Utilizing biodegradable porous silicon membranes as a novel design for lung-on-a-chip microfluidic devices to investigate extracellular matrix interactions.

利用可生物降解的多孔硅膜作为片上肺微流体装置的新颖设计来研究细胞外基质相互作用。

• 批准号: 10400222
• 负责人: David James Blake
• 金额: $ 9.75万
• 依托单位: FORT LEWIS COLLEGE
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-05-01 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10400222
• 关键词: Address; Alveolar; Anatomy; Animal Model; Animals; Apoptosis; Architecture; Atomic Force Microscopy; Basic Science; Biochemical; Biocompatible Materials; Biodegradation; Biological Models; Biomanufacturing; Biomedical Engineering; Cell Culture Techniques; Cells; Characteristics; Collagen; Complex; Crete; Cues; Cultured Cells; Data; Development; Devices; Disease; Disease Progression; Elastin; Endothelial Cells; Epithelial Cells; Extracellular Matrix; Extracellular Matrix Proteins; Extravasation; Flow Cytometry; Functional disorder; Future; Goals; Growth Factor; Human; Human Characteristics; Hypoxia; Immune; Immunofluorescence Microscopy; In Vitro; Inflammatory; Knowledge; Lead; Lipopolysaccharides; Lung; Lung diseases; Mediator of activation protein; Membrane; Microfluidic Microchips; Microfluidics; Modeling; Organ; Outcome; Pathogenesis; Pathway interactions; Peptide Hydrolases; Pharmacologic Substance; Physiology; Prevention; Production; Property; Public Health; Pulmonary Hypertension; Research; Reverse Transcriptase Polymerase Chain Reaction; Role; Safety; Scanning Electron Microscopy; Silicon; Structure; Surface; Techniques; Therapeutic; Thinness; Translating; Xenobiotics; base; biomaterial compatibility; body system; candidate identification; cell type; cigarette smoke; cytokine; design; drug candidate; extracellular; fibrotic lung disease; flexibility; human disease; human model; immunoregulation; in vitro Model; in vivo; innovation; insight; live cell imaging; macrophage; materials science; nanomaterials; novel; organ on a chip; pathogenic microbe; pre-clinical; prospective; success; three dimensional structure; toxicant; translational impact; translational therapeutics

Project Summary Identifying the cellular pathways that promote disease or which prospective therapies are effective relies upon appropriate mammalian models. Therefore, there is an urgent need for advanced human model systems that can accurately reproduce human anatomy and physiology to help predict human disease progression and as- sess potential treatment options. The long-term goal is to utilize a novel in vitro lung-on-a-chip (LOAC) microflu- idic device to predict how xenobiotics lead to inflammatory, fibrotic and immunomodulatory pulmonary diseases in humans. The overall objective is to create the first fully organic LOAC that is structurally supported by a cell- derived extracellular matrix (ECM) and includes innate immune cells to simulate organ-level functionality. The rationale for the proposed research is to employ the unique properties of porous silicon (PSi) not previously explored to revolutionize the field of material science in the fabrication of microfluidic platforms that incorporates dynamic ECM changes. Guided by strong preliminary data, the overall objective will be accomplished by pursing the following three specific aims: 1) Identify the optimal parameters and cellular mechanisms to dissolve ultrathin porous silicon during long-term culture; 2) Determine the extent to which co-cultured cells within the LOAC se- crete and create their own ECM; and 3) Develop a multicellular alveolar structure to activate immune cells leading to extravasation and ECM remodeling using an in vitro model of pulmonary hypertension. Under the first aim, the working hypothesis based on preliminary data is that human macrophages (MACs) are essential to modify and dissolve PSi. Dissolution rates of PSi will be quantified through scanning electron microscopy (SEM) and surface analysis will be completed by atomic force microscopy (AFM) to reproducibly create flexible, structurally intact membranes. Under the second aim, the working hypothesis is that endothelial cells (ECs) will express and secrete cell-derived ECM proteins. Secretion of de novo synthesized ECM components will be quantified through RT-PCR, confocal immunofluorescence (IF) microscopy and AFM. The third aim based on preliminary data in- dicate epithelial cells (EPCs), ECs and MACs can be successfully co-cultured and are viable during long-term culture on PSi membranes. The working hypothesis is in the presence of hypoxic conditions, MACs will become activated and release soluble mediators leading to apoptosis of ECs and increased ECM remodeling that will be quantified through confocal IF microscopy and live cell imaging. The proposed research is innovative, in our opinion, because it represents a substantive departure from the status quo by utilizing the unique characteristics of PSi, which is a biocompatible and biodegradable material. In addition, utilizing PSi provides the capability to create a cell specific ECM that will release biochemical cues including growth factors and extracellular protein- ases. The proposed research is significant because it is expected to have broad translational importance in the prevention and treatment of a wide range of pulmonary diseases. Finally, therapeutic advancements in the de- velopment of PSi nanomaterials and factors that lead to degradation in vivo are expected.

项目概要 确定促进疾病的细胞途径或哪些前瞻性疗法有效依赖于 合适的哺乳动物模型。因此，迫切需要先进的人体模型系统 可以准确地再现人体解剖学和生理学，以帮助预测人类疾病的进展和as- 评估潜在的治疗方案。长期目标是利用新型体外肺芯片 (LOAC) 微流控技术 idic 装置可预测异生物质如何导致炎症、纤维化和免疫调节性肺部疾病 在人类中。总体目标是创建第一个完全有机的 LOAC，其结构由细胞支撑 衍生的细胞外基质（ECM），并包括先天免疫细胞来模拟器官水平的功能。这 拟议研究的基本原理是利用多孔硅（PSi）以前没有的独特性能 探索在微流体平台的制造中彻底改变材料科学领域，该平台结合了 动态 ECM 变化。以强有力的初步数据为指导，通过追求实现总体目标 以下三个具体目标： 1) 确定溶解超薄物质的最佳参数和细胞机制 长期培养过程中的多孔硅； 2) 确定 LOAC 内共培养细胞的程度 具体并创建自己的 ECM； 3) 开发多细胞肺泡结构以激活免疫细胞 使用肺动脉高压体外模型进行外渗和 ECM 重塑。在第一个目标下， 基于初步数据的工作假设是人类巨噬细胞（MAC）对于改变 并溶解PSi。 PSi 的溶解速率将通过扫描电子显微镜 (SEM) 进行量化， 表面分析将通过原子力显微镜 (AFM) 完成，以可重复地创建灵活的结构 完整的膜。在第二个目标下，工作假设是内皮细胞（EC）将表达并 分泌细胞来源的 ECM 蛋白。从头合成的 ECM 成分的分泌将通过以下方式进行量化 RT-PCR、共焦免疫荧光 (IF) 显微镜和 AFM。第三个目标基于初步数据—— 表明上皮细胞 (EPC)、EC 和 MAC 可以成功共培养，并且可以长期存活 PSi 膜上的培养。工作假设是在缺氧条件下，MAC 会变成 激活并释放可溶性介质，导致 EC 凋亡并增加 ECM 重塑，这将 通过共焦 IF 显微镜和活细胞成像进行量化。拟议的研究具有创新性，在我们看来 意见，因为它代表着利用独特特征与现状的实质性背离 PSi 是一种生物相容性和可生物降解材料。此外，利用 PSi 还能够 创建细胞特异性 ECM，释放生化信号，包括生长因子和细胞外蛋白 - 情况。拟议的研究意义重大，因为预计它将在以下领域具有广泛的转化重要性： 预防和治疗多种肺部疾病。最后，治疗方面的进展 PSi纳米材料的发展以及导致体内降解的因素是值得期待的。