High accuracy optical growth assay of 3D cellular systems

3D 细胞系统的高精度光学生长测定

基本信息

批准号：
10330571
负责人：
Rashid Bashir
金额：
$ 45.82万
依托单位：
UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-02-01 至 2024-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10330571
关键词：
3-Dimensional Adopted Algorithms Anatomy Basic Science Biological Assay Biomedical Engineering Cell Therapy Cells Cellular biology Clinic Clinical Clinical Research Collaborations Coulter counter Disease Drug Targeting Escherichia coli Extracellular Matrix Proteins Fluorescence Fluorescence Microscopy Growth Image Individual Interference Microscopy Kinetics Label Laboratories Letters Light Mammalian Cell Measurement Measures Methodology Methods Microscope Microscopy Multimodal Imaging Neoplasm Metastasis Nuclear Optics Organ Organoids Osmotic Pressure Performance Phase Population Process Quantitative Microscopy Regulation Research Research Personnel Specimen Subcellular structure System Techniques Technology Thick Time Tissue Model Translating Variant Vision Work base biomedical scientist cell growth drug development human disease human model imaging modality in vivo instrument interest mathematical analysis multidisciplinary novel screening targeted treatment tool

项目摘要

Project Summary Growth regulation of mammalian cells has been described as "One of the last big unsolved problems in cell biology". The ability to measure accurately the growth rate of single cells has been the main obstacle in answering this question. From a clinical perspective, the basic understating of cell growth kinetics and how it is modulated by disease and treatment will allow for more targeted drug development. In recent years, there has been a significant interest in multidisciplinary work by biomedical engineers and scientists with a vision of developing 3D ex vivo tissue models of human organ function, anatomy, and disease. These 3D cellular systems are referred interchangeably as organoid, organotypic, or spheroid (spherical organoid). Organoids self-assemble under proper conditions, i.e., when relevant components, such as extracellular matrix (ECM) proteins, are present. Organoids are well documented to better recapitulate aspects of in vivo organ function and human disease. The common tool for analysis of such systems has been confocal (fluorescence) microscopy of fixed specimens. However, this approach does not reveal structural information in the center of the construct and, most importantly, is limited in terms of time-lapse imaging. There is a critical need for revealing subcellular structures in label-free mode with high contrast, which allows for dynamic, non- destructive imaging. At the same time, quantifying the dry mass of the organoid and its cellular components will inform on the basic organ function and disease, with and without treatment. Despite this critical need, a unified, easy-to-use methodology to measure the growth rate of individual cells and 3D constructs is lacking. Until recently, the state-of-the-art method to assess a single cell growth curve was using Coulter counters to measure the volume of a large number of cells, in combination with careful mathematical analysis. For relatively simple cells such as Escherichia coli (E. coli), traditional microscopy techniques have also been used to assess growth in great detail. In this type of method the assumption is that volume is a good surrogate for mass; however, this assumption is not always valid, for example due to variations in osmotic pressure. We propose to develop a practical dry mass assay for 2D cell populations, as well as 3D organoids, based on a novel imaging method developed in our laboratory: Spatial Light Interference Microscopy (SLIM) for 2D cultures and Gradient Light Interference Microscopy (GLIM) for 3D organoids. SLIM/GLIM takes advantage of the fact that optical phase delay accumulated through a live cell is linearly proportional to the dry mass (non-aqueous content) of the cell. Due to its particular interferometric principle, GLIM significantly suppresses multiple scattering and, as result, is capable of imaging thick specimens such as organoid/spheroids. The project aims to optimize and translate the composite SLIM/GLIM technology into a cell growth assay instrument that can be broadly adopted by researchers in both the research and pharma markets.

项目概要哺乳动物细胞的生长调节被描述为“细胞中最后一个未解决的大问题之一” 生物学”。准确测量单细胞生长速率的能力一直是该领域的主要障碍回答这个问题。从临床角度，对细胞生长动力学及其原理的基本理解受疾病和治疗的调节将允许更有针对性的药物开发。近年来，生物医学工程师和科学家对多学科工作产生了浓厚的兴趣。科学家们的愿景是开发人体器官功能、解剖学和疾病的 3D 离体组织模型。这些 3D 细胞系统可互换地称为类器官、器官型或球体（球形类器官）。类器官在适当的条件下自组装，即当相关组件，例如存在细胞外基质（ECM）蛋白。类器官有详细记录，可以更好地概括各个方面体内器官功能和人类疾病。分析此类系统的常用工具是共焦固定样本的（荧光）显微镜检查。然而，这种方法并没有揭示结构信息结构的中心，最重要的是，在延时成像方面受到限制。有一个关键的需要以高对比度的无标记模式揭示亚细胞结构，这允许动态、非破坏性成像。同时，量化类器官的干质量及其细胞成分将告知基本器官功能和疾病，无论是否接受治疗。尽管存在这一迫切需求，但仍需要一种统一、易于使用的方法来测量单个细胞的生长速度和缺乏 3D 构造。直到最近，评估单细胞生长曲线的最先进方法是使用库尔特计数器测量大量细胞的体积，并结合仔细的数学分析。对于大肠杆菌（E. coli）等相对简单的细胞，传统的显微镜检查技术也被用来详细评估生长情况。在这种类型的方法中，假设是体积可以很好地替代质量；然而，这个假设并不总是有效，例如由于渗透压的变化。我们建议开发一种适用于 2D 细胞群以及 3D 类器官的实用干质量测定法，基于我们实验室开发的一种新颖的成像方法：空间光干涉显微镜 (SLIM) 用于 2D 培养，梯度光干涉显微镜 (GLIM) 用于 3D 类器官。纤薄/闪亮利用通过活细胞累积的光学相位延迟是线性的这一事实与细胞的干质量（非水含量）成正比。由于其特殊的干涉测量原理上，GLIM 显着抑制多重散射，因此能够对厚层成像样本，例如类器官/球体。该项目旨在优化和转化复合材料将SLIM/GLIM技术打造为可被研究人员广泛采用的细胞生长检测仪器在研究和制药市场。

项目成果

期刊论文数量（8）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（1）

Network science characteristics of brain-derived neuronal cultures deciphered from quantitative phase imaging data

DOI：
10.1038/s41598-020-72013-7
发表时间：
2020-09
期刊：
Scientific Reports
影响因子：
4.6
作者：
Chenzhong Yin;Xiongye Xiao;Valeriu Balaban;M. Kandel;Y. J. Lee;G. Popescu;P. Bogdan
通讯作者：
Chenzhong Yin;Xiongye Xiao;Valeriu Balaban;M. Kandel;Y. J. Lee;G. Popescu;P. Bogdan

Cell-to-cell influence on growth in large populations

DOI：
10.1364/boe.10.004664
发表时间：
2019-09-01
期刊：
BIOMEDICAL OPTICS EXPRESS
影响因子：
3.4
作者：
Kandel, Mikhail E.;Lu, Wenlong;Popescu, Gabriel
通讯作者：
Popescu, Gabriel

Large-scale phase retrieval.

DOI：
10.1038/s41377-021-00616-4
发表时间：
2021-09-03
期刊：
Light, science & applications
影响因子：
0
作者：
Popescu G
通讯作者：
Popescu G

Engineering geometrical 3-dimensional untethered in vitro neural tissue mimic

工程几何三维无束缚体外神经组织模拟

DOI：
10.1073/pnas.1916138116
发表时间：
2019
期刊：
Proceedings of the National Academy of Sciences
影响因子：
0
作者：
Pagan-Diaz, Gelson J.;Ramos-Cruz, Karla P.;Sam, Richard;Kandel, Mikhail E.;Aydin, Onur;Saif, M. Taher;Popescu, Gabriel;Bashir, Rashid
通讯作者：
Bashir, Rashid

Epi-illumination gradient light interference microscopy for imaging opaque structures