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Yutong Jing, Kai Liao, Zekai Cui, Yalan Zhou, shenyang li, Shibo Tang, Jiansu Chen; Modeling macular corneal dystrophy: iPSC-derived corneal stromal-like cells with heterozygous mutations in CHST6 gene. Invest. Ophthalmol. Vis. Sci. 2020;61(7):2913.
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© ARVO (1962-2015); The Authors (2016-present)
In order to advanced understanding the pathogenesis of macular corneal dystrophy (MCD), we created a cell model in vitro by using induced pluripotent stem cells (iPSCs), which carrying compound heterozygous mutations in the CHST6 gene.
In this sturdy, we recruited an MCD family with three siblings. The phenotype of each patient was demonstrated by detailed clinical examination. Genetic analysis by targeting next-generation sequencing (NGS), which verified compound heterozygous variation in the CHST6 gene (NM_021615): c.62_63delTinsGA (p. L21delinsRfs) and c.C892T (p. Q298X). Generation of MCD-iPSCs from specific urine cells and passed through neural crest stem cells (NCSCs) contribute to the corneal stromal-like cells (CSCs). CSCs characteristic was confirmed by immunofluorescence staining and real-time polymerase chain reaction(qPCR).
The pedigree of the MCD-family. Genotype information of compound heterozygous mutations performed sanger sequencing. Pre-operative photograph of the eyes of II.5 demonstrating ill-defined corneal stromal despaired. Confocal microscopy images (Fig. 1,2 A-B). The morphology of MCD-iPSC, which be able to differentiated into all three layers (Fig. 3A). We adopted adherent culture(2D) and three-dimensional (3D) spheroid culture of NCSC differentiation medium, qPCR displayed 3D state promoted the expression of the characteristic markers of NCSCs (Fig. 3B). The characterization of iPSC-Derived CSC (Fig. 3C).
NCSCs undergo general and coordinated movements away from folds of the neural ectoderm to different regions of the embryo to give rise to ocular cells. We use patient-derived iPS cells via regulating NCSCs induction to simulating a MCD disease model in vitro. This information serves as platform for further analyses of the molecular networks involved in MCD, and provide insights into cell-based therapies, drug discovery.
This is a 2020 ARVO Annual Meeting abstract.
Fig. 1. The pedigree of the family (A). Square indicates male and circle indicates female. Dark symbol means affected member, all examined individuals were marked in the side. The arrow above the patient represents the proband. Ito IV means generations and 1 to 13 means the ID number of each member. The genotype information of compound heterozygous mutations (B) .Fig. 2. Clinical phenotypes.
Fig. 3. Characterization of differentiation cell
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