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Céline Portal, Qing Liu, Peter Y Lwigale, Carlo Iomini; Primary cilia deficiency in neural crest cells leads to Anterior Segment Dysgenesis (ASD) by disruption of the Indian Hedgehog signaling pathway in the periocular mesenchyme. Invest. Ophthalmol. Vis. Sci. 2019;60(9):4154.
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Anterior segment dysgenesis (ASD) represents one of the main causes of congenital opacity and leads to glaucoma in ~50% of the cases. Several ciliopathies display conditions affecting tissues of the anterior segment (AS) of unknown pathogenesis. Here we test the hypothesis that assembly of primary cilia in neural crest cells (NCC), from which most of the AS tissue derives, is required for normal AS development.
We utilize mouse genetics tools to inactivate genes or to detect hedgehog activity in the NCC. Phenotypic analysis of AS in ciliary mutants was assessed on eye sections of paraffin or plastic embedded tissue. Cell proliferation in the periocular mesenchyme (POM) was assessed by bromodeoxyuridine labeling. Components of the Hedgehog (Hh) pathway such as cilia and Smoothened (Smo) as well as nerves and blood vessels were detected by immunostaining. Gene expression was assessed by RT-qPCR on dissected eye tissues or FACS sorted cells using genetically activable fluorescent markers.
Because somatic mutations of ciliogenic genes, including IFT88, lead to midgestation death, we obtained Wnt1-Cre;Ift88fx/fx conditional mutants by ablating IFT88 in NCC. The disruption of primary cilia in the NCC leads to significant reduction of the anterior chamber and cornea diameter, thinner stroma thickness with abnormal keratocytes, reduced mesenchymal cell accumulation at the iridocorneal angle, disorganized corneal innervation and corneal neovascularization. Gene expression analysis revealed reduction of Hh target genes, as well as genes encoding for transcription factors linked to the Axenfeld-Rieger Syndrome, like Pitx2 and Foxc1. By using a reporter mouse (Gli1-LacZ) we show a dramatic reduction of the Hh activity and cell proliferation in POM cells surrounding the optic nerve and the retinal pigmented epithelium including the iridocorneal angle area in cilia mutants. Importantly, ablation of Smo in the NCC leads to a similar but more severe AS phenotype compared to the one observed in cilia mutants. Finally, we show that the main activator of the Hh pathway in the POM is Indian Hh.
Primary cilia play a pivotal role in AS development by promoting the Hh signaling in a subset of POM cells and are involved in the etiology of ocular disorders derived from ASD.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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