Bruch's Membrane-On-A-Chip to Study Drusen Formation in AMD

ABSTRACT Age-related macular degeneration (AMD) is a major cause of blindness in developed nations. This debilitating condition currently impacts 196 million people worldwide, with projections indicating a rise to 288 million by 2050. The majority of affected individuals experience the early and intermediate stages of "dry" AMD, for which no treatments are available due to an incomplete understanding of the disease mechanisms. In these stages, extensive lipoprotein-rich deposits, known as drusen, accumulate in the macula, impairing vision. Although drusen are a hallmark of AMD, the exact mechanism of their formation remains unclear. Drusen develop in the extracellular matrix (ECM) between the basal lamina of the retinal pigment epithelium and the inner collagenous layer of Bruch’s membrane (BrM). Our lab has shown that the glycosaminoglycan, heparan sulfate (HS), is elevated in patients with early and intermediate AMD and regulates lipoprotein retention. Alterations in glycosaminoglycans are observed with aging and in various diseases, including atherosclerosis. These changes have been shown to regulate the retention of lipoproteins in the ECM, leading to the aggregation of lipoproteins and the formation of atheromas. Alteration of the retention of lipoproteins in BrM has the potential to reveal novel therapeutics that could remove drusen in the early stages of AMD. Nevertheless, our progress had been hindered by the lack of meaningful model systems allowing for quantitative analysis of BrM binding dynamics in the context of AMD. To investigate this chemistry, our lab developed a novel Quartz Crystal Microbalance assay specific to human BrM tissue (QCM-BrM), enabling precise binding kinetics studies under more natural conditions than previously possible. In this exploratory research proposal using QCM-BrM, we aim to investigate the determinants of lipoprotein retention in BrM, including apolipoprotein composition (SA1) and BrM HS content (SA2), and explore the pharmacologic potential of novel therapeutics (SA3). The QCM-BrM technology represents a significant advancement in the study of ECM biology, particularly within the context of AMD. The purpose of this application is to investigate the utility and versatility of the QCM- BrM technology using lipoprotein and HS binding in BrM as a proof of principle for investigating natural ECM-on-a-chip. Future studies will explore the impact of AMD-risk alleles and protease activity on BrM ECM kinetics and investigate binding kinetics of other drusen associated proteins and extracellular vesicles in BrM aggregation. In addition, we plan to collaborate with ECM biology researchers to extend our BrM-on-a-chip technology to additional ECM systems, such as the arterial vessel wall in atherosclerosis.