TY - JOUR T1 - Cilia-associated wound repair mediated by IFT88 in retinal pigment epithelium. JF - Sci Rep Y1 - 2023 A1 - Ning, Ke A1 - Bhuckory, Mohajeet B A1 - Lo, Chien-Hui A1 - Sendayen, Brent E A1 - Kowal, Tia J A1 - Chen, Ming A1 - Bansal, Ruchi A1 - Chang, Kun-Che A1 - Vollrath, Douglas A1 - Berbari, Nicolas F A1 - Mahajan, Vinit B A1 - Hu, Yang A1 - Sun, Yang KW - Animals KW - Cilia KW - Ciliopathies KW - Disease Models, Animal KW - Humans KW - Mice KW - Microtubule-Associated Proteins KW - Retinal Degeneration KW - Retinal Pigment Epithelium KW - Tumor Suppressor Proteins AB -

Primary cilia are conserved organelles that integrate extracellular cues into intracellular signals and are critical for diverse processes, including cellular development and repair responses. Deficits in ciliary function cause multisystemic human diseases known as ciliopathies. In the eye, atrophy of the retinal pigment epithelium (RPE) is a common feature of many ciliopathies. However, the roles of RPE cilia in vivo remain poorly understood. In this study, we first found that mouse RPE cells only transiently form primary cilia. We then examined the RPE in the mouse model of Bardet-Biedl Syndrome 4 (BBS4), a ciliopathy associated with retinal degeneration in humans, and found that ciliation in BBS4 mutant RPE cells is disrupted early during development. Next, using a laser-induced injury model in vivo, we found that primary cilia in RPE reassemble in response to laser injury during RPE wound healing and then rapidly disassemble after the repair is completed. Finally, we demonstrated that RPE-specific depletion of primary cilia in a conditional mouse model of cilia loss promoted wound healing and enhanced cell proliferation. In summary, our data suggest that RPE cilia contribute to both retinal development and repair and provide insights into potential therapeutic targets for more common RPE degenerative diseases.

VL - 13 IS - 1 ER - TY - JOUR T1 - Chorioretinal atrophy following voretigene neparvovec despite the presence of fundus autofluorescence. JF - Mol Genet Genomic Med Y1 - 2022 A1 - Kolesnikova, Masha A1 - Lima de Carvalho, Jose Ronaldo A1 - Parmann, Rait A1 - Kim, Angela H A1 - Mahajan, Vinit B A1 - Tsang, Stephen H A1 - Sparrow, Janet R KW - Atrophy KW - Child KW - cis-trans-Isomerases KW - Female KW - Humans KW - Leber Congenital Amaurosis KW - Mutation KW - Retinal Degeneration AB -

INTRODUCTION: Leber congenital amaurosis (LCA) type 2, due to disease-causing variants in RPE65, is characterized by severe visual loss in early infancy. Current treatments include voretigene neparvovec-rzyl (VN) for RPE65-associated LCA. Herein, we present the long-term follow-up of a patient treated with VN using quantitative autofluorescence (488 nm excitation).

CASE REPORT: A 9-year-old girl with a diagnosis of LCA with biallelic variants in RPE65 presented for evaluation. The patient underwent VN treatment at the age of 11. The patient returned to clinic at age of 19 at which time imaging revealed evidence of chorioretinal atrophy. Quantitative autofluorescence performed prior to gene therapy and at 6- and 8-year follow-up revealed a central area of fundus autofluorescence.

DISCUSSION: This case report demonstrates acquisition of fundus autofluorescence at 6- and 8-year follow-up despite the development of chorioretinal atrophy.

VL - 10 IS - 11 ER - TY - JOUR T1 - CRISPR Repair Reveals Causative Mutation in a Preclinical Model of Retinitis Pigmentosa. JF - Mol Ther Y1 - 2016 A1 - Wu, Wen-Hsuan A1 - Tsai, Yi-Ting A1 - Justus, Sally A1 - Lee, Ting-Ting A1 - Zhang, Lijuan A1 - Lin, Chyuan-Sheng A1 - Bassuk, Alexander G A1 - Mahajan, Vinit B A1 - Tsang, Stephen H KW - Animals KW - Clustered Regularly Interspaced Short Palindromic Repeats KW - CRISPR-Cas Systems KW - Cyclic Nucleotide Phosphodiesterases, Type 6 KW - Disease Models, Animal KW - DNA Repair KW - Electroretinography KW - Exons KW - Gene Editing KW - Genetic Loci KW - Homologous Recombination KW - Mice KW - Mice, Transgenic KW - Mutation KW - Photoreceptor Cells, Vertebrate KW - Retinal Degeneration KW - Retinitis Pigmentosa KW - RNA, Guide AB -

Massive parallel sequencing enables identification of numerous genetic variants in mutant organisms, but determining pathogenicity of any one mutation can be daunting. The most commonly studied preclinical model of retinitis pigmentosa called the "rodless" (rd1) mouse is homozygous for two mutations: a nonsense point mutation (Y347X) and an intronic insertion of a leukemia virus (Xmv-28). Distinguishing which mutation causes retinal degeneration is still under debate nearly a century after the discovery of this model organism. Here, we performed gene editing using the CRISPR/Cas9 system and demonstrated that the Y347X mutation is the causative variant of disease. Genome editing in the first generation produced animals that were mosaic for the corrected allele but still showed neurofunction preservation despite low repair frequencies. Furthermore, second-generation CRISPR-repaired mice showed an even more robust rescue and amelioration of the disease. This predicts excellent outcomes for gene editing in diseased human tissue, as Pde6b, the mutated gene in rd1 mice, has an orthologous intron-exon relationship comparable with the human PDE6B gene. Not only do these findings resolve the debate surrounding the source of neurodegeneration in the rd1 model, but they also provide the first example of homology-directed recombination-mediated gene correction in the visual system.

VL - 24 IS - 8 U1 - http://www.ncbi.nlm.nih.gov/pubmed/27203441?dopt=Abstract ER - TY - JOUR T1 - Quantitative autofluorescence as a clinical tool for expedited differential diagnosis of retinal degeneration. JF - JAMA Ophthalmol Y1 - 2015 A1 - Marsiglia, Marcela A1 - Lee, Winston A1 - Mahajan, Vinit B A1 - Zernant, Jana A1 - Delori, François C A1 - Tsang, Stephen H A1 - Sparrow, Janet R KW - Adult KW - ATP-Binding Cassette Transporters KW - Diagnosis, Differential KW - DNA KW - DNA Mutational Analysis KW - Fluorescein Angiography KW - Fundus Oculi KW - Humans KW - Male KW - Mutation KW - Optical Imaging KW - Retina KW - Retinal Degeneration KW - Retinitis Pigmentosa VL - 133 IS - 2 U1 - http://www.ncbi.nlm.nih.gov/pubmed/25375877?dopt=Abstract ER - TY - JOUR T1 - Calpain-5 mutations cause autoimmune uveitis, retinal neovascularization, and photoreceptor degeneration. JF - PLoS Genet Y1 - 2012 A1 - Mahajan, Vinit B A1 - Skeie, Jessica M A1 - Bassuk, Alexander G A1 - Fingert, John H A1 - Braun, Terry A A1 - Daggett, Heather T A1 - Folk, James C A1 - Sheffield, Val C A1 - Stone, Edwin M KW - Amino Acid Sequence KW - Base Sequence KW - Calpain KW - Cell Line KW - Cells, Cultured KW - Choroid Diseases KW - Exome KW - Exons KW - Eye Diseases, Hereditary KW - Female KW - Gene Expression KW - Genetic Linkage KW - Humans KW - Male KW - Models, Molecular KW - Molecular Sequence Data KW - Mutation KW - Pedigree KW - Phenotype KW - Photoreceptor Cells, Vertebrate KW - Protein Conformation KW - Protein Transport KW - Retinal Degeneration KW - Sequence Alignment AB -

Autosomal dominant neovascular inflammatory vitreoretinopathy (ADNIV) is an autoimmune condition of the eye that sequentially mimics uveitis, retinitis pigmentosa, and proliferative diabetic retinopathy as it progresses to complete blindness. We identified two different missense mutations in the CAPN5 gene in three ADNIV kindreds. CAPN5 encodes calpain-5, a calcium-activated cysteine protease that is expressed in retinal photoreceptor cells. Both mutations cause mislocalization from the cell membrane to the cytosol, and structural modeling reveals that both mutations lie within a calcium-sensitive domain near the active site. CAPN5 is only the second member of the large calpain gene family to cause a human Mendelian disorder, and this is the first report of a specific molecular cause for autoimmune eye disease. Further investigation of these mutations is likely to provide insight into the pathophysiologic mechanisms of common diseases ranging from autoimmune disorders to diabetic retinopathy.

VL - 8 IS - 10 ER - TY - JOUR T1 - Seroreactivity against aqueous-soluble and detergent-soluble retinal proteins in posterior uveitis. JF - Archives of ophthalmology Y1 - 2011 A1 - Ko, Audrey C A1 - Brinton, Jason P A1 - Mahajan, Vinit B A1 - Zimmerman, Bridget A1 - Brinton, Gregory S A1 - Stone, Edwin M A1 - Folk, James C A1 - Mullins, Robert F KW - Adolescent KW - Adult KW - Autoantibodies KW - Autoantigens KW - Blotting, Western KW - Electrophoresis, Polyacrylamide Gel KW - Eye Proteins KW - Female KW - Humans KW - Immunoglobulin A KW - Immunoglobulin G KW - Immunoglobulin M KW - Male KW - Middle Aged KW - Retina KW - Retinal Degeneration KW - Solubility KW - Uveitis, Posterior KW - Young Adult AB -

To characterize the seroreactivity against retinal proteins in patients with posterior uveitis, retinal disease of noninflammatory origin, and healthy controls.

VL - 129 IS - 4 U1 - http://www.ncbi.nlm.nih.gov/pubmed/21482867?dopt=Abstract ER - TY - JOUR T1 - Transgenic mice carrying the H258N mutation in the gene encoding the beta-subunit of phosphodiesterase-6 (PDE6B) provide a model for human congenital stationary night blindness. JF - Human mutation Y1 - 2007 A1 - Tsang, Stephen H A1 - Woodruff, Michael L A1 - Jun, Lin A1 - Mahajan, Vinit A1 - Yamashita, Clyde K A1 - Pedersen, Robert A1 - Lin, Chyuan-Sheng A1 - Goff, Stephen P A1 - Rosenberg, Thomas A1 - Larsen, Michael A1 - Farber, Debora B A1 - Nusinowitz, Steven KW - Adult KW - Animals KW - Cyclic Nucleotide Phosphodiesterases, Type 6 KW - Disease Models, Animal KW - DNA Mutational Analysis KW - Electroretinography KW - Female KW - Humans KW - Male KW - Mice KW - Mice, Inbred DBA KW - Mice, Transgenic KW - Mutation KW - Night Blindness KW - Phosphoric Diester Hydrolases KW - Retinal Degeneration KW - Transgenes AB -

Mutations in the beta-subunit of cGMP-phosphodiesterase (PDE6beta) can lead to either progressive retinal disease, such as human retinitis pigmentosa (RP), or stationary disease, such as congenital stationary night blindness (CSNB). Individuals with CSNB in the Rambusch pedigree were found to carry the H258N allele of PDE6B (MIM# 180072); a similar mutation was not found in RP patients. This report describes an individual carrying the H258N allele, who presented with generalized retinal dysfunction affecting the rod system and a locus of dysfunction at the rod-bipolar interface. Also described are preclinical studies in which transgenic mice with the H258N allele were generated to study the pathophysiological mechanisms of CSNB. While Pde6b(rd1)/Pde6b(rd1) mice have severe photoreceptor degeneration, as in human RP, the H258N transgene rescued these cells. The cGMP-PDE6 activity of dark-adapted H258N mice showed an approximate three-fold increase in the rate of retinal cGMP hydrolysis: from 130.1 nmol x min(-1) x nmol(-1) rhodopsin in wild-type controls to 319.2 nmol x min(-1) x nmol(-1) rhodopsin in mutants, consistent with the hypothesis that inhibition of the PDE6beta activity by the regulatory PDE6gamma subunit is blocked by this mutation. In the albino (B6CBA x FVB) F2 hybrid background, electroretinograms (ERG) from H258N mice were similar to those obtained from affected Rambusch family members, as well as humans with the most common form of CSNB (X-linked), demonstrating a selective loss of the b-wave with relatively normal a-waves. When the H258N allele was introduced into the DBA background, there was no evidence of selective reduction in b-wave amplitudes; rather a- and b-wave amplitudes were both reduced. Thus, factors other than the PDE6B mutation itself could contribute to the variance of an electrophysiological response. Therefore, caution is advisable when interpreting physiological phenotypes associated with the same allele on different genetic backgrounds. Nevertheless, such animals should be of considerable value in further studies of the molecular pathology of CSNB.

VL - 28 IS - 3 U1 - http://www.ncbi.nlm.nih.gov/pubmed/17044014?dopt=Abstract ER -