一个中国全色盲家系 CNGB3基因新突变

目的:探讨全色盲一家系的致病基因突变。方法:采用家系调查研究方法,于2018年11月对河南省立眼科医院收集的来自河南省洛阳市的汉族全色盲一家系进行基因测序。详细采集患者的病史资料,对患者及其家系成员进行最佳矫正视力(BCVA)测定,采用裂隙灯显微镜和前置镜检查眼前节和眼底,采用客观和主觉验光法对受检者进行屈光度检查,采...     

Macular maldevelopment in ATF6-mediated retinal dysfunction

ABSTRACT

Background: Achromatopsia has been previously associated with mutations in the ATF6 gene. Rod-monochromatism, foveal hypoplasia, and disruption of the subfoveal photoreceptor layer are described as phenotypical features. We report detailed structural and electrophysiological assessment of two patients from two families, one manifesting severe macular maldevelopment and one with foveal hypoplasia.

Materials and methods: The patients underwent a complete ophthalmic examination including electroretinography (ERG), spectral-domain optical coherence tomography (SD-OCT), fundus autofluorescence, and fundus photography. Genetic testing was performed by next-generation sequencing.

Results: In one patient, fundoscopy and SD-OCT revealed well-demarcated coloboma-like excavated lesions at the central macula of both eyes. Genetic analysis identified a novel homozygous p.Asp140Ter mutation in the ATF6 gene. The second patient had foveal hypoplasia in association with a homozygous ATF6 mutation affecting a splice donor site (c.1187 + 5G>C). In both patients, electrophysiological assessment showed normal rod-specific (DA 0.01) and dark-adapted bright white-flash ERGs (DA 10.0). 30 Hz flicker ERGs were undetectable. There were low-amplitude single-flash photopic ERGs (LA 3.0) with timing and shape suggesting S-cone origin.

Conclusions: The findings, particularly a case with severe macular maldevelopment, may expand on the phenotype previously associated with ATF6-mediated achromatopsia. In addition, the comprehensive electrophysiological assessment suggests that preserved S-cone activity can be detected in this particular molecular sub-type of cone dysfunction.     

An international standard for electroretinography

An international standard procedure for electroretinography (ERG) has finally been established. It represents a minimum protocol to record five basic signals (rod response, scotopic maximal response, oscillatory potentials, single-flash cone response, 30-Hz flicker response). Individual laboratories may continue to do additional or specialized procedures, but the hope is that all laboratories will include standard responses in their routine protocols so ERG data can be compared throughout the world for patient care and for research.Supported in part by NIH-NEI Research Grant EY01678.     

Novel causative variants in patients with achromatopsia

ABSTRACT

Purpose: To report five novel genetic variants in seven unrelated consanguineous families with achromatopsia (ACHM).

Methods: Patients were examined with multimodal retinal imaging and full-field electroretinography (ffERG). Genetic testing was conducted using next-generation sequencing (NGS).

Results: Three novel homozygous variants were detected in CNGA3: a missense c.967G > C (p.Ala323Pro) variant was detected in exon 8 (one patient), a splice site variant c.101 + 1G > A in intron 2 (three patients), and a splice site variant c.395 + 1G > T in intron 4(one patient). Another two novel variants were found in PDE6C: a homozygous missense variant c.1899C > A (p.His633Gln) in exon 15 (one patient) and a homozygous splice site variant c.1072-1G > C in intron 7 (one patient). Mutation segregation assessment was possible in 3 of the 7 families. All patients had nonrecordable ffERG 30-Hz flicker responses, reduced single-flash cone responses but preserved rod responses. Patients presented with variable degrees of foveal outer retinal layer loss and variable patterns of foveal hyperautofluorescence.

Conclusions: These novel variants expand the genotypes associated with ACHM and may help in future therapy development for ACHM.     

早发性严重视网膜营养不良一家系TULP1和CNGB1基因突变

目的:确定早发性严重视网膜营养不良（EOSRD）一家系的致病基因突变。方法:回顾性临床研究。2018年8月于河南省立眼科医院检查确诊的一个汉族EOSRD家系中1例患者及3名家系成员纳入研究。详细采集患者病史后,对受试者行最佳矫正视力（BCVA ）、裂隙灯显微镜联合前置镜、眼底彩色照相、频域光相干断层扫描（SD-OCT）...     

Development of a technique for recording the focal rod ERG

The scotopic ERG recorded in response to a focal stimulus has a double b-wave. The first wave results from direct focal stimulation of the retina, and the second originates in the peripheral retina in response to scattered light. The aim of this study was to assess two possible protocols for the isolation of the focal rod response, namely 'Subtraction' and 'Background Adaptation' techniques. The Subtraction technique involved the recording of a full-field response, which matched the b-wave elicited by intraocular light scatter, and the subtraction of this full-field response from the initial trace to isolate the focal component. In the Background Adaptation technique an adapting surround was used to suppress the response from the peripheral retina. Focal rod responses were isolated with both techniques. However, the Background Adaptation technique was found to more reliably elicit a focal response with a measurable a-wave, and was also considerably less time consuming than the Subtraction technique.     

Empiric limits of rod photocurrent component underlying a-wave response in the electroretinogram

The corneally recorded rod photocurrent component (photoresponse) underlying the a-wave feature of the electroretinogram was analyzed. The results set empiric limits on critical photoresponse variables. Measurements were obtained from four normal adult subjects on a-wave amplitude, a-wave velocity, b-wave amplitude, b-wave implicit time and b-wave height above baseline. At high intensity, interference from the b-wave component was minimized and the amplitude of the saturated photoresponse component was approximated by the a-wave feature. At lower intensities, the a-wave feature represented progressively less of the underlying photoresponse amplitude. Photoresponse amplitude saturation was signaled by the abrupt slowing of the rate of decline of b-wave peak latency and occurred at an intensity about 2.5 log units above the first appearance of the b-wave. At the intensity of photoresponse saturation, the peak amplitude of the a-wave feature was only about 25% of the maximum amplitude of the underlying photoresponse component. A-wave leading edge velocity was found to increase up to 3 log units above the intensity of photoresponse amplitude saturation and to provide a good estimate of photoresponse velocity at higher intensities. A cascaded low-pass filter model with modifications to accommodate amplitude and timing nonlinearities was used to generate a set of probable underlying photoresponses from the analysis of a-wave amplitude and velocity. Movement of the a-wave leading edge to the left at higher intensities in algebraic combination with a static b-wave leading edge above the intensity of photoresponse amplitude saturation was found to explain the second rise of the b-wave amplitude function and the decline of b-wave amplitude above baseline at high intensities. This analysis provides a basis for modeling the underlying photoresponse on a biochemical level and for interpreting photoreceptor damage in disease states.Supported in part by a grant from the Ethel Brown Foerderer Foundation, Children's Hospital of Philadelphia, and by a grant from the Nina and Paul MacKall Trust.     

Visual dysfunction in normotensive glaucoma

Color vision (desaturated D—15), contrast sensitivity (Vistech 6500) and pattern electroretinograms (transient and steady state) were measured in nine patients with diagnosed normotensive glaucoma. Results were compared to those from visually normal controls subjects (n = 73) and patients with primary open-angle glaucoma (n = 51). Patients with normotensive glaucoma exhibited significant contrast sensitivity and pattern electroretinogram deficits similar to those evident in patients with primary open-angle glaucoma. However, patients with normotensive glaucoma exhibited significantly better color vision than did patients with primary open-angle glaucoma. These results indicate that the pattern of visual impairment associated normotensive glaucoma and primary open-angle glaucoma is not identical. Different pathologic mechanisms mediating visual loss in the two diseases could explain these differences.Abbreviations ANOVA analysis of variance - CCS color confusion score - IOP intraocular pressure - NTG normotensive glaucoma - POAG primary open-angle glaucoma     

Comparison of conventional ERG parameters and high-intensity A-wave analysis in a clinical setting

Computational analysis of high-intensity a-waves yields direct information about the rod and cone receptor potential. However, it is not clear whether such information adds materially to the diagnostic value of the standard ERG in a routine clinical setting. We recorded both conventional ISCEV standard and computational high intensity ERG parameters from 38 patients referred to a clinical laboratory for ERG testing, and also from eight normal volunteers. The patients were grouped as: (1) macular dysfunction; (2) diffuse cone dysfunction; (3) diffuse rod-cone dysfunction. The results showed moderate variation in both conventional and computational parameters, but in general a similar pattern of normality or abnormality for both among the disease groups. There were only a few outlying subjects for which one or the other approach seemed more sensitive. We conclude that a-wave analysis is an important tool for clinical research and the study of special patients, but adding it to the standard ERG protocol does not, at our present state of knowledge, add markedly to clinical evaluations in a routine clinical setting.     

Rod and rod mediated function in patients with β-thalassemia major

An electroretinographic (ERG) study was undertaken to test the hypothesis that scotopic retinal function is altered in transfused thalassemics on chronic Deferoxamine (DFO). ERG a- and b-wave responses and dark adapted visual thresholds were obtained from 11 patients with β-thalassemia major, ages 7 to 38 (median 17) years. A quantitative model of the activation of phototransduction was fitted to the a-waves to estimate the gain of the transduction processes and the saturated amplitude of the rod photoresponse. From b-wave stimulus/response functions, the saturated b-wave amplitude and an index of b-wave sensitivity (log σ) were calculated. The patients' data were compared to those of normal subjects. The relations of the ERG parameters to age, average ferritin level, and duration of transfusion without DFO as well as other clinical parameters were examined. Longitudinal measures of b-wave responses and dark adapted visual thresholds, available for nine of the patients, were examined for significant change over time. For all patients both the gain and saturated amplitude of the rod response are normal. In two patients log σ is below the 99% prediction interval for normal. One has low scotopic visual sensitivity. The duration of transfusion therapy unprotected by DFO chelation therapy was correlated with log σ. These results suggest iron accumulation rather than DFO toxicity underlies scotopic dysfunction in older thalassemics, some of whom may have had extended periods of transfusion without the protection of chelation. Thus, monitoring of retinal function is recommended in such patients. This revised version was published online in July 2006 with corrections to the Cover Date.     

The local cone and rod system function in early age-related macular degeneration

To compare cone and rod system function in patients with early age- related macular degeneration (ARMD) and control group using multifocal electroretinogram (MERG) and perimetry, to investigate whether there is rod system dysfunction in the central retina in ARMD. Cone-mediated MERG, photopic sensitivity, rod-mediated MERG, and scotopic sensitivity in 16 eyes of control subjects and 24 eyes of early dry-form ARMD were measured with VERIS Science ^TM 4.0 and Octopus 101 perimetry. The latencies and average response densities of the summed responses and five ring retinal regions, average sensitivity of all locus and eight ring retinal regions in control eyes were compared with those in ARMD. Mean scotopic and photopic sensitivity of ARMD patients were significantly lower than that of normal controls. Sotopic sensitivity reduced more than photopic sensitivity and the greatest deficit was 2.5–5.0°. The amplitudes of N₁ and P₁ wave in one ring (5.0°) of rod MERG were significantly lower of ARMD patients than that of normal subjects. Our results suggest that rod function decreased and the parafoveal rod cells were predominantly damaged in ARMD. The rod function testing in macula may be a useful tool to diagnose and measure the fundus dysfunction of ARMD.     

重度非增生型糖尿病视网膜病变mfERG的定量分析

目的研究重度非增生型糖尿病视网膜病变（NPDR）多焦视网膜电图（mfERG）的特征及临床意义。方法30例（40眼）重度NPDR患者为NPDR组和35例（35眼）正常人为对照组。以国际分期作为NPDR诊断纳入标准,mfERG记录过程遵循国际临床视觉电生理学会的标准化方案,每个受试者在接受检查前均取得知情同意。结果与对照组相比,NPDR组患者mfERG2～5环的P1波、N1波反应密度明显下降,差异均有统计学意义（P〈0.05～0.01）;mfERG第Ⅱ象限和第Ⅲ象限的P1波、N1波反应密度明显降低,差异均有统计学意义（P〈0.05～0.01）。NPDR患者mfERG3～5环P1波、N1波隐含时较对照组明显延长,差异均有统计学意义（P〈0.05～0.01）;第Ⅰ象限和第Ⅲ象限隐含时显著延迟,差异均有统计学意义（P〈0.05～0.01）。结论NPDR可导致视网膜黄斑区视功能的损伤,mfERG能够客观、定量地反映黄斑区功能损害的程度。     

A new cause for difficulty in seeing at night

A syndrome is described in which patients complain of inability to see at night, although their rod and cone thresholds are normal. In such persons the rods, when dark-adapted, elevate cone flicker thresholds by 2 log units or more, and this condition is the basis of their complaint. Patients are often thought to be hysterics or to have congenital stationary night blindness.     

眼球钝挫伤ERG与病理改变的关系

检查轻，重度兔眼挫伤后ＥＲＧａ，ｂ波及ＯＰｓ波的改变，用镧示踪法观察血视外屏障及其它组织结构改变。轻型挫伤ＥＲＧｂ波暂时降低，无示踪剂渗漏，光感受器外节轻度破坏。重型挫伤ＥＲＧａ，ｂ波及ＯＰｓ波明显降低，视风膜色素上皮细胞及外层视网膜细胞严得损害，伴有血视网膜外屏障破坏。结果提示眼球挫伤后视力的预后和组织损伤的程度有直接关系。     

Deletional analysis of the rod photoreceptor cell peripherin/RDS carboxy-terminal region

The C-terminal region of peripherin/rds contains three predicted alpha-helical domains. One of these domains, corresponding to amino acids 311-322, form an amphiphilic alpha-helix previously shown to promote membrane fusion. The present studies were conducted to determine how the additional alpha-helical regions of the peripherin/rds C-terminus affect complex formation with rom-1, glycosylation, intracellular localization and membrane fusion properties. Bovine peripherin/rds and rom-1 were epitope tagged with an amino-terminal FLAG-tag or amino-terminal hemagglutinin (HA)-tag, respectively, and cloned into the pCI-neo expression vector for transient transfection into COS cells. Similarly, four C-terminal peripherin/rds truncation mutants (Delta1, Delta2, Delta3 and Delta4), corresponding to deletions of -19, -29, -39 and -59 amino acids were designed to disrupt the alpha-helical domains. Immunofluorescence microscopy and enzymatic digestions demonstrated that full-length peripherin/rds and the four C-terminal deletion mutants were localized to intracellular membranes and were all Endo-H sensitive. Western blotting and immunoprecipitation studies showed that the FLAG-tagged bovine peripherin/rds (full-length) was expressed as a 76kDa dimer, which associates with HA-tagged rom-1 to form a higher order complex. The deletion mutants were also able to associate with rom-1. However, when analyzed using non-denaturing tricine electrophoresis, full-length peripherin/rds and the Delta1, Delta2 and Delta3 mutants formed homo-oligomeric complexes, while the Delta4 mutant appeared to form only homodimers suggesting a region upstream of amino acid 300 may be involved in C-terminal interactions. Membrane fusion was then evaluated using fluorescence resonance energy transfer (RET) techniques. Intracellular COS cell membranes containing full-length peripherin/rds fused with rod outer segment plasma membrane vesicles. This fusion was inhibited with the addition of a synthetic peptide (PP-5) corresponding to the fusion domain of peripherin/rds. In contrast, fusion was negligible with any of the C-terminal truncation mutants. Collectively, these results suggest that in addition to the fusion domain, other regions of the peripherin/rds C-terminus are required for fusion. Most interesting is the observation that the last 19amino acids, a region downstream of the fusion peptide that is deleted in the Delta1 mutant, appear to be necessary for fusion. This region corresponds to the epitope for anti-peripherin/rds monoclonal antibody 2B6, which is shown to partially inhibit peripherin/rds mediated membrane fusion.