To develop and characterize a novel mouse model of granular corneal dystrophy type II (GCD2) using CRISPR/Cas9 technology and explore the underlying pathogenesis of transforming growth factor-beta-induced protein (TGFBIp) aggregation. CRISPR/Cas9 technology was employed to introduce the R124H mutation in the TGFBI gene of mice. Genomic sequencing and polymerase chain reaction confirmed the mutation. Phenotypic characteristics were evaluated through slit-lamp examination, optical coherence tomography, histological analysis, electron microscopy, and immunofluorescence, comparing wild-type (WT), heterozygous (HE), and homozygous (HO) mice. Transcriptome sequencing was conducted to identify the pathogenesis of GCD2. The findings were further validated through western blotting and transmission electron microscopy. The R124H mutation in TGFBI was successfully introduced, with breadcrumb-like deposits observed in the corneas of mutant mice, with HO mice displaying more severe phenotypes than HE mice. TGFBIp levels were elevated in HE and HO mice (both P < 0.001). Histological and electron microscopy analyses revealed abnormal collagen arrangement and TGFBIp deposits in the corneal stroma of the HE and HO mice. Transcriptome analysis indicated that the TGFBI-R124H mutation was associated with impaired autophagy, endocytosis, and extracellular matrix signaling. Additional experiments confirmed autophagy-related markers LC3 and SQSTM1 were upregulated in the corneas of mutant mice, accompanied by increased autophagosome formation in corneal keratocytes, indicating impaired autophagy flux in HE and HO mice. We established a GCD2 mouse model caused by the R124H mutation using CRISPR/Cas9, providing a reliable platform for understanding pathogenesis for GCD2. Lattice corneal dystrophy (LCD) is an inherited disorder of the eye characterized by the deposition of amyloid resulting in steadily progressive loss of vision. These deposits create linear, “lattice-like” opacities arising primarily in the central cornea, while the peripheral cornea is often spared. They are radially oriented and are accompanied by gradual, superficial opacification of the cornea. Recurrent epithelial erosions are often present, causing ocular irritation and additional vision loss. The erosions may appear before any noticeable stromal deposits. LCD belongs to a broader family of corneal dystrophies and has several subtypes, as described below. Type I LCD (LCD1), also known as classic lattice corneal dystrophy or Biber-Haab-Dimmer dystrophy, is the primary form of LCD. It is autosomal dominant and results from mutations in the transforming human growth factor beta-induced (TGFBI) gene. Although TGFBI and its protein transcript are found throughout the body, there are no known systemic effects outside of the ocular pathology for which it is named. It usually presents in the first or second decade of life. The LCD variants are subtypes of LCD caused by a variety of mutations on the TGFBI gene. The variants were formerly described as types IA, III, IIIA, IIIB, IV, V, VI, VII, and polymorphic corneal amyloidosis. These are variations of the same disease process that causes type I LCD, with minor changes in their phenotypic features. Specific phenotypic patterns are traceable to specific mutations in the TGFBI gene, which resulted in their being initially described as separate diseases. They are now considered to be subtypes of the same disease, in which type I is the classic and most common presentation. The variants are often geographically specific and are occasionally traceable to single founder mutations. LCD type II is no longer included among the corneal dystrophies as it is a primarily systemic disorder with ophthalmologic features. While it was initially included in the LCD family because of the lattice-like ocular deposits, it is now more accurately described as familial amyloid polyneuropathy (FAP) type IV, FAP Finnish type, FAP Gelsolin type, or Meretoja syndrome. Systemic symptoms include neuropathy (due to amyloid infiltration of nerves), facial paralysis, and extreme skin laxity. It tends to present in the twenties. Another disease process often mistakenly included in the LCD family is granular corneal dystrophy type II (GCD type II). Also called combined granular-lattice dystrophy or Avellino dystrophy, GCD type II was formerly considered a hybrid of granular and lattice dystrophies since it exhibits symptoms of both diseases. It is characterized by both granular and branching linear deposits that make it challenging to distinguish it from the LCDs. Physical exam findings that differentiate it from the lattice dystrophies are further discussed in this article under the name Avellino dystrophy, to minimize confusion with type II LCD.

To develop an automatic algorithm to analyze dystrophic lesions on photographic images of corneal dystrophy. The dataset included 32 images of corneal dystrophy. The dystrophic area was manually segmented twice. Manually labeled dystrophy areas were compared with automatically segmented images. First, we manually removed the light reflex from the image of the cornea. Using an automatic approach, we extracted the brown color of the iris. Then, the program detected the circular region of the pupil and the corneal surface. A whitish dystrophy area was defined based on the image intensity on the iris and the pupil. The sliding square kernel was applied to clearly define the dystrophic region. For the manual analysis and the twice automatic approach, the Dice similarity was 0.804 and 0.801, respectively. The Pearson correlation coefficient was 0.807 and 0.806, respectively. The total number of distinct dystrophic areas showed no significant difference between the manual and automatic approaches according to the Wilcoxon signed-rank test (p < 0.0001, both). We proposed an automatic algorithm for detecting the dystrophy areas on photographic images with an accuracy of approximately 0.80. This system can be applied to detect and predict the progression of corneal dystrophy.

Granular and lattice corneal dystrophies (GCDs & LCDs) are autosomal dominant inherited disorders of the cornea. Due to genetic heterogeneity and large genes, unraveling the mutation is challenging. Patients underwent comprehensive clinical examination, and targeted next-generation sequencing (NGS) was used for mutation detection. Co-segregation and in silico analysis was accomplished. Patients suffered from GCD. NGS disclosed a known pathogenic variant, c.371G>A (p.R124H), in exon 4 of TGFBI. The variant co-segregated with the phenotype in the family. Homozygous patients manifested with more severe phenotypes. Variable expressivity was observed among heterozygous patients. The results, in accordance with previous studies, indicate that the c.371G>A in TGFBI is associated with GCD. Some phenotypic variations are related to factors such as modifier genes, reduced penetrance and environmental effects.

Many types of inherited renal diseases have ocular features that occasionally support a diagnosis. The following study describes an unusual example of a 40-year-old woman with granular corneal dystrophy type II complicated by renal involvement. These two conditions may coincidentally coexist; however, there are some reports that demonstrate an association between renal involvement and granular corneal dystrophy type II. Granular corneal dystrophy type II is caused by a mutation in the transforming growth factor-β-induced (TGFBI) gene. The patient was referred to us because of the presence of mild proteinuria without hematuria that was subsequently suggested to be granular corneal dystrophy type II. A kidney biopsy revealed various glomerular and tubular basement membrane changes and widening of the subendothelial space of the glomerular basement membrane by electron microscopy. However, next-generation sequencing revealed that she had no mutation in a gene that is known to be associated with monogenic kidney diseases. Conversely, real-time polymerase chain reaction, using a simple buccal swab, revealed TGFBI heteromutation (R124H). The TGFBI protein plays an important role in cell-collagen signaling interactions, including extracellular matrix proteins which compose the renal basement membrane. This mutation can present not only as corneal dystrophy but also as renal disease. TGFBI-related oculorenal syndrome may have been unrecognized. It is difficult to diagnose this condition without renal electron microscopic studies. To the best of our knowledge, this is the first detailed report of nephropathy associated with a TGFBI mutation.

Clinical, instrumental, and genetic findings are reported in Italian families with Type II Granular Corneal Dystrophies (GCD2) presenting an initial unusual presentation of a Granular Corneal Dystrophy Type I (GCD1) phenotypic spectrum in female descendants. Slit-lamp examinations showed the typical phenotypic features of GCD2 in both mothers and a phenotypic appearance of GCD1 in both daughters. Despite the different phenotypic onset, the genetic diagnostic testing revealed the presence of a mutation in the TGFB-I gene, typical of GCD2 in both cases, excluding GCD1. Patients who were clinically suspected of corneal dystrophy need a genetic confirmatory testing for certain diagnosis. Genetic test may help to find the specific mutation distinguishing between different phenotypic spectra with relative diagnostic and prognostic implications. The study demonstrates that the phenotypic spectrum of genetically confirmed granular corneal dystrophies in patients may change over time. Since the R124H mutation has also been described in clinically asymptomatic individuals prior to LASIK, who then develop dramatic deposition, suggesting that this particular mutation and phenotype may be sensitive to, precipitated, or modified by central cornea trauma, a careful familial anamnesis excluding cornel dystrophies and specific preoperative genetic test are recommended prior to LASIK.