Osteocyte Signaling Within Mineralized Lacuna-Canaliculi Microenvironment

矿化腔隙-小管微环境中的骨细胞信号传导

基本信息

批准号：
10240448
负责人：
Pranav Soman
金额：
$ 16.01万
依托单位：
SYRACUSE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-09-01 至 2023-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10240448
关键词：
3D Print Ablation Biochemical Biocompatible Materials Body Fluids Bone Diseases Bone Matrix Calcium Signaling Cell Line Cells Cellular biology Characteristics Confocal Microscopy Dendritic Cells Diffusion Disease Evaluation Fluorescence Microscopy Functional disorder Gap Junctions Gases Gelatin Goals Homeostasis Hybrids Hydrogels Hypoxia Incubators Individual Lasers Location Measures Mechanical Stimulation Mechanics Methacrylates Methods Microfluidic Microchips Microfluidics Minerals Modeling Morphology Mus Nutrient Nutrient Canals Optics Orthopedic Procedures Osteocytes Osteon Pathology Patients Periodicity Physiological Play Printing Property Refractory Risk Signal Transduction Speed Structure Technology Testing Time Time Study Work base bone bone cell calcification cell injury design high risk hormonal signals in vitro Model in vivo inhibitor/antagonist insight mechanical force mechanotransduction new technology novel therapeutics preservation shear stress skeletal skeletal abnormality therapeutic target

项目摘要

Summary Although it is widely accepted that osteocytes regulate bone homeostasis by sensing, integrating and transducing mechanical and hormonal signals, characterization of dynamic signaling within the osteocyte network has been challenging due to its location embedded within the bone matrix. Osteocytes reside within a mineralized lacunar-canalicular (MLC) structure allowing sensing of mechanical forces and transduction this signal through gap-junctions and secreted exchange of soluble biochemical signals. The MLC structure modulates access of essential nutrients between vasculature and entombed osteocytes in a spatially gradient manner. New understanding on osteocyte signaling will be necessary to develop new therapeutics for treating diseases that involve osteocyte dysfunction. To that end, the goal of this work is to develop a new in vitro model that will not only mimic the in vivo like MLC structure, but also facilitate the study of signaling dynamics within an osteocyte network upon targeted mechanical stimulation or cell damage. The hypothesis that, “the nutrient gradient that osteocyte encounter is a function of the mineralized lacunar-canalicular (MLC) structure, which in turn regulates their signal propagation dynamics”, will be tested using three specific aims. Aim 1 will use a Hybrid Laser Printing (HLP) platform to develop a microfluidic chip that mimics the MLC structure with associated gradient nutrient transport properties. Aim 2 will identify experimental conditions to generate osteocyte network within MLC chips using the mouse MLO-Y4 osteocyte cell line. Aim 3 will characterize propagation characteristics of calcium signaling (amplitude, range, velocity, refractory period, spike-synchrony) within osteocyte networks upon targeted mechanical stimulation, cell-damage, ablation of cell-cell connections, or in the presence of signaling inhibitors. In summary, individual and combined effects of (i) MLC structure-induced gradient nutrient access (ii) mineralized matrix, (iii) environmental hypoxia, and (iv) single cell manipulation, on calcium signaling dynamics will provide new insights into osteocyte mechanotransduction. In the long term, this model can be extended to patient-specific cells to screen therapeutics that target skeletal pathologies associated with osteocyte malfunctions.

概括尽管广泛认为骨细胞通过传感，整合和机械和马信号转导，骨细胞内动态信号的表征由于其位置嵌入了骨基质中，因此受到挑战。骨细胞位于矿化的lacunar-canscular（MLC）结构，允许机械力的灵敏度并翻译这通过间隙 - 缝线和固体生化信号的分泌交换信号。 MLC结构调节在空间梯度中的脉管系统和吞噬骨细胞之间的必需营养方式。对于开发用于治疗的新疗法将需要对骨细胞信号传导的新了解涉及骨细胞功能障碍的疾病。为此，这项工作的目标是开发一种新的体外模型这不仅会模仿体内的MLC结构，还可以促进研究信号动力学的研究靶向机械刺激或细胞损伤后的骨细胞网络。假设“营养素骨细胞相遇的梯度是矿化lacunar-cancancular（MLC）结构的函数，该结构在转向调节其信号传播动力学”将使用三个特定目标进行测试。AIM1将使用混合动力激光打印（HLP）平台开发一个微流体芯片，该芯片模拟MLC结构与相关的结构梯度营养转运特性。 AIM 2将确定实验条件以生成骨细胞网络使用小鼠MLO-Y4骨细胞系中的MLC芯片中。 AIM 3将表征传播特征钙信号传导（幅度，范围，速度，难治性周期，尖峰同步）在整骨细胞网络中有针对性的机械刺激，细胞破坏，细胞 - 细胞连接的消融或存在信号抑制剂。总而言之，（i）MLC结构诱导的梯度养分的单个和组合效应访问（ii）矿化基质，（iii）环境缺氧和（iv）单细胞操作，钙信号传导动力学将为骨细胞机械转移提供新的见解。从长远来看，这个模型可以是扩展到患者特异性细胞到筛查治疗，以靶向与骨细胞故障。