Objective To observe the clinical features, phenotypes and genotypes in a Chinese family with choroideremia (CHM). Methods A Chinese four-generation family (15 members) with CHM, including 5 patients (4 males/1 female), 2 female carriers and 8 healthy members, was enrolled in this study. Initially all family members underwent best corrected visual acuity (BCVA), indirect ophthalmoscopy, fundus fluorescein angiography, optical coherence tomography (OCT), visual field and full view electroretinogram (ERG). BCVA was followed up for 3 years. Venous blood samples were collected, and all of the 15 coding exons and flanking intron regions were amplified in the proband by polymerase chain reaction followed by direct sequencing. Protein structure was modeled based on the protein data bank and mutations in DeepView v4.0.1 to predict the effect of the mutations. A total of 180 healthy volunteers were enrolled as control group to matching CHM gene sequences. Results The visual acuity (VA) of 3/4 adult male patients began to decrease at less than 10, 10 and 30 years old, the average BCVA was 0.43. There were characteristic signs and symptoms of CHM including narrow visual field, extinguished rod and cone response in ERG, disappeared junction line and intermediate line of photoreceptor inner segment/outer segment on OCT. After 3 years, the mean BCVA decreased to 0.11. The BCVA of one young male patient was 1.0 in both eyes with minor changes fundus and visual field. The VA of the female patient began to decrease at 50 years old, her BCVA of two eyes were 0.5 and 0.25, respectively. The fundus changes were typical of CHM, with relative scotomas in the peripheral visual field of OD, and big scotomas in the OS. After 3 years, her mean BCVA decreased to 0.2. Of 2 female carriers, one had minor fundus changes (patches of pigmentary deposits, atrophy spots of retinal pigment epithelium cells), and the other was normal. A novel heterozygous c.1837G＞A mutation in exon 15 of CHM was detected in the proband, which resulted in the substitution of serine by proline at codon 613 (p.D613N). Based on molecular modeling, the misfolded protein caused by the mutation might destabilize the structure of the helix that potentially could affect the global stability of the Rep-1 protein. Conclusions A novel c.1837G＞A (p.D613N) mutation may be the causative mutation for CHM in this family. Female CHM carriers may have some signs and symptoms.
ObjectiveTo determine the pathogenic gene mutation in a family with incomplete congenital quiescent night blindness (CSNB) of Schubert-Bornschein type. MethodsA retrospective clinical study. In February 2021, one patient and his parents and elder brother from a Han Chinese incomplete CSNB of Schubert-Bornschein type family diagnosed by clinical and genetic examination at Henan Provincial People's Hospital were included in the study. The patient’s medical history, family history were inquired; best corrected visual acuity (BCVA), color vision, fundus color photography, full-field electroretinogram (ERG), and frequency domain optical coherence tomography (OCT) were examined in detail. Five ml of the subject’s peripheral venous blood was collected and the whole genome DNA was extracted. The genomic DNA of the subject was library constructed, and all-exon probes were polymerized for capture. The suspected pathogenic mutation site was verified by Sanger, and the pathogenicity of the gene mutation site was determined by parallel bioinformatics analysis. ResultsThe BCVA of both eyes of the proband (Ⅱ2) was 0.4; the color vision test could not recognize the red color. Fundus examination showed no obvious abnormalities. The retina thickness in the macular area of both eyes was slightly thinned. ERG examination of the whole field showed that the amplitude of ERG b wave was significantly reduced under the stimulation of binocular dark adaptation 3.0 and showed a negative waveform. The mother of the proband (Ⅰ2) had normal BCVA, color vision, fundus color photography, and frequency domain OCT examination. The full-field ERG examination showed that the amplitude of each eye reaction was slightly reduced, and the amplitude of the dark adaptation shock potential was significantly reduced. Genetic testing showed that the proband (Ⅱ2) had a c.1761dupC hemizygous mutation in exon 14 of the voltage-dependent calcium channel α1F subunit gene (CACNA1F gene). The results of protein sequence homology analysis showed that the site was highly conserved in multiple species; the results of bioinformatics analysis showed that the CACNA1F gene c.1761dupC (pY588fs) subsequently had a frameshift mutation and became a stop at position 10. Codons appear translational termination in the conserved regions of the protein. According to the standards and guidelines of the American College of Medical Genetics and Genomics, the mutation was judged to be a possible pathogenic variant. The mother of the proband (Ⅰ2) was a carrier of this site mutation. The clinical and genetic test results of the father and elder brother of the proband were not abnormal. ConclusionCACNA1F gene c.1761dupC is the pathogenic mutation site of the Schubert-Bornschein type incomplete CSNB family.
Inherited eye disease is a heterogeneous group of eye disorders caused by genetic defects, which has many genetic characteristics, such as multiple inheritance modes and numerous gene variation types. Over the past few decades, genetic testing has improved significantly, with more and more known disease-causing gene variants identified. With the rapid development of high-throughput sequencing technology, clinical diagnosis and treatment of eye genetic diseases have been accelerated, and molecular diagnosis of eye genetic diseases has become an important step in accurate diagnosis and treatment. How to correctly select and evaluate each kind of genetic testing technology, reasonably standardize the use of genetic testing technology, and provide patients with more accurate genetic counseling are problem that clinicians need to seriously consider.
ObjectiveTo evaluate macular microvessel changes in familial exudative vitreoretinopathy (FEVR) by optical coherence tomography angiography. MethodsCross-sectional clinical case-control study. From November 2019 to November 2020, 21 FEVR patients (41 eyes) from Weifang Eye Hospital were selected; 17 healthy volunteers (28 eyes) with the same age and gender as FEVR group were selected as normal control group. According to the best corrected visual acuity (BCVA) 1.0 and ＜1.0, FEVR group was divided into normal visual acuity group (27 eyes) and visual acuity decreased group (14 eyes). All enrollees received BCVA and OCTA. BCVA was performed with an international standard visual acuity chart, which was converted to logarithm of the minimum angle of resolution (logMAR) vision. The OCTA instrument was used to scan the macular area of all the examined eyes in the range of 3 mm×3 mm, 6 mm×6 mm, and the blood vessel density (VD) and blood perfusion density (PD) within the range of 3 mm×3 mm, 6 mm×6 mm were measured and the area, circumference, and morphological index of the foveal avascular zone (FAZ) within the range of 6 mm×6 mm. Quantitative data were compared between groups by independent sample t test. Statistical data were compared by χ2 test. The area under curve (AUC) of each index was determined according to receiver operating characteristic curve (ROC curve), and the predictive value of each index was evaluated. ResultsIn the macular area of 6 mm×6 mm, VD, PD, FAZ area and FAZ perimeter of FEVR group were all lower than those of normal control group, and the differences were statistically significant (t=−3.350, −2.387, −3.519, −3.029; P＜0.05). In macular area of 3 mm×3 mm and 6 mm×6 mm, compared with normal vision group and vision loss group, both VD and PD decreased. The differences were statistically significant (t=2.088, 2.114, 2.160, 2.545; P＜0.05). In the macular area of 6 mm×6 mm , the FAZ morphological index of the two groups was significantly different (t=2.409, P＜0.05). ROC curve analysis showed that all the indicators had low diagnostic value for FEVR (AUC＜0.5). ConclusionThere are microvascular abnormalities in macular area in FEVR patients, and the decrease of blood vessels and the change of FAZ shape may be related to the loss of visual acuity.
ObjectiveTo explore the light response, retinal inflammation and apoptosis of the retinal ganglion cells (RGCs) 1 year after the new type of channelrhodopsin PsCatCh2.0 was transfected into the retina of rd1 mice. MethodsTwenty-four male rd1 mice were randomly divided into rd1 experimental group and rd1 control group, 12 mice in each group. 1.5 μl of recombinant adeno-associated virus (rAAV)2/2-cytomegalovirus (CMV)-PsCatCh2.0-enhanced green fluorescent protein (EGFP) was injected into the vitreous cavity 1 mm below the corneoscleral limbus of mice in the rd1 experimental group, and the same dose of recombinant virus was injected 2 weeks later at temporal side 1 mm below the corneoscleral limbus. One year after virus injection, the light response of RGCs expressing PsCatCh2.0 was recorded by patch clamp technique; the expression of PsCatCh2.0 in the retina was evaluated by immunofluorescence staining; the transfection efficiency of recombinant virus was evaluated by the transfection efficiency of virus and the number of RGCs. Hematoxylin-eosin staining was performed to measure the inner retinal thickness. Western blotting was used to detect the protein expression of nuclear factor (NF)-κB p65 in retina; real-time quantitative polymerase chain reaction was used to detect the relative expression of tumor necrosis factor (TNF)-α, interleukin (IL)-6 and Bax mRNA. Terminal deoxynucleotidyl transferase kit was used to observe the apoptosis of retinal cells in each group of mice. ResultsOne year after the intravitreal injection of recombinant virus, PsCatCh2.0-expressing RGCs can still generate 30 pA photocurrent. The virus PsCatCh2.0-EGFP was mainly transfected into RGCs, and partly transfected into amacrine cells, almost no transfection was seen in bipolar and horizontal cells. There were no significant differences in the number of RGCs and thickness of the inner retina between the rd1 experimental group and the rd1 control group (F=14.35, 0.05; P＞0.05), while the rd1 experimental group NF-κB p65 protein expression, TNF-α and IL-6 mRNA quantification were significantly lower than those of rd1 control group (F=4.61, 5.91, 5.78; P＜0.05). The number of red fluorescent apoptotic cells in the retina of mice in the rd1 experimental group was less than that in the rd1 control group, and the Bax mRNA expression was lower than that in the rd1 control group, and the difference was statistically significant (F=7.52, P＜0.01). ConclusionOne year after intravitreal injection of recombinant virus, the PsCatCh2.0 expressing RGCs can still generate photocurrent. Long term transfection and expression of PsCatCh2.0 has no obvious cytotoxic effect on RGCs, nor it increases the inflammatory effect of the retina of rd1 mice with retinal degeneration.
The human hereditary retinal degeneration is one of the main cause of irreversible blindness in the world. the mechanisms leading to retinal photoreceptor degeneration are not entirely clear. However, microglia acting as innate immune monitors are found to be activated early in retinal degeneration in many retinitis pigmentosa animal models. These activated microglia are involved in phagocyte rod cell fragments of degenerated retina, and also produce high levels of cytotoxic substances such as pro-inflammatory cytokines and chemokines, which aggravate the death of adjacent healthy photoreceptor cells. It suggests that microglia activation plays an important role in photoreceptor degeneration. At the same time, a series of studies have confirmed that some drugs can prevent or reduce neuronal death and slow the occurrence and progression of retinal degeneration by interfering with abnormal activation of microglia. It is expected to be a new choice for the treatment of hereditary retinal degeneration.
Inherited retinal diseases (IRD) are a group of genetic disorders with high genetic and clinical heterogeneity. Patients with IRD may have their clinical diagnosis confirmed by genetic testing. Over the past 30 years, rapid advances in molecular genetics have raised the disease-causing gene variant detection rate and the accuracy of genetic testing, which provide hope to patients. The genetic diagnosis of patients with IRD is complicated due to the overlapping clinical phenotypes, and the fact that different variants lead to different phenotypes and severity even of the same gene. It is very important to overall evaluate the clinical phenotype of patients, precisely select genetic testing methods, and reasonably define disease-causing genes and variants during genetic diagnosis, which can guide the patient's subsequent treatment and provide genetic counseling.
Inherited retinal diseases (IRDs) are a group of severe retinal degenerative diseases leading to permanent visual impairment. IRDs are the major cause of irreversible blindness in children and working age groups. Gene therapy is a new clinical treatment method and currently the only clear and effective treatment for IRDs, while, there are still risks in clinical research and application. How to standardize perioperative management and reduce the potential risks of treatment is one of the keys to ensure the safety and effectiveness of treatment. However, there is no systematic and standardized guidance on the perioperative management for IRDs gene therapy. Therefore, in order to standardize the perioperative management, the Fundus Disease Group of Ophthalmology Society of Chinese Medical Association and Chinese Medical Doctor Association organized domestic experts to put forward standardized opinions on the perioperative management of IRDs gene therapy in China after repeated discussion and combined with domestic and foreign research experience, so as to provide clinicians with reference and application in clinical research and practice.
Rapid development of recently emerging precision medicine techniques represented by gene therapy has brought hope for the treatment of rare blinding eye diseases such as inherited retinal diseases (IRDs) for which there was no effective treatment previously. Although the globally growth of clinical trials for IRDs has increased rapidly over the past decade, due to the highly genetic and clinical phenotypic heterogeneity, as well as limited data on epidemiology and natural history of the disease, along with severe loss of vision function of majority patients for which the established measurements may not be appropriate, such studies lack standard outcome measurements and endpoints to asses clinical meaningful effectiveness, posing great challenges in terms of study design and evaluation of treatment efficacy, as well as clinical practice application. At present, there is no systematic nor standardized guidance on safety measures, clinical outcomes and endpoints of visual function for clinical trial design in IRDs. Therefore, in order to standardize the validated evaluation of IRDs clinical efficacy outcome measurements and endpoints, the Fundus Disease Group of Chinese Medical Association Ophthalmology Branch and Fundus Disease Committee of Chinese Medical Doctor Association Ophthalmology Branch organized domestic experts to put forward consensus and recommendations on standardizing outcome measurements and endpoints for clinical study design in IRDs, aiming to advance the study design of IRDs natural history research and clinical trials and to effectively evaluate disease progression and intervention efficacy. Along with the development of medical science and clinical trials, relevant content will be improved and updated accordingly.