Myocilin Mutations Are Not a Major Cause of Primary Congenital Glaucoma in Iranian Patients

Purpose: To assess the frequency of mutations in the Myocilin (MYOC) gene in Iranian patients affected with primary congenital glaucoma (PCG). Methods: The individuals evaluated herein are among a larger cohort of 100 patients who had previously been screened for CYP1B1 mutations. Eighty subjects carried mutations in CYP1B1, but the remaining 20 patients who did not, underwent screening for MYOC mutations for the purpose of the study. MYOC exons in the DNA were polymerase chain reaction (PCR) amplified and sequenced. Sequencing was performed using PCR primers, the ABI big dye chemistry and an ABI3730XL instrument. Sequences were analyzed by comparing them to reference MYOC sequences using the Sequencher software. Results: Four MYOC sequence variations were observed among the patients, but none of them were considered to be associated with disease status. Three of these variations were single nucleotide polymorphisms already reported not to be disease causing, the fourth variation created a synonymous codon and did not affect any amino acid change. Conclusion: In this cohort, MYOC mutations were not observed in any Iranian subject with PCG. It is possible that in a larger sample, a few subjects carrying disease causing MYOC mutations could have been observed. But our results show that the contribution of MYOC to PCG status in Iran is small if any

Variable Expressivity of Wolfram Syndrome in a Family with Multiple Affected Subjects

Purpose: To study the genetic basis and clinical manifestations of Wolfram syndrome in a multi-affected family. Methods: Complete clinical examinations including urological, ophthalmic, neurological, and endocrinologic assessment were performed for three affected family members. Genomic DNA was extracted from peripheral blood leukocytes with salting out method and all WFS1 exons and their flanking regions were sequenced. Candidate variation was screened for segregation in the pedigree by Sanger sequencing. Results: A known pathogenic missense mutation in WFS1 gene (c.1885C>T which leads to p.Arg629Trp in the encoded protein) was identified in all affected individuals. Both clinical and genetic investigations confirmed Wolfram syndrome diagnosis with variable phenotypic features. Conclusion: Identical mutations in the Wolfram syndrome causative gene can lead to variable manifestations of the syndrome even in the same family. Although the medical findings and clinical examination are imperative for the diagnosis of Wolfram syndrome, genetic testing is useful to confirm the diagnosis, especially in cases with possible reduced penetrance of the characteristic signs

Mutation Screening of Six Exons of ABCA4 in Iranian Stargardt Disease Patients

Purpose: Stargardt disease type 1 (STGD1) is a recessively inherited retinal disorder that can cause severe visual impairment. ABCA4 mutations are the usual cause of STGD1. ABCA4 codes a transporter protein exclusively expressed in retinal photoreceptor cells. The gene contains 50 exons. Mutations are most frequent in exons 3, 6, 12, and 13, and exons 10 and 42 each contain two common variations. We aimed to screen these exons for mutations in Iranian STGD1 patients. Methods: Eighteen STGD1 patients were recruited for genetic analysis. Diagnosis by retina specialists was based on standard criteria, including accumulation of lipofuscin. The six ABCA4 exons were PCR amplified and sequenced by the Sanger method. Results: One or more ABCA4-mutated alleles were identified in 5 of the 18 patients (27.8%). Five different mutations including two splice site (c.1356+1G>A and c.5836-2A>G) and three missense mutations (p.Gly1961Glu, p.Gly1961Arg, and p.Gly550Arg) were found. The p.Gly1961Glu mutation was the only mutation observed in two patients. Conclusion: As ABCA4 mutations in exons 6, 12, 10, and 42 were identified in approximately 25% of the patients studied, these may be appropriate exons for screening projects. As in other populations, STDG1 causative ABCA4 mutations are heterogeneous among Iranian patients, and p.Gly1961Glu may be relatively frequent

Choroidal Thickness in Different Types of Inherited Retinal Dystrophies

Purpose: To compare the choroidal thickness among eyes with retinitis pigmentosa (RP), Stargardt disease, Usher syndrome, cone-rod dystrophy, and healthy eyes of sex- and age-matched individuals. Methods: In this comparative study, 503 eyes with RP (n = 264), cone-rod dystrophy (n = 109), Stargardt disease (n = 76), and Usher syndrome (n = 54) were included. To validate the data, 109 healthy eyes of 56 sex- and age-matched individuals were studied as controls. Choroidal imaging was performed using enhanced depth imaging-optical coherence tomography. Choroidal thickness was measured manually using MATLAB software at 13 points in nasal and temporal directions from the foveal center with the interval of 500 μm and the choroidal area encompassing the measured points was calculated automatically. Results: The mean age was 36.33 ± 13.07 years (range, 5 to 72 years). The mean choroidal thickness at 13 points of the control eyes was statistically significantly higher than that in eyes with RP (P < 0.001) and Usher syndrome (P < 0.05), but not significantly different from that in eyes with Stargardt disease and cone-rod dystrophy. Among different inherited retinal dystrophies (IRDs), the choroidal thickness was the lowest in eyes with RP (P < 0.001). Choroidal thickness in the subfoveal area correlated negatively with best-corrected visual acuity (r = −0.264, P < 0.001) and the duration of ocular symptoms (r = −0.341, P < 0.001) in all studied IRDs. No significant correlation was observed between the subfoveal choroidal thickness and central macular thickness (r = −0.24, P = 0.576). Conclusion: Choroidal thinning in four different types of IRDs does not follow a similar pattern and depends on the type of IRD and the duration of ocular symptoms. A larger cohort is required to verify these findings

Erratum – Carrier Status for p.Gly61Glu and p.Arg368His CYP1B1 Mutations Causing Primary Congenital Glaucoma in Iran

This is an Erratum to "Carrier Status for p.Gly61Glu and p.Arg368His CYP1B1 Mutations Causing Primary Congenital Glaucoma in Iran" [J Ophthalmic Vis Res 2021;16(4):574–581] and does not have an abstract. Please download the PDF or view the article HTML

Carrier Status for p.Gly61Glu and p.Arg368His CYP1B1 Mutations Causing Primary Congenital Glaucoma in Iran

Purpose: To estimate carrier frequencies of CYP1B1 mutations p.Gly61Glu and p.Arg368His, respectively, in Talesh and the east of Guilan province in Iran with a maximum error of 2%. Previously, it was shown that these CYP1B1 mutations may be relatively prevalent in these regions. Methods: Population-based screenings were performed. DNA was extracted from saliva samples of 1036 individuals from Talesh and 3029 individuals from the east of Guilan. P.Gly61Glu and p.Arg368His screenings were performed, respectively, by RFLP and ARMS-based PCR protocols. For confirmation, the DNA of individuals with mutations was sequenced using the Sanger protocol. Results: Nine individuals from Talesh (0.86%; 95%CI: 0.45–1.64%) carried the p.Gly61Glu mutation, and 73 from the east of Guilan (2.41%; 95%CI: 1.91–3.04%) carried p.Arg368His. There was no significant difference in frequencies between urban and rural regions of the various cities, nor among four cities within the east of Guilan. Conclusion: The frequencies of p.Gly61Glu carriers in Talesh and of p.Arg368His carriers in the east of Guilan were within the 95% confidence interval of a previous study based on screenings of fewer individuals. The reliability of the recent estimates is higher, as the confidence interval for p.Gly61Glu decreased from 6.5% to 1.19% and the interval for p.Arg368His decreased from 4% to 1.13%. Based on the new findings, the maximum expected frequency of p.Gly61Glu carriers in Talesh is 1.64%, and of p.Arg368His carriers in the east of Guilan is 3%. The need for performing premarital screenings in the respective cities can be evaluated

The Inhibitory Effect of Connective Tissue Growth Factor Antibody on Postoperative Fibrosis in a Rabbit Model of Trabeculectomy

Purpose: To compare the efficacy of subconjunctival injection of an anti-connective tissue growth factor antibody (anti-CTGF) versus mitomycin-C (MMC) and placebo in reducing scar formation in a rabbit model of trabeculectomy. Methods: A total of 14 rabbits were included. Nine rabbits underwent trabeculectomy with subconjunctival injections of either anti-CTGF antibody, MMC, or balanced salt solution (BSS), each administered in three eyes, before peritomy. The anti-CTGF group received a repeated dose of the antibody five days after surgery. All nine rabbits were euthanized on day 14; the globes were stained with hematoxylin & eosin, Masson’s Trichrome, and immunohistochemistry for detecting alpha-smooth muscle (α-SMA) actin. RNA extraction was performed on five eyes of the remaining rabbits which included one eye without any surgery, one eye 5 hr after trabeculectomy without any injection, one eye five days after trabeculectomy without any injection, and two eyes five days after trabeculectomy with administration of MMC and BSS, respectively. Results: The mean bleb area in the anti-CTGF, MMC, and control groups was 3.8 ± 1.45, 5.9 ± 1.4, and 3.5 ± 1.9 mm2, respectively. Collagenous tissue was found to occupy the bleb area by 13.7%, 13.5%, and 18.5%, respectively. This ratio was significantly higher in the BSS group (P = 0.04). The expression of CTGF mRNA after 5 hr and five days in eyes undergoing trabeculectomy were significantly more pronounced as compared to the unoperated eye. The mean H-SCORE of α-SMA-immune reactive cells calculated as the grade of staining multiplied by the percentage of immune stained cells was 14.6, 10.22, and 140.58 in the anti-CTGF, MMC, and control groups, respectively. While the control eyes had a significantly higher score (Ps < 0.001), the anti-CTGF and MMC groups were comparable (P = 0.87). Conclusion: Based on the results of this animal study, the anti-CTGF antibody injection resulted in a significant reduction in collagenous tissue and myofibroblast cells after trabeculectomy

Screening of common CYP1B1 mutations in Iranian POAG patients using a microarray-based PrASE protocol

Purpose: The gene coding cytochrome P4501B1 (CYP1B1) has been shown to be a major cause of primary congenital glaucoma in the Iranian population. More recently it was shown to also be important in juvenile-onset open angle glaucoma (JOAG). We aimed to further investigate the role of CYP1B1 in a larger cohort of primary open angle glaucoma (POAG) patients which included late-onset patients. We also aimed to set up a microarray based protocol for mutation screening with an intent of using the protocol in a future population level screening program. Methods: Sixty three POAG patients, nine affected family members, and thirty three previously genotyped primary congenital glaucoma (PCG) patients were included in the study. Clinical examination included slit lamp biomicroscopy, IOP measurement, gonioscopic evaluation, fundus examination, and measurement of perimetry. G61E, R368H, R390H, and R469W were screened by a protocol that included multiplexed allele specific amplification in the presence of a protease (PrASE), use of sequence tagged primers, and hybridization to generic arrays on microarray slides. The entire coding sequences of CYP1B1 and myocilin (MYOC) genes were sequenced in all individuals assessed by the microarray assay to carry a mutation. Intragenic single nucleotide polymorphism (SNP) haplotpes were determined for mutated alleles. Results: Genotypes assessed by the array-based PrASE methodology were in 100 concordance with sequencing results. Seven mutation carrying POAG patients (11.1) were identified, and their distribution was quite skewed between the juvenile-onset individuals (5/21) as compared to late-onset cases (2/42). Four of the seven mutation carrying Iranian patients harbored two mutated alleles. CYP1B1 mutated alleles in Iranian PCG and POAG patients shared common haplotypes. MYOC mutations were not observed in any of the patients. Conclusions: The PrASE approach allowed reliable simultaneous genotyping of many individuals. It can be an appropriate tool for screening common mutations in large sample sizes. The results suggest that CYP1B1 is implicated in POAG among Iranians, notably in the juvenile-onset form. Contrary to POAG patients studied in other populations, many mutation harboring Iranian patients carry two mutated alleles. We propose an explanation for this observation. © 2008 Molecular Vision

A large, curated, open-source stroke neuroimaging dataset to improve lesion segmentation algorithms.

Accurate lesion segmentation is critical in stroke rehabilitation research for the quantification of lesion burden and accurate image processing. Current automated lesion segmentation methods for T1-weighted (T1w) MRIs, commonly used in stroke research, lack accuracy and reliability. Manual segmentation remains the gold standard, but it is time-consuming, subjective, and requires neuroanatomical expertise. We previously released an open-source dataset of stroke T1w MRIs and manually-segmented lesion masks (ATLAS v1.2, N = 304) to encourage the development of better algorithms. However, many methods developed with ATLAS v1.2 report low accuracy, are not publicly accessible or are improperly validated, limiting their utility to the field. Here we present ATLAS v2.0 (N = 1271), a larger dataset of T1w MRIs and manually segmented lesion masks that includes training (n = 655), test (hidden masks, n = 300), and generalizability (hidden MRIs and masks, n = 316) datasets. Algorithm development using this larger sample should lead to more robust solutions; the hidden datasets allow for unbiased performance evaluation via segmentation challenges. We anticipate that ATLAS v2.0 will lead to improved algorithms, facilitating large-scale stroke research