Mechanism of calcium phosphate stone formation in engineered 3D tubule

项目摘要/摘要 肾钙磷(CaP)和CaOx+CaOx混晶生物矿化结果 在肾结石等疾病中，在美国，影响超过10%的成年人，占超过 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5年内达到近50%。尽管在过去一个世纪里进行了广泛的研究， 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 开发一种可模拟动态生物的新型微流控工作台进行成矿 体内肾小管系统的条件。我们假设模仿体内CAP的过程 在微流体中创建的这个体外肾小管模型中的沉积将使我们能够系统地评估 分析各动力因素对帽晶形成的贡献及其多因素机制 微环境提示(例如，细胞调节、流体流体动力学和物理化学相互作用)。 此外，我们认为，原位表征技术和活体类3D管的结合 微流体中产生的微环境将使我们能够更好地了解 在分子和细胞水平上形成盖层结石。我们的程序是新颖的方法，它检查 持续流动的活体肾小管内帽状结石的形成机制。与之前的不同 我们建议对肾液的成分和肾脏中的固体矿物沉积进行原位测绘 具有连续流体流动的管状结构，这将使我们能够捕获最相关的分子， 传统方法无法获得的细胞和流体动力学信息。此外，我们的 专门的体外工具可以剖析体内复杂事件的细节，彻底改变我们的理解 在系统层面上的石头形成。这项研究计划涉及两个目标：i)了解 三维体外肾流体流动影响下细胞与冠状晶体的相互作用 在微流体中设计的肾小管结构，以及ii)了解生理学和 临床相关分子在保持，溶解，生长的帽和/或CaOx晶体内的MF- 以体外肾小管结构为基础。拟议的研究将引导我们：1)找到预防和治疗的策略 肾脏钙结石疾病；ii)对软组织中异常的帽子沉积有了新的认识(即， 骨骼外钙化)；以及iii)在理解分子和 在血管和其他类似的细胞微环境中形成帽状结石的药理学基础。 由于这种方法是如此之新，我们正在申请一项探索性拨款，以开发使能技术 用来阐明结石形成的一般机制。我们的目标是证明 从基础科学、工程学和生物学的角度论证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.