Mechanistic Study of Developmental Neurotoxicity on 3D Cultured Stem Cell Microarrays

3D 培养干细胞微阵列的发育神经毒性机制研究

基本信息

批准号：
8944604
负责人：
MOO-YEAL LEE
金额：
$ 32.45万
依托单位：
CLEVELAND STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-07-01 至 2019-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8944604
关键词：
ABCB1 gene ABCG2 gene Acute Adult Adverse effects Affect Animal Model Apoptosis Astrocytes Binding Biological Assay Biological Markers Biomimetics Brain Cell Culture System Cell Culture Techniques Cell Differentiation process Cell Surface Receptors Cell physiology Cells Cellular Morphology Cytochrome P450 Cytochromes Data Set Development Embryo Encapsulated Enzymes Extracellular Matrix Glutathione Goals Growth Health Hepatocyte Human Hydrogels Image In Vitro Individual Ion Channel Lead Measures Membrane Potentials Metabolism Methodology Modeling Molecular National Institute of Environmental Health Sciences National Institute of General Medical Sciences Necrosis Neurites Neurons Nuclear Outcome Oxidative Stress Pathway interactions Pharmaceutical Preparations Pharmacologic Substance Physiological Poison Recombinants Research Robotics Role Shapes Staging Stem Cell Development Stem cells Subfamily lentivirinae System Technology Testing Therapeutic Time Tissues Toxic Environmental Substances Toxic effect Toxicokinetics Toxicology Work base biochip cytotoxic cytotoxicity developmental disease developmental neurotoxicity developmental toxicity developmental toxicology disorder prevention drug candidate drug discovery high throughput screening improved in vivo inhibitor/antagonist miniaturize mitochondrial membrane nerve stem cell neurogenesis neuronal cell body neurotoxicity neurotoxicology oligodendrocyte lineage preclinical evaluation public health relevance receptor response screening tool toxicant trait

项目摘要

DESCRIPTION: There is a critical need for improved in vitro human toxicology testing to screen and identify potential toxic compounds, the levels at which they are lethal, and evaluate their developmental toxicity on human neural stem cells (NSCs) at the cellular and molecular levels. In particular, understanding mechanisms underlying human toxicity and the role of compounds including environmental toxicants and drug candidates is one of major goals of federal agencies such as NIEHS, NIGMS, and EPA, and has important implications in human health and disease prevention. Although animal models and primary human cells have been extensively used for such toxicology studies, their use is limited by high species variability with little or no predictability of relevance to humans, the instability of primary cells, and insufficient supply of human primary cells including NSCs for high-throughput screening. Therefore, there is an urgent need to develop in vitro strategies to rapidly assess compound-induced toxicity, and accurately predict adverse responses in vivo. Using three-dimensional (3D) cultured, human NSCs, together with high-throughput methodology and high-content imaging (HCI), we propose to decipher the cellular and molecular mechanisms underlying the effects of toxic model compounds. The core hypotheses are: (i) miniaturized 3D NSC microarrays can maintain high neuronal cell functions by better mimicking in vivo microenvironments; (ii) blocking ion channels and transporters on NSCs can modulate cell differentiation and cytotoxicity; (iii) physiological in vivo effects of compounds and their mechanistic actions on NSCs could be replicated and elucidated in vitro via high-throughput, high- content cell function analysis; and (iv) such miniaturized 3D cell culture systems can be used to facilitate mechanistic toxicology assays, which in turn can improve predictability of toxicity in vivo. The specific aims of the proposed work are to (1) demonstrate high neuronal cell functions on 3D NSC microarrays within biomimetic microenvironments in high throughput on the chip, (2) investigate mechanistic actions of various classes of compounds on NSC microarrays using high-throughput, ion channel and transporter assays, and (3) establish HCI assays on 3D NSC microarrays to investigate mechanistic profiles of toxicity by compounds and their metabolites. This information is essential to better understand the mechanistic basis of pharmaceutical toxicology on embryonic and adult human cells and tissues, and prioritize environmental toxicants based on their potential adverse effects on humans.

描述：迫切需要改善体外人类毒理学测试，以筛选和鉴定潜在的毒性化合物，其致死水平，并评估其在细胞和分子水平上对人神经元干细胞（NSC）的发育毒性。特别是，了解人类毒性的基础机制以及包括环境有毒物质和候选药物在内的化合物的作用是NIEHS，NIGMS和EPA等联邦机构的主要目标之一，并且对预防人类健康和疾病具有重要意义。尽管动物模型和原代人细胞已被广泛用于这种毒理学研究，但它们的使用受到高物种可变性的限制，几乎没有或没有与人相关的可预测性，原代细胞的不稳定性以及不足的供应人类原代细胞，包括用于高通量筛查的NSC。因此，迫切需要制定体外策略来快速评估复合诱导的毒性，并准确预测体内不良反应。使用三维（3D）培养的人类NSC，以及高通量方法论和高含量成像（HCI），我们建议破译有毒模型化合物作用的细胞和分子机制。核心假设为：（i）微型3D NSC微阵列可以通过更好地模仿体内微环境来维持高神经元细胞功能；（ii）在NSC上阻断离子通道和转运蛋白可以调节细胞分化和细胞毒性；（iii）物理化合物的体内影响及其对NSC的机械作用可以通过高通量，高含量细胞功能分析在体外复制和阐明；（iv）这种微型3D细胞培养系统可用于促进机械毒理学测定，进而可以提高体内毒性的可预测性。拟议工作的具体目的是（1）在芯片上高吞吐量中的3D NSC微阵列上表现出高神经元细胞的功能，（2）研究各种化合物在NSC微阵列上使用高容器，离子通道和转运仪和（3）建立HCI的机构进行的机械作用，并在HC上进行了调查。化合物及其代谢产物的毒性。该信息对于更好地了解胚胎和成人人类细胞和组织的药物毒理学的机理基础至关重要，并根据对人类的潜在不利影响对环境毒物进行优先级。