Mechanism of calcium phosphate stone formation in engineered 3D tubule

项目总结/摘要 肾脏磷酸钙（CaP）和CaP加草酸钙（CaOx）混合晶体生物矿化结果 在美国，肾结石等疾病影响超过10%的成年人，占美国人口的10%以上。 每年在这个国家造成50亿美元的经济损失。肾结石的复发率往往很高， 在五年内达到近50%。尽管在过去的世纪进行了广泛的研究， 晶体成核、生长和聚集成石头的机制仍然知之甚少。这 该计划旨在提高我们对这种生物控制的起源和机制的理解。 通过开发一种新型的基于微流体的工作台，可以模拟动态生物矿化， 体内肾小管系统的条件。我们假设，模拟体内钙磷代谢过程 在微流体中产生的这种离体管状模型内的沉积将使我们能够系统地评估 通过分析各动力学因素对CaP晶体形成的贡献， 微环境线索（例如，细胞调节、流体流体动力学和生理化学相互作用）。 此外，我们认为，结合原位表征技术和体内类3D管状 在微流体中产生的微环境将使我们能够更好地理解 钙磷结石形成的分子和细胞水平。我们的计划是新颖的，在其方法来研究 CaP结石形成的机制在体内样小管与连续肾流体流。不同于以往 尝试，我们建议原位映射肾液和固体矿物沉积物的组成，在肾 管状结构与连续流动的流体，这将使我们能够捕捉最相关的分子， 细胞和流体动力学信息，这是传统方法无法获得的。此外，我们的 专门的体外工具可以剖析复杂的体内事件的细节，彻底改变我们的理解 在系统层面上形成石头。这项研究计划涉及两个目标：一）了解 在离体三维肾流体动力学流动的影响下细胞与CaP晶体的相互作用 在微流体中工程化的肾小管结构，和ii）为了理解生理和 在MF内的CaP和/或CaOx晶体的保留、溶解、生长中的临床相关分子， 基于离体肾小管结构。这项拟议中的研究将引导我们i）找到预防和治疗的策略 肾脏中的钙结石疾病; ii）获得对软组织中异常CaP沉积的新认识（即， 骨骼外钙化）;和iii）在理解分子和 在血管和其他类似的细胞微环境中CaP结石形成的药理学基础。 因为这种方法是如此的新，我们正在申请一笔探索性的赠款，以开发这种使能技术。 阐明石头形成的一般机制所需的。我们的目标是展示 从基础科学、工程学和生物学的角度来研究我们的方法的可行性。

Project Summary/Abstract 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.