Aging, aniridia, and injury: transcriptomic perspectives on the molecular pathology of the ocular lens
| Author(s) | Faranda, Adam P. | |
| Date Accessioned | 2023-10-09T17:45:56Z | |
| Date Available | 2023-10-09T17:45:56Z | |
| Publication Date | 2023 | |
| SWORD Update | 2023-09-20T19:15:49Z | |
| Abstract | By helping to focus incoming light, the lens of the eye has an essential role in the vertebrate visual system. Consisting of two basic cell types, the lens has a simple yet sophisticated internal architecture and biochemistry. The growth of the mammalian lens continues indefinitely over a lifetime as new cell layers accumulate over older ones in a precise pattern of cell packing, via a differentiation process that culminates with near-total elimination of cellular organelles. Without the ability to construct new cellular components, cellular integrity in deeper layers can become compromised over time if cumulative insults exceed the capacity of the protective mechanisms maintained by the outermost metabolically active cells. Disruption of lens cell differentiation or failure of these protective mechanisms can lead to protein insolubility, damage to cell membranes, morphological defects or disorganization of cellular packing that produces opacities or refractive aberrations. The accumulation of opacified lens tissue results in a cataract, a partial or complete opacification of the lens. While genetic factors and exposure to various environmental hazards can directly induce lens opacities or increase one’s risk, one of the most important risk factors for cataract is aging. ☐ Aging is a complex biological process associated with physiological deterioration that involves transcriptome-wide changes in gene expression. RNA Sequencing (RNA-Seq) is an established technology for profiling transcriptomic shifts related to aging and other drivers of cataract formation. In Chapter 3, I address the impact of unique lens cell characteristics, such as the small size of the lens epithelial cell population, and the presence of partially degraded RNA in lens fiber cells, on RNA-Sequencing data generation and analysis. Chapter 3 explores an approach for incorporating various bioinformatic tools into a cohesive pipeline, evaluates the performance of these tools on several lens data sets, and shows how variation in RNA integrity may introduce certain unavoidable biases. The pipeline described in this chapter was subsequently used to investigate the consequences of aging on the mouse lens, and the impact of aging on the response of lens cells to cataract surgery. ☐ The micro-surgical restoration of vision in cataract patients is nothing short of a medical miracle, however a small but significant fraction of patients develop posterior capsular opacification (PCO), a vision compromising sequelae of cataract surgery requiring surgical follow up. Inflammatory signaling and fibrotic cell-transformation have a central role in driving the development of PCO in cataract patients. These pathways are attractive therapeutic targets for PCO prevention and can themselves lead to cataract formation. The mouse lens-aging study documented in Chapter 4 was originally motivated by a comment from a fellow lens biologist that young mice are prone to inflammation, and the inflammatory lens-injury response we observe in mice less than 5 months old may not occur in older mice or older humans for that matter. Not only did this study categorically demonstrate that the lens in geriatric mice is equally susceptible to injury dependent inflammation, it also revealed age-dependent increases in basal expression of certain inflammatory markers. These and other transcriptomic shifts observed in the aging mouse lens affecting oxidative stress response genes, and key regulators of fiber cell differentiation, may have important roles in age related cataract. ☐ The intent of Chapter 5 was to advance our knowledge of the aging biology of the lens from mice to humans. While there are acknowledged limitations of this study’s design, it was possible to identify age dependent changes in human-lens gene expression affecting genes that are likely relevant to cataract formation, for example we observe upregulation of the transcription factor KLF4 in aging humans, and its murine homolog Klf4 in aging mice. Klf4 regulates lens maturation in mice, and single nucleotide polymorphism (SNP) mutations at the KLF4 locus are associated with age-related cataract in humans. ☐ While most people maintain lens clarity beyond their 50’s, genetic abnormalities can dramatically accelerate the process, one example being PAX6 haploinsufficiency. A keystone in the process of embryonic eye and lens development, the transcription factor PAX6 continues to have a vital role in the lifelong maintenance of lens homeostasis. With only a single functional copy of PAX6, haploinsufficient individuals develop a pan-ocular syndrome known as aniridia. Juvenile cataract is one of several of the sequelae of aniridia that compromise vision, and frequently occurs at ages as young as 1 to 3 years old. To address a significant knowledge gap in the molecular etiology of aniridic cataract, Chapter 6 examines the impact of Pax6 haploinsufficiency on the mouse lens transcriptome. This analysis identified profound changes in gene expression that imply a radical disruption of the normal fiber cell differentiation process, which may be responsible for the visible lens defects we observe, and lead to the activation of inflammatory and fibrotic markers. My analysis shows that PAX6 haploinsufficiency leads to transcriptomic changes in the lens that normally characterize the response of wildtype lens cells to injury. These findings, which include an inflammatory response, support the hypothesis that PAX6 haploinsufficient lenses persist in a chronically wounded state. Throughout this work, inflammation appears to be a common thread that unifies the lens pathology associated with aging, injury and aniridia. | |
| Advisor | Duncan, Melinda K. | |
| Degree | Ph.D. | |
| Department | University of Delaware, Center for Bioinformatics and Computational Biology | |
| DOI | https://doi.org/10.58088/a3br-d897 | |
| Unique Identifier | 1416354164 | |
| URL | https://udspace.udel.edu/handle/19716/33482 | |
| Language | en | |
| Publisher | University of Delaware | |
| URI | https://login.udel.idm.oclc.org/login?url=https://www.proquest.com/dissertations-theses/aging-aniridia-injury-transcriptomic-perspectives/docview/2866692493/se-2?accountid=10457 | |
| Keywords | Inflammation | |
| Keywords | Lens | |
| Keywords | Transcriptomics | |
| Keywords | Fibrotic markers | |
| Keywords | Genetic abnormalities | |
| Title | Aging, aniridia, and injury: transcriptomic perspectives on the molecular pathology of the ocular lens | |
| Type | Thesis |