Using animal models to examine mechanisms and treatments of Retinitis Pigmentosa

Sara Kathryn Mayer

doi:10.25820/etd.006854

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Dissertation

Using animal models to examine mechanisms and treatments of Retinitis Pigmentosa

Sara Kathryn Mayer

University of Iowa

Doctor of Philosophy (PhD), University of Iowa

Autumn 2023

DOI: 10.25820/etd.006854

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Abstract

Retinitis pigmentosa (RP) is a progressive genetic disorder in which retinal degeneration leads to blindness. It occurs in 1 in 4,000 people worldwide. RP can be caused by mutations in over 300 genes and presents as either syndromic or non-syndromic. Syndromic RP is characterized by retinal degeneration in combination with defects in other organ systems. Non-syndromic RP is characterized by retinal degeneration alone. Early stages of RP involve the death of rod photoreceptors via apoptosis, causing loss of peripheral and night vision. This is followed by the death of cone photoreceptors, followed by the loss of central vision. Mechanisms of RP are not well understood; however, there is an association with oxidative stress. I studied Bardet Biedl Syndrome Type 10 (BBS10) as an example of an autosomal recessive syndromic form of RP. In addition to retinal degeneration, patients can experience obesity, polydactyly, and kidney disease. BBS10 is caused by mutations in the gene BBS10, which encodes a chaperonin-like complex protein. This complex is responsible for BBSome assembly, while the BBSome itself is necessary for primary cilium maintenance and function. Mutations in BBS10 cause up to 20% of all BBS cases, meaning treatments for specifically BBS10 would benefit a large portion of BBS patients. To investigate treatment options, a mouse model was developed. Mice lacking Bbs10, Bbs10-/- mice, have similar phenotypes to humans with BBS10, including obesity and progressive retinal degeneration, demonstrated by electroretinogram (ERG), optical coherence tomography (OCT), and visually guided swim assay. In humans, some forms of BBS are more severe than others and BBS10 is more severe than BBS1. BBS1 is another commonly mutated gene that causes BBS; the BBS1 protein is a part of the BBSome itself. I determined that Bbs10-/- mice have more severe retinal degeneration compared to a mouse model of BBS1, Bbs1M390R/M390R mice, similar to the relationship observed in humans. Given the association of RP with oxidative stress, I decided to combat retinal degeneration by targeting oxidative stress with the antioxidant N-acetylcysteine, which increases levels of glutathione. When given N-acetylcysteine, retinal degeneration in Bbs10-/- was slowed. As a comparison experiment, buthionine sulfonimine, which decreases levels of glutathione, was given to Bbs10-/- mice. This treatment did not have a noticeable effect. I also studied an autosomal dominant, non-syndromic form of RP caused by mutations in the SNRNP200 gene. SNRNP200 encodes a core splicing factor necessary for the catalytic step in pre-mRNA splicing. It is unknown how mutations in a vital splicing factor cause a retina specific disorder. To investigate the molecular defects associated with SNRNP200-associated RP and discover potential treatments, Drosophila models were developed and characterized. Drosophila has an orthologue of SNRNP200 that I will refer to as dSnrnp200, encoding a protein with 74% amino acid identity and 86% similarity. Depletion of dSnrnp200 using RNAi in the developing eye causes a rough eye phenotype that is due to increased apoptosis. Human RP mutations that cause the amino acid substitutions T731I and S1087L were modeled in Drosophila dSnrnp200. These mutants display photoreceptor defects that include abnormal photoreceptor spacing and mitochondrial defects. Consistent with these defects, adult flies have abnormal electroretinogram wave forms. Treatment with N-acetylcysteine the rough eye phenotype in the flies depleted for dSnrnp200 and the retinal defects in flies possessing mutant alleles. Taken together, BBS10 and SNRNP200-assoicated RP can be accurately modeled in animals such as mice and flies, respectively. Here, I show that the Bbs10-/- mice recapitulate aspects of human BBS10. I also show that Drosophila models of SNRN200-associated RP have photoreceptor defects. Both model organisms responded positively to treatment with antioxidant N-acetylcysteine. This suggests N-acetylcysteine is a potential treatment for BBS10 and SNRNP200-associated RP as well as for RP caused by other gene mutations.

Retinitis Pigmentosa

Bardet-Biedl Syndrome

Drosophila

Mouse

SNRNP200

Details

Title: Subtitle: Using animal models to examine mechanisms and treatments of Retinitis Pigmentosa
Creators: Sara Kathryn Mayer
Contributors: Arlene V Drack (Advisor)
Lori L Wallrath (Advisor)
Sheila A Baker (Committee Member)
Alina V Dumitrescu (Committee Member)
Val C Sheffield (Committee Member)
Richard J H Smith (Committee Member)
Resource Type: Dissertation
Degree Awarded: Doctor of Philosophy (PhD), University of Iowa
Degree in: Genetics
Date degree season: Autumn 2023
Publisher: University of Iowa
DOI: 10.25820/etd.006854
Number of pages: xix, 215 pages
Grant note: I am grateful for the funding sources that made my PhD thesis research possible. This includes the NIH T32 Genetics Training Grant and Philanthropy funds. (vi)
Language: English
Date submitted: 11/28/2023
Description illustrations: illustrations, tables, graphs
Description bibliographic: Includes bibliographical references (pages 185-215).
Public Abstract (ETD): Retinitis pigmentosa (RP) is a genetic disorder which causes loss of vision, specifically through the death of the cells responsible for vision called photoreceptors. People first will lose their night vision and can progress to a loss of daytime vision, which can ultimately lead to blindness. People can be diagnosed with RP through clinical tests such as an electroretinogram (ERG). RP is present in approximately 1 in 4,000 people worldwide. RP has many genetic causes and disease progression is not fully understood. There is evidence that damaging molecules called reactive oxygen species contribute to cell death in RP due to a process called oxidative stress. This can arise due to abnormal cell function which can lead to cell damage and death. To study RP and its causes, animal models are used. We can also use these models to investigate potential treatments for RP, which can slow or even reverse blindness. Here I used two different animal models for two forms of RP, BBS10 and SNRNP200- associated RP in mice and fruit flies, respectively. Both models were able to mirror aspects of their respective human disease, including retinal cell loss. I then treated these models with the antioxidant N-acetylcysteine, NAC, which will combat oxidative stress. I saw that upon NAC treatment, retinal defects were lessened in both models, suggesting that treatments with antioxidants may be a path forward to help those with RP.
Academic Unit: Interdisciplinary Graduate Program in Genetics
Record Identifier: 9984546541902771

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