Mechanism of calcium phosphate stone formation in engineered 3D tubule

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Renal Calcium phosphate (CaP) and CaP plus Calcium Oxalate (CaOx) mixed crystal biomineralization results in diseases such as nephrolithiasis, which affect over 10% of adults in the United States, accounting for over $5 billion in economic costs in this country each year. The recurrence rate of nephrolithiasis is often high, reaching almost 50% over a 5-year period. Despite extensive research being conducted over the past century, the mechanism by which crystals nucleate, grow, and aggregate into stones is still poorly understood. This program seeks to improve our understanding of the origin and mechanism of such biologically controlled mineralization by developing a novel microfluidic-based workbench that can simulate the dynamical biological conditions of an in vivo renal tubular system. We hypothesize that mimicking the process of in vivo CaP deposition within this ex vivo tubular model created in microfluidics will enable us to systematically evaluate the multifactorial mechanism of CaP crystal formation by analyzing the contribution of each dynamic microenvironmental cue (e.g., cellular regulations, fluidic hydrodynamics and physiochemical interactions). Further, we believe that the combination of in situ characterization techniques and in vivo-like 3D tubular microenvironment generated in microfluidics will enable us to achieve a better understanding of the process of CaP stone formation at molecular and cellular level. Our program is novel in its approach to examine the mechanism of CaP stone formation in in vivo-like tubules with continuous renal fluidic flow. Unlike previous attempts, we propose to in situ map the compositions of renal fluids and solid mineral depositions in renal tubular structures with continuous flow of fluids, which will enable us to capture the most relevant molecular, cellular and hydrodynamic information that are not attainable by conventional approaches. Further, our specialized ex vivo tools can dissect the details of the complex in vivo event, revolutionizing our understanding of stone formation at the systems level. This research program involves two objectives: i) To understand the cellular interaction with CaP crystals under the influence of hydrodynamic renal fluid flow within 3D ex vivo renal tubular structure engineered in microfluidics, and ii) To understand the role of physiologically and clinically relevant molecules in the retention, dissolution, growth of CaP and/or CaOx crystals within the MF based ex vivo renal tubular structure. The proposed study will lead us to i) find strategies to prevent and treat calcium stone disease in kidney; ii) gain new insights on the abnormal CaP deposition in soft tissues (namely, extra-skeletal calcification); and iii) open up exciting research fronts in understanding the molecular and pharmacological basis of CaP stone formation in vascular and other similar cellular microenvironments. Because the approach is so new, we are requesting an exploratory grant to develop the enabling techniques required for the elucidation of the general mechanism of stone formation. Our objective is to demonstrate the feasibility of our approach from a fundamental science, engineering and biological perspective.