An Arg124Cys mutation in transforming growth factor ?-induced gene associated with lattice corneal dystrophy type I in a Chinese pedigree

Purpose: To identify a clinical and genetic form of a large Chinese family with an autosomal?dominant lattice corneal dystrophy type I (LCD I). Methods: The patients’ eyes were examined on the basis of slit?lamp microscopy, and other clinical records were also collected. Genomic DNA was extracted from peripheral leukocytes of the affected patients and their unaffected family members. Each previous reported mutation of the transforming growth factor ??induced gene (TGFBI) gene was amplified by touch?down polymerase chain reaction and directly sequenced to verify the disease?causing mutation. Results: Typical clinical features of LCD I were found by slit?lamp photography in these affected Chinese pedigrees. A heterozygous single base?pair transition from C to T (c.418 C > T), leading to amino acid substitution Arg124Cys (R124C) in the encoded TGFBI protein, was detected in all of the eighteen affected patients. The same mutation was not found in unaffected family members. Conclusion: The R124C mutation hot spot, which was relatively rare in China, was responsible for LCD I in the large family. Molecular genetic analysis of TGFBI gene can offer an accurate diagnosis of patients with lattice corneal dystrophies in the clinical treatment.

Lithium chloride promotes the proliferation and autophagy of corneal stromal fibroblasts by TGFBI / 国际眼科杂志(Guoji Yanke Zazhi)

@#AIM: To study the expressions of TGFBI and microtubule-associated protein 1 light chain 3 alpha(LC3)in granular corneal dystrophy, and the influences of lithium chloride(LiCl)on corneal stromal fibroblast cell proliferation by TGFBI. <p>METHODS: Immunohistochemistry and Western-blot assays were used to detect the expression levels of TGFBI and LC3 in corneal dystrophy and normal corneal tissues. TGFBI overexpression vector was transfected into corneal stromal fibroblasts, and then the cells were treated with 5, 10, 20, 40mmol/L LiCl for different times(0, 1, 6, 12h), and Western-blot assay was performed to evaluate the expression levels of TGFBI and LC3, and CCK-8 assay was carried out to assess cell proliferation activity.<p>RESULTS: TGFBI and LC3 were highly expressed in corneal tissues of patients with corneal dystrophy. TGFBI overexpression inhibited the proliferation ability of corneal stromal fibroblasts(<i>P</i><0.05). LiCl inhibited the expression levels of TGFBI and LC3, and enhanced the cell proliferation activity in corneal stromal fibroblasts transfected with mutant TGFBI(<i>P</i><0.05).<p>CONCLUSION: LiCl promoted the proliferation and autophagy of corneal stromal fibroblasts, and its mechanism may be related to down regulated expressions of TGFBI and LC3.

Clinical observation and gene mutation detection in a pedigree of retinal lattice degeneration combined with granular corneal dystrophy / 中华眼底病杂志

Objective To investigate the clinical manifestations and gene mutation of a pedigree with retinal lattice degeneration and granular corneal dystrophy (GCD) type 2.Methods Ten members in 3 generations of a pedigree with retinal lattice degeneration and GCD2 were included in the study, including 6 patients (3 males and 3 females) and 4 healthy family members. All members underwent visual acuity, slit lamp microscope, three-mirror lens, fundus color photography, optical coherence tomography, and corneal endothelial cells counting. Genomic DNA was extracted from peripheral venous blood (2 ml) from all the subjects and their spouses, who had no related inherited diseases. The next generation sequencing method was used to detect the mutation sites of transforming growth factor β (TGFBI), and all results underwent Sanger verification.Results Among the 12 eyes of 6 patients, the visual acuity was FC/20 cm-1.0. In the superficial central corneal stroma, snowflake-like deposits were observed in three cases (6 eyes), and a small amount of granular deposits were observed in three cases (6 eyes). Corneal endothelial cell counts were normal. Retinal lattice degeneration were observed in 3 cases, 6 eyes (including 3 cases of rhegmatogenous retinal detachment in 4 eyes); retinal thinning without obvious lattice degeneration in 4 eyes of 2 patients. Nystagmus in 1 patient and fundus examination showed no significant abnormalities. DNA sequencing results showed that the proband and 4 patients had missense mutation of TGFBI gene in exon 4 c.371G> A, the mutation site corresponding to the amino acid change encoded by TGFBI gene No. 124 Amino acids, from arginine to histidine (p.R124H). Patients with this mutation have varying degrees of clinical phenotype.Conclusions The mutation of c.701G> A (p.R124H) in TGFBI gene is the causative gene of GCD in this pedigree. The patients with this mutation have different clinical phenotypes.

Research Progress of Granular Corneal Dystrophy / 现代生物医学进展

Granular corneal dystrophy is a rare autosomal dominant genetic disease in clinic.Due to the TGFBI mutation on the 5q31 chromosome,the TGFBIp abnormally aggregates in the Bowman layer and the matrix layer and metabolic disorders,patients' bilateral cornea arise opacity,making visual acuity Progressive impairment.At present,there are at least 66 TGFBI mutations,at least 10 of which are related to granular corneal dystrophy,due to variation in genotype and the difference between homozygous and heterozygous,the patients' phenotype shows a significant difference.Along with the improvement of people's cognition,and the application of laser scanning confocal microscope and the gene diagnosis,More and more patients get the correct diagnosis,Current treatment methods mainly include corneal transplantation and laser ablation,patients are not satisfied because of the postoperative recurrence and aggravated.Due to the establishment of granular corneal dystrophy animal model,lithium and gene therapy will show a good application prospects.

Mutation Analysis of the TGFBI Gene in Consecutive Korean Patients With Corneal Dystrophies

BACKGROUND: Mutations in the transforming growth factor beta-induced gene (TGFBI) are major causes of genetic corneal dystrophies (CDs), which can be grouped into TGFBI CDs. Although a few studies have reported the clinical and genetic features of Korean patients with TGFBI CD, no data are available regarding the frequency and spectrum of TGFBI mutations in a consecutive series of Korean patients with clinically diagnosed CDs. METHODS: Patients with any type of CD, who underwent both ophthalmologic examination and TGFBI gene analysis by Sanger sequencing at a tertiary care hospital in Seoul, Korea from 2006 to 2013, were enrolled in this study. RESULTS: Among a total of 89 patients, 77 (86.5%) were diagnosed as having clinical TGFBI CD. Seventy-three out of 74 patients (98.6%) with granular CD type 2 (GCD2), had the p.R124H mutation. Of particular note, one patient with rapidly progressive CD had the p.R124H mutation as well as a novel nonsense variant with unknown clinical significance (p.A179*). In three patients with lattice CD type 1 (LCD1), one known mutation (p.R124C) and two novel variants (p.L569Q and p.T621P) in the TGFBI gene were identified. CONCLUSIONS: This study provides epidemiological insight into CDs in a Korean population and reaffirms that GCD2 is the most common TGFBI CD phenotype and that p.R124H is the only mutation identified in patients with GCD2. In addition, we broaden the spectrum of TGFBI mutations by identifying two novel missense variants in patients with LCD1.

A Comparative Study of the Diagnostic Performance of the GENEDIA Avellino Corneal Dystrophy Mutation Detection Kit and Screening Master Mix and a Direct Sequencing Method to Detect Mutations in the TGFB1 Gene / 임상검사와정도관리

BACKGROUND: Mutations in the gene encoding transforming growth factor-beta induced (TGFBI) are associated with corneal dystrophies. We evaluated the diagnostic performance of the GENEDIA Avellino corneal dystrophy (ACD) mutation detection kit and GENEDIA corneal dystrophy screening master mix (Green Cross Medical Science Co., Korea) by comparing it with an in-house sequencing method. METHODS: The study group consisted of 40 patients with Avellino corneal dystrophy (ACD) and 40 patients suspected to suffer from ACD; 40 healthy individuals were used as the control. All samples used for this study were previously obtained. All results obtained using the kit were evaluated for sensitivity, specificity, and detection limit. RESULTS: The sensitivity of the GENEDIA ACD kit was 100.0% with a positive mean+/-2SD Ct (cycle threshold) value of 25.87+/-1.24 and an excellent coefficient of variation value of 0.02 in ACD group. All normal control samples were negative, indicating a specificity of 100% for the GENEDIA kit. The detection limit was set at a DNA concentration of >0.2 ng/microL. Direct sequencing results obtained using the GENEDIA master mix and the in-house method agreed for all 20 ACD samples. Additional R555W mutation detected in four ACD-suspected samples were suggestive of the diagnosis of granular corneal dystrophy type I. CONCLUSIONS: The GENEDIA ACD detection kit and master mix showed acceptable results, demonstrating high sensitivity and specificity, and may be considered for clinical application. Furthermore, the GENEDIA master mix was useful for the detection of mutations in exons 4 and 12 of the TGFBI gene.

Long-Term Outcomes of Penetrating Keratoplasty in Treating Macular Corneal Dystrophy, TGFBI Dystrophy, and Fuchs' Dystrophy

PURPOSE: To compare long-term outcomes of penetrating keratoplasty in treating macular corneal dystrophy, transforming growth factor beta induced (TGFBI) dystrophy, and Fuchs' dystrophy. METHODS: The present retrospective study describes the postoperative results of 123 eyes in 70 patients; 53 eyes with macular corneal dystrophy, 20 eyes with TGFBI dystrophy, and 50 eyes with Fuchs' dystrophy. All patients received penetrating keratoplasty. The outcome measures included recurrence rate, graft rejection, graft survival rate and corneal endothelial density and other complications. RESULTS: As compared with the other groups, TGFBI dystrophy had the highest recurrence rate (p-value < 0.0001). Fuchs' dystrophy had the lowest graft survival rate (p-value = 0.03). Corneal endothelial cell density was lowest in Fuchs' dystrophy 10 years postoperatively (p-value = 0.0006), but there was no significant difference in corneal endothelial cell density between the 3 groups 1 and 5 years postoperatively. There was no significant difference between the 3 groups in other complications such as graft rejection, secondary glaucoma and persistent epithelial defect (p-value = 0.809, p-value = 0.584, p-value = 0.972, respectively). CONCLUSIONS: Penetrating keratoplasty showed different long-term outcomes in treating 3 types of corneal dystrophies. The difficulty may depend on the pathogenesis of corneal dystrophies.

Clinical Manifestations of Avellino Corneal Dystrophy Diagnosed by Non-invasive Genetic Test

PURPOSE: To introduce a new genetic method for the diagnosis of Avellino corneal dystrophy (ACD), which is non-invasive and can be easily performed on an outpatient basis, and to evaluate the relationship between the degree of corneal opacity and age or sex. METHODS: A genetic study was performed on 11 patients who had a specific corneal opacity by slit-lamp examination and on four normal patients by using a specific adhesive tape to obtain epidermal keratinocytes. Corneal dystrophy was diagnosed according to the genetic study. RESULTS: All 11 patients were confirmed as having heterozygous ACD. Heterozygous ACD patients were classified into five stages: trace, mild, moderate, severe, or very severe, based on slit-lamp photography status. Corneal stages had no relationship with sex (p=0.982), but the severity of ACD increased with age (p=0.005). CONCLUSIONS: A non-invasive sticker-type genetic study kit, the "U-gene test" is a good method to diagnose corneal dystrophy genetically. Avellino corneal dystrophy becomes more severe over time but has no relationship with sex.