3D human lung model to study lung disease and formation of fibrosis

用于研究肺部疾病和纤维化形成的 3D 人体肺部模型

• 批准号: 8415381
• 负责人: JOAN E NICHOLS
• 金额: $ 31.01万
• 依托单位: UNIVERSITY OF TEXAS MED BR GALVESTON
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-07-24 至 2014-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8415381
• 关键词: A549; Absorbable Gelatin Sponge; Acute; Alveolar; Amniotic Fluid; Animal Model; Apoptosis; Biochemical; Biological; Biological Assay; Biology; Bioreactors; Cell Adhesion; Cell Communication; Cell Culture Techniques; Cell Death; Cell Differentiation process; Cell Line; Cell Proliferation; Cell Survival; Cell model; Cells; Collagen Type IV; Complex; Confocal Microscopy; Culture Media; Dendritic Cells; Dependency; Detection; Development; Disease; Elderly; Embryo; Endothelial Cells; Endothelium; Environment; Enzyme-Linked Immunosorbent Assay; Epithelial Cells; Epithelium; Evaluation; Exposure to; Fetal Lung; Fibroblasts; Fibrosis; Flow Cytometry; Functional disorder; Human; Human Cell Line; Human Engineering; Image; Imaging Techniques; Immune; Immune response; Immunoprecipitation; In Vitro; Infection; Inflammatory; Influenza; Influenza A Virus, H1N1 Subtype; Influenza A Virus, H5N1 Subtype; Injury; Label; Laboratories; Lead; Leukocytes; Life; Lung; Lung diseases; Measurement; Methodology; Methods; Microscopy; Modeling; Monitor; Morphology; Mus; Myofibroblast; Neuraminidase inhibitor; Organ; Organ Model; Oseltamivir; Pathogenesis; Pathogenicity; Pathway interactions; Phenotype; Physiology; Population; Process; Production; Pulmonary Fibrosis; Quantum Dots; Rattus; Reliance; Research Personnel; Respiratory physiology; Role; Serial Passage; Somatic Cell; Source; Stem cells; Structure of parenchyma of lung; System; Technology; Testing; Tissue Engineering; Tissue Model; Tissues; Toxic effect; Toxicology; Trachea; Umbilical vein; Validation; Work; Wound Healing; Xenobiotics; alveolar type II cell; base; cell injury; cell motility; cell type; complex biological systems; cytokine; drug discovery; drug testing; efficacy testing; embryonic stem cell; established cell line; human embryonic stem cell; human stem cells; human tissue; immortalized cell; improved; in vivo; lung development; lung injury; macrophage; matrigel; microbial; monolayer; progenitor; research study; response; scaffold; stem; three-dimensional modeling; two-dimensional; two-photon; whole body imaging

DESCRIPTION (provided by applicant): Current toxicity or pathogenesis studies rely heavily on traditional cell culture methods and animal models. While these systems provide much basic information, a human 3D organ model composed of native cell types interacting in a physiologically relevant way would more closely approximate in vivo conditions. Development of improved in vitro organ models would enable researchers to construct and analyze complex biological systems especially as they relate to organ development, disease pathogenesis, and toxicology and drug discovery. The development of such a model requires identifying the appropriate matrix material, cell types and microenvironment. Our hypothesis is that human embryonic stem or progenitor cells can be cultured on scaffolds to create an in vitro human lung model. It is possible to introduce leukocytes to the system to further mimic the in vivo environment and to study the roles of fibroblasts/myofibroblasts and macrophages/dendritic cells in the development of pulmonary fibrosis. The central hypothesis of this project is that human stem or progenitor cells can be on natural acellular lung or other commercially available scaffolds to create an in vitro human lung model which can be used to study lung disease and the development of fibrosis after microbial infection or exposure to xenobiotic agents. In this project, our aims are to (1) Produce a long term sustainable three dimensional (3D) human pulmonary model and (2) Test the validity of the model using exposure to H5N1, known to cause development of acute conditions and fibrosis in the lung or H1N1 2009 suspected of causing similar but less severe responses. Sham exposure or exposure to circulating strains of influenza A that do not elicit these responses will be used as controls as will addition of neuraminidase inhibitors Tamiflu and Amantidine. In order to produce the best lung model various matrices such as Gelfoam, Matrigel and collagen-IV will be examined for the ability to allow cell adhesion and sustainability. Several matrix materials will be considered, however, acellular (AC) human lung may be the best candidate for a fibrosis model. Recent work in our laboratory has shown this matrix is able to support differentiation of mouse embryonic stem cells toward cells comprising lung tissue with an appropriate morphology. Both AC whole rat trachea-lung and small sections of AC human lung will be tested for efficacy. For a human model, cell sources to be considered include cell lines and primary cells. Among the human cell lines to be trialed are A549 cells (Type II pneumocytes), HUV-EC-C or HUVEC-CS (human umbilical vein endothelial cells developed into a cell line), 13Lu (fetal lung cells) and human embryonic stem cells. Primary and progenitor cells such as HUVEC (primary cells), human amniotic fluid cells, and somatic lung progenitor cells may be used in a supporting role or to populate the matrix. The growth medium will be assessed for efficient proliferation and differentiation of cells into lung tissue. PUBLIC HEALTH RELEVANCE: We intend to develop engineered human lung tissue to use as a model. The model will be grown using human stem cells, human derived cell lines, or primary cells seeded onto small pieces of scaffold which will support three dimensional tissue development. These tissue pieces will have many of the types of cells found in the lung and will be used to study lung development, microbial lung infections and diseases such as fibrosis which sometimes results from lung infections.

描述(由申请人提供)：目前的毒性或发病机制研究严重依赖于传统的细胞培养方法和动物模型。虽然这些系统提供了许多基本信息，但由自然细胞类型以生理相关的方式相互作用组成的人体3D器官模型将更接近体内的条件。改进的体外器官模型的发展将使研究人员能够构建和分析复杂的生物系统，特别是当它们与器官发育、疾病发病机制、毒理学和药物发现有关时。这种模型的开发需要确定合适的基质材料、细胞类型和微环境。我们的假设是，人类胚胎干细胞或祖细胞可以在支架上培养，以创建一个体外人类肺模型。有可能将白细胞引入该系统，以进一步模拟体内环境，并研究成纤维细胞/肌成纤维细胞和巨噬细胞/树突状细胞在肺纤维化发展中的作用。该项目的中心假设是，人类干细胞或祖细胞可以在天然无细胞肺或其他商业可获得的支架上建立体外人类肺模型，可用于研究微生物感染或暴露于异物后肺部疾病和纤维化的发展。在这个项目中，我们的目标是(1)建立一个长期可持续的三维(3D)人体肺部模型，以及(2)使用H5N1病毒暴露来测试模型的有效性。H5N1病毒已知会导致肺部急性疾病和纤维化的发展，或者2009年H1N1流感疑似引起类似但不那么严重的反应。假暴露或暴露于不会引起这些反应的甲型流感流行毒株将被用作对照，神经氨酸酶抑制剂达菲和阿曼替丁的加入也是如此。为了制作最好的肺模型，将检查各种基质，如明胶泡沫、Matrigel和IV型胶原，以确定其允许细胞黏附和可持续性的能力。几种基质材料将被考虑，然而，无细胞(AC)人肺可能是纤维化模型的最佳候选者。我们实验室最近的工作表明，这种基质能够支持小鼠胚胎干细胞向含有适当形态的肺组织的细胞分化。将对AC大鼠全肺和AC人肺的小切片进行有效性测试。对于人体模型，要考虑的细胞来源包括细胞系和原代细胞。将被试验的人类细胞系包括A549细胞(II型肺细胞)、HuV-EC-C或HUVEC-CS(人脐静脉内皮细胞发展成细胞系)、13Lu(胎肺细胞)和人胚胎干细胞。原代和祖细胞，如HUVEC(原代细胞)、人羊水细胞和体细胞肺祖细胞可用于支持作用或填充基质。将对该生长介质进行有效的细胞增殖和分化为肺组织的评估。 公共卫生相关性：我们打算开发工程化的人类肺组织作为模型。该模型将使用人类干细胞、人类衍生细胞系或种植在支持三维组织发育的小块支架上的原代细胞进行培养。这些组织块将具有许多在肺中发现的细胞类型，并将用于研究肺发育、微生物肺部感染以及有时由肺部感染引起的纤维化等疾病。