Project 3 - Mechanisms of extra- and intra-cellular calcification

PROJECT 3 SUMMARY Mechanisms of extra- and intra-cellular calcification. Recent findings from our laboratory showed that extracellular calcification is an integral part of the hallmark lesions of age-related macular degeneration (AMD). Calcified deposits rich in the calcium phosphate mineral hydroxyapatite (HAP) accumulate in the space under the retinal pigment epithelium (RPE) in the back of the eye, where nutrient uptake and clearance of unwanted material usually are exchanged between the blood circulation and the RPE cells. Inhibition of this exchange process can lead to rapid progression to an end-stage disease characterized by irreversible visual loss. Apart from AMD, sub-RPE deposit formation is also associated with other diseases such as Alzheimer’s disease (AD). Based on recent findings, we proposed a new model for sub-RPE deposit initiation and growth where calcification plays a pivotal role. We know from clinical and basic science studies that calcification, and the associated interactions with proteins and lipids, have a negative effect on disease progression. We also learned that understanding how different mineralization forms contribute to diseases is critical. However, we do not yet know how calcification occurs and what intra- and extracellular molecules and pathways contribute to mineral deposition in the sub-RPE space. This project will study the cellular and systemic contributors to extracellular mislocalized (ectopic) calcification using a cellular model system we developed to study the initiation and progression of the mineral-laden sub-RPE deposits. We will also develop a significantly improved, new 3D microfluidic cell model to study calcification, as sub-RPE deposit formation, development and progression of AMD and AD are linked to the choroidal endothelium and its degeneration. The results will be validated using human postmortem eye tissues from patients with AMD and AD. The validation and the experimental plan using the 2D and 3D cell culture systems will provide a robust foundation for developing new intervention strategies centered on the calcification of the sub-RPE deposits. Apart from AMD and AD, these treatment strategies will also be applicable for other diseases in which sub-RPE deposits play a role.