin mRNA and (rhod)opsin were measured in central and peripheral retina of cows and pigmented rats. Published ROS length and/or rhodopsin data from albino rats, cows and monkeys were re-analyzed. Logistic growth curves were fitted to the newly obtained and published data. Within a species, growth in central and peripheral regions was compared. RESULTS: The logistic growth curves fit all the data well and provide an excellent view of the developmental increases in ROS length, opsin mRNA and (rhod)opsin in each retinal region. Within a species, the growth curves for ROS length, opsin mRNA and (rhod)opsin concentration are superimposable. The age at which ROS length reaches 50% of its adult value is invariant with eccentricity. An exception to this pattern is the simian parafoveal ROS, which appears to have a delayed course of development. CONCLUSIONS: The hypothesis is disproved. Unlike rod photoreceptor cell genesis, ROS development is invariant with retinal eccentricity. Primate parafoveal ROS appear to have a different pattern of development.     

Prevalence of 2314delG mutation in Spanish patients with Usher syndrome type II (USH2)

The Usher syndrome (USH) is a group of autosomal recessive diseases characterized by congenital sensorineural hearing loss and retinitis pigmentosa. Three clinically distinct forms of Usher syndrome have so far been recognized and can be distinguished from one another by assessing auditory and vestibular function. Usher syndrome type II (USH2) patients have congenital moderate-to-severe nonprogressive hearing loss, retinitis pigmentosa, and normal vestibular function. Genetic linkage studies have revealed genetic heterogeneity among the three types of USH, with the majority of USH2 families showing linkage to the USH2A locus in 1q41. The USH2A gene (MIM 276901) has been identified: three mutations, 2314delG, 2913delG, and 4353-54delC, were initially reported in USH2A patients, the most frequent of which is the 2314delG mutation. It has been reported that this mutation can give rise to typical and atypical USH2 phenotypes. USH2 cases represent 62% of all USH cases in the Spanish population, and 95% of these cases have provided evidence of linkage to the USH2A locus. In the present study, the three reported mutations were analyzed in 59 Spanish families with a diagnosis of USH type II. The 2314delG was the only mutation identified in our population: it was detected in 25% of families and 16% of USH2 chromosomes analyzed. This study attempts to estimate the prevalence of this common mutation in a homogeneous Spanish population.     

Prevalence of 2314delG mutation in Spanish patients with Usher syndrome type II (USH2)

The Usher syndrome (USH) is a group of autosomal recessive diseases characterized by congenital sensorineural hearing loss and retinitis pigmentosa. Three clinically distinct forms of Usher syndrome have so far been recognized and can be distinguished from one another by assessing auditory and vestibular function. Usher syndrome type II (USH2) patients have congenital moderate-to-severe nonprogressive hearing loss, retinitis pigmentosa, and normal vestibular function. Genetic linkage studies have revealed genetic heterogeneity among the three types of USH, with the majority of USH2 families showing linkage to the USH2A locus in 1q41. The USH2A gene (MIM 276901) has been identified: three mutations, 2314delG, 2913delG, and 4353-54delC, were initially reported in USH2A patients, the most frequent of which is the 2314delG mutation. It has been reported that this mutation can give rise to typical and atypical USH2 phenotypes. USH2 cases represent 62% of all USH cases in the Spanish population, and 95% of these cases have provided evidence of linkage to the USH2A locus. In the present study, the three reported mutations were analyzed in 59 Spanish families with a diagnosis of USH type II. The 2314delG was the only mutation identified in our population: it was detected in 25% of families and 16% of USH2 chromosomes analyzed. This study attempts to estimate the prevalence of this common mutation in a homogeneous Spanish population.     

The electroretinogram components in Abyssinian cats with hereditary retinal degeneration

PURPOSE: To examine phototransduction using the a-wave and other aspects of retinal function with the intraretinal b- and c-waves at different stages of an inherited photoreceptor degeneration in Abyssinian cats. METHODS: Vitreal and intraretinal ERGs were recorded from eight dark-adapted, anesthetized Abyssinian cats. Brief bright flashes were used to elicit vitreal a- and b-waves. Longer, weaker flashes were used to elicit intraretinal b- and c-waves. Stages 1 through 4 of the disease were characterized ophthalmoscopically. Parameters of the Lamb and Pugh a-wave model (a(max), A, and t(eff)) for the Abyssinian cats were compared with those for normal cats. Light microscopy was used to count photoreceptor nuclei. RESULTS: The maximum a-wave amplitude, a(max), was significantly smaller in stage 1, and continued to decrease (stage 1: 50% of normal, stage 2: 28%, stage 3: 27%; and stage 4: unrecordable). There was a small, but not significant, decrease in the amplification constant A from 0.24 +/- 0.11 s(-2) in normal cats to 0.16 +/- 0.08 s(-2) in Abyssinian cats. The intraretinal b- and c-wave amplitudes decreased most dramatically during the early stage of the disease. Affected animals had fewer photoreceptors than unaffected Abyssinians or control animals. The number of photoreceptors declined most rapidly in the inferior periphery. CONCLUSIONS: The amplitudes of all ERG components were already reduced significantly by stage 1 and progressively declined. The lack of major changes in a-wave model parameters indicates that the degeneration is probably not due to a mutation in transduction proteins. Losses of photoreceptor function were larger than losses of photoreceptor nuclei.     

Angiotensin II does not contract bovine retinal resistance arteries in vitro

The effect of angiotensin II was studied in vitro on ring segments of bovine retinal resistance arteries (i.d. 126-271 microns) and posterior ciliary arteries (i.d. 207-1153 microns). Although the retinal resistance arteries were responsive to 5-hydroxytryptamine, prostaglandin F2 alpha, and changes in extracellular K(+)-concentration, they did not, in contrast to the posterior ciliary arteries, contract to cumulative or single doses of angiotensin II. In the latter arteries, angiotensin II induced a small concentration dependent contraction, 5% of maximal 125 mM K(+)-induced response, with a pD2-value of 9.3. The single addition of 10(-6) M angiotensin II increased the maximal vessel response of the posterior ciliary arteries three times to angiotensin II. Tachyphylaxis was pronounced in the posterior ciliary arteries, in which the response to angiotensin II could not be repeated. Indomethacin (10(-5) M), methylene blue (3 x 10(-6) M), or removal of endothelium did not make the retinal resistance arteries responsive to angiotensin II. Retinal arteries precontracted with 30 mM potassium did not respond to angiotensin II. Angiotensin II did not potentiate the 5-hydroxytryptamine- and noradrenaline concentration-response characteristics of both retinal resistance and posterior ciliary arteries. Although angiotensin II-receptors have been detected in bovine retinal vascular smooth muscle using radioligand-binding technique, the present results suggest that these receptors are non-functional in respect to regulation of retinal resistance artery tone.     

Permeability of retinal capillaries in rats with inherited retinal degeneration.

The permeability of retinal capillaries in RCS rats with inherited retinal degeneration was investigated with horseradish peroxidase used as a tracer. Five-week-old rats showed typical degeneration of photoreceptor cells and accumulation of outer segment debris, but retinal capillaries were not permeable to peroxidase. At 10 weeks of age, capillaries in the inner retina appeared normal, but many in the outer retina were leaky. Peroxidase activity in these latter vessels was demonstrable in the basal laminae of endothelial cells and pericytes and in vesicles on the luminal and abluminal sides of the endothelium. Tracer also permeated the intercellular spaces in the surrounding area. The number of leaky capillaries in the outer retina increased during the course of the dystrophy. The site of the leak in permeable capillaries has not yet been established; it may be due to an alteration of the endothelial cell junctions or of transcellular vesicular transport. In the choriocapillaris, peroxidase permeated Bruch's membrane and the basal infoldings between adjacent pigment epithelial cells; tracer progression along the intercellular spaces was blocked at the zonulae occludens at the apicolateral border. The RCS rat may be a useful model for studying the morphological basis of changes in capillary permeability associated with retinal degeneration.     

Pigmentary retinal degeneration in patients with HTLV-I-associated myelopathy

Ophthalmological evaluations were made of the records of a series of 38 patients with HTLV-I-associated myelopathy, a chronic progressive myelopathy caused by human T-lymphotropic virus type I (HTLV-I). Four patients with no contributory family history showed pigmentary degenerative changes of the retina and choroid. Two of the patients (73-year-old woman, 68-year-old woman) had a progressive visual loss and night blindness with morphologic and functional features of diffuse pigmentary retinal degeneration. The other two patients (59-year-old man, 72-year-old man) complained of recently developed visual loss with sectorial or regional retinochoroidal atrophy. These elderly patients claimed that they had been healthy until a few years before presentation, not only visually but also neurologically. It was concluded, together with an epidemiologic consideration, that the coexistence of pigmentary retinal degeneration and HTLV-I-associated myelopathy is not simply chance but indicates a close association between the two conditions. It is proposed that HTLV-I infection might be a primary causative factor of degenerative changes of the retina and choroid, although the pathogenesis remains to be defined.     

The two-stage mutation model in retinal hemangioblastoma

Background: The two-stage mutation model involving successive inactivation of both alleles of a tumor suppressor gene was originally proposed by Knudson, who analyzed the age incidence curves for unilateral and bilateral retinoblastoma, and suggested that hereditary tumors arise by a single somatic event superimposed on a defective genetic background and sporadic tumors by a two-stage somatic process. In this study, the age-incidence curve of patients with retinal hemangioblastoma with and without associated von Hippel-Lindau disease were analyzed. Methods: We reviewed the literature between 1964 and 1998 to find all reported cases of retinal hemangioblastoma and classified patients in a type A group (n=223) when associated with von Hippel-Lindau disease and a type B group (n=30) when not associated with von Hippel-Lindau disease. We analyzed and compared the age incidence of these two groups. Results: There was a statistically significant difference between the mean age at diagnosis of retinal hemangioblastoma in the two groups, i.e., 48.4±16.6 years for type B patients and 24.9±12.0 years for type A patients (p=0.0001). The age incidence curve for type A retinal hemangioblastoma fit a first-order equation (log S=0.411-0.034t) with r=0.97, indicating a single somatic mutation, whereas the age incidence curve for type B retinal hemangioblastoma fit a second-order equation (log S=0.184-2.25×10-4t2) with r=0.97, indicating two somatic mutations. Conclusions: Type B (sporadic) retinal hemangioblastoma may arise from two separate somatic mutations inactivating both alleles at the von Hippel-Lindau locus, whereas patients with von Hippel-Lindau disease (type A) inherit a defective allele and require only one additional somatic mutation.     

CRX基因突变影响视紫红质转录表达和NRL蛋白稳定性的细胞学研究

目的在细胞水平上,探讨存在和缺乏神经视网膜亮氨酸拉链（NRL）的条件下,视锥-视杆细胞同源异形框基因（CRX）突变型对调控视紫红质基因转录表达的影响,并分析CRX突变型对CRX和NRL蛋白稳定性的影响。方法实验研究。分别构建携带野生型和c.C766T（p.Gln256X）无义突变型的CRX基因,以及共表达NRL基因和牛视紫红质基因启动子萤光素酶（pBR130-luc）的表达载体,分别转染体外培养的293T细胞和ARPE-19细胞,再行双荧光素酶检测分析和Western blotting实验。以持家蛋白GAPDH作为内参照。结果与野生型CRX蛋白的表达导致牛视紫红质启动子表达5倍的激活率相比,CRX/c.C766T（p.Q256X）突变所造成的相应激活率降至1/4。共表达野生型CRX和NRL基因后,激活率增高至30倍,而共转染CRX/c.C766T和NRL后,牛视紫红质蛋白的活性降至1/13。以持家蛋白GAPDH作为内参,可见存在CRX/c.C766T突变时,CRX蛋白的稳态水平大幅下降,且CRX/c.C766T突变型对NRL蛋白的稳态有一定的影响,出现NRL增高的现象。结论在细胞水平上,CRX基因的c.C766T（p.Q256X）突变可降低所调控的视紫红质蛋白的表达,并对CRX蛋白和NRL蛋白的稳态均造成一定的影响。     

The phenotype of early-onset retinal degeneration in persons with RDH12 mutations

PURPOSE: To describe the retinal dystrophy phenotype associated with mutations in RDH12, the gene encoding a retinoid dehydrogenase/reductase expressed in the photoreceptor cells. METHODS: Sixteen persons from 12 families with pathogenic RDH12 mutations on both alleles were studied. Retinal phenotypes were characterized by ophthalmic examination, including psychophysical and standardized electrophysiological methods and multifocal electroretinography (mfERG). RESULTS: The retinal disease in persons with RDH12 mutations in the homozygous (p.G127X, p.Q189X, p.Y226C, p.A269GfsX1, and p.L274P) or compound heterozygous state (p.R65X/p.A269GfsX1, p.H151D/p.T155I, p.H151D/p.A269GfsX1) was diagnosed initially as Leber congenital amaurosis (LCA) or early-onset retinitis pigmentosa. These individuals appeared to share a common clinical picture, independent of the type of mutation, characterized by poor, yet useful visual function in early life, followed by progressive decline due to both rod and cone degeneration. Marked pigmentary retinopathy, including bone spicules in the peripheral retina, was present in all persons older than age 6, and pronounced maculopathy was evident in persons older than 7 years. A unique view into the progressive nature of the disorder was achieved by evaluation of seven affected persons from three consanguineous families, all carrying the homozygous p.Y226C mutation. CONCLUSIONS: Ophthalmic findings in persons with RDH12 mutations suggest that RDH12 loss-of-function results in a characteristic form of early and progressive rod-cone degeneration distinct from that caused by mutations in other LCA genes. From our data, it seems likely that various clinical designations appropriately describe the diagnosis in these persons, including early-onset retinitis pigmentosa, LCA type II, and childhood retinal dystrophy.     

Human diabetic serum does not stimulate bovine retinal endothelial cell growth in culture

Bovine retinal microvascular endothelial cells were cultured with serum from nondiabetic controls, patients with background, preproliferative, proliferative and inactive (treated) proliferative diabetic retinopathy. Using Coomassie Blue to measure cellular protein, we found no significant increase in retinal endothelial cell growth with serum from patients with preproliferative or proliferative retinopathy.