Three iPSC-derived cell models for the study of Stargardt Disease and Retinitis Pigmentosa.

Abstract

Purpose : Stargardt disease (STGD) and Retinitis Pigmentosa (RP) are two of the most prevalent inherited retinal dystrophies (IRD). The first one is caused by recessive mutations in the ABCA4 gene, an ABC-transporter expressed in both retinal pigmented epithelia (RPE) cells and photoreceptors. Loss of ABCA4 function or expression leads to the accumulation of bisretinoid compounds and loss of central vision that can evolve further to the periphery.
RP is the most common form of rod-cone dystrophy, which starts with night blindness followed by a progressive loss of the peripheral visual field. More than 100 genes have been identified as the cause of either autosomal recessive or dominant RP. RHO - the gene encoding for the specific opsin of rods – is one of the most prevalent in the dominant forms. Specific mutations carried by IRD patients can be modeled using induced Pluripotent Stem Cells (iPSC).

Methods : We generated 3 different iPSC-derived cell models: RPE, photoreceptor precursors (PhRPs) and Retinal Organoids (RO) derived from iPSC of 3 patients affected by STGD disease or RP (with a dominant RHO mutation) and 2 healthy individuals. Using qPCR, Western Blot, and immunochemistry, along with other cellular biology methods, we analyzed the phenotype caused by these mutations.

Results : The effect of patient-specific mutations induces aberrant splicing, reduced mRNA expression and reduced protein expression of ABCA4 while showing early signs of autophagy and reduced function on RPE, PhRPs and RO. The dominant mutation in the RHO gene shows accumulation of the rhodopsin protein in rod photoreceptors, and early signs of endoplasmic reticulum (ER) stress and autophagy.

Conclusions : iPSC are a wonderful tool for the investigation of retinal diseases, being able to differentiate into the different cellular types of the retina. While some models with a faster differentiation time can be useful for detection of mRNA or protein expression, more complicated models such as RO are necessary for a deeper insight into the pathophysiology of each disease.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.

 

33 week-old RO obtained from a patient with RP. The photoreceptor layer is found on the external side of the organoid while the rest of neuronal cells are organized in the inner zone.

 

Segment of a 33 week-old RO showing the rod-specific marker Rhodopsin (red) and the cone-specific marker Opsin M/L (green). Nuclei are marked in blue.