Anchorage-independent culture for maintaining vocal fold epithelial stem cells

用于维持声带上皮干细胞的非贴壁依赖性培养

• 批准号: 9903281
• 负责人: Xudong Shi
• 金额: $ 19.13万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-04-01 至 2022-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9903281
• 关键词: 3-Dimensional; Address; Adherent Culture; Affect; Biological Assay; Biology; Bioreactors; Caring; Cell Culture Techniques; Cell Differentiation process; Cell Line; Cell Separation; Cells; Cellular biology; Clinical; Collaborations; Communities; Culture-independent methods; Data; Development; Developmental Biology; Diagnosis; Disease; Dysphonia; Environment; Epithelial; Epithelial Cells; Epithelium; Fibroblasts; Functional disorder; Generations; Goals; Growth; Human; Human Cell Line; Immunologics; Impairment; In Vitro; Ion Transport; Larynx; Maintenance; Medicine; Methodology; Methods; Mucous Membrane; Natural regeneration; Patients; Performance; Phenotype; Physiological; Population; Quality of life; Regenerative Medicine; Reporting; Reproducibility; Research; Resources; Source; Standardization; System; Techniques; Testing; Tissue Engineering; Tissues; United States; Voice; Water; Work; bone; epithelial stem cell; experimental study; in vivo; novel; pluripotency; pre-clinical; regenerative; self-renewal; stem cell biology; stem cells; stemness; tool; vibration; vocal cord

Project Summary Voice impairment (dysphonia) affects an estimated 20 million people in the United States and, despite advances in diagnosis and treatment, remains a challenging clinical problem with a negative impact on patient quality-of-life. Advances in the emerging field of VF mucosal biology have resulted in a number of powerful and widely used in vitro experimental tools, such as human VF fibroblast cell lines and vibrational bioreactors; however, there remain limited cell biology resources specific to the VF epithelium and its constituent cells, hampering advances in VF epithelial biology and the understanding of epithelial diseases. A central reason for this lack of progress is the technical challenges associated with VF epithelial cell isolation, maintenance, and expansion in vitro. A reliable and reproducible culture methodology is needed to overcome these challenges. In the proposed work, we will introduce and validate a promising in vitro technique – anchorage-independent culture – for the isolation and maintenance of VF epithelial stem cells. Our team’s prior work in other epithelial systems, and pilot data generated using VF epithelial cells, demonstrate that this technique results in the formation of three-dimensional culture spheres that are enriched for tissue- specific stem cells. In the proposed research, we will characterize the makeup, growth, maintenance, and passage of VF spheres, as well as test the capacity of sphere-contained stem cells for in vitro differentiation and as a cell source for VF tissue engineering. This work, conducted using human cells, will result in a validated and reproducible in vitro methodology that will advance progress in VF epithelial (stem) cell biology and facilitate ongoing and future research amongst the wider scientific community. This research effort represents a powerful collaboration between experts in VF mucosal biology and stem cell biology and has broad applicability to both basic scientific research and regenerative medicine of the larynx.

项目概要 据估计，美国有 2000 万人受到声音障碍（发声困难）的影响，尽管 诊断和治疗的进步，仍然是一个具有挑战性的临床问题，对患者产生负面影响 生活质量。 VF 粘膜生物学新兴领域的进展已经产生了许多强大且 广泛使用的体外实验工具，如人VF成纤维细胞系和振动生物反应器； 然而，针对 VF 上皮及其组成细胞的细胞生物学资源仍然有限， 阻碍了室颤上皮生物学的进展和对上皮疾病的理解。一个核心原因是 这种缺乏进展是与 VF 上皮细胞分离、维护和治疗相关的技术挑战。 体外扩增。需要可靠且可重复的培养方法来克服这些挑战。在 在拟议的工作中，我们将介绍并验证一种有前途的体外技术——锚定无关 培养——用于分离和维持 VF 上皮干细胞。我们团队之前在其他上皮细胞方面的工作 系统和使用 VF 上皮细胞生成的试验数据表明，该技术导致 形成富含组织特异性干细胞的三维培养球。在提议的 研究中，我们将表征 VF 球体的构成、生长、维护和通过，以及测试 球体干细胞体外分化的能力以及作为 VF 组织细胞来源的能力 工程。这项工作使用人体细胞进行，将产生经过验证且可重复的体外 该方法将推动 VF 上皮（干）细胞生物学的进展，并促进当前和未来的发展 更广泛的科学界的研究。这项研究工作代表了强有力的合作 VF 粘膜生物学和干细胞生物学专家之间的合作，对这两个基础领域都有广泛的适用性 喉科学研究和再生医学。