Directing Collective Epithelial Morphology in Space and Time Using a Light-Based Carving Tool

使用基于光的雕刻工具指导空间和时间上的集体上皮形态

• 批准号: 9809041
• 负责人: Daniel A Harrington
• 金额: $ 15.4万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-07-01 至 2021-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9809041
• 关键词: 3-Dimensional; Ablation; Acinus organ component; Acute; Affect; Aging; Architecture; Autoimmune Responses; Basement membrane; Biological; Biomanufacturing; Caliber; Cell Culture Techniques; Cell Proliferation; Cell Size; Cell Survival; Cells; Cellular Morphology; Cellular Spheroids; Cessation of life; Clinical; Coculture Techniques; Complex; Connective Tissue; Crosslinker; Cues; Dental Care; Dental caries; Deposition; Development; Disease; Duct (organ) structure; Ductal; Elements; Encapsulated; Engineering; Epithelial; Epitopes; Etiology; Extracellular Matrix; Face; Feasibility Studies; Fibroblasts; Financial Hardship; Future; Gel; Gland; Goals; Grant; Growth; Growth Factor; Harvest; Head and Neck Cancer; Human; Hyaluronic Acid; Hydrogels; In Vitro; Integrins; Laboratories; Lasers; Length; Ligands; Light; Major salivary gland structure; Maps; Matrix Metalloproteinases; Membrane Proteins; Mesenchymal; Methods; Microscopy; Morphology; Myoepithelial; Neurons; Neurotransmitter Receptor; Neurotransmitters; Oral health; Organ; Pain; Palliative Care; Pathway interactions; Patients; Pattern; Peptide Signal Sequences; Phenotype; Physics; Pilot Projects; Polymers; Porosity; Positioning Attribute; Printing; Proliferating; Quality of life; Radial; Radiation induced damage; Radiation therapy; Regenerative Medicine; Resolution; Salivary; Salivary Gland Tissue; Salivary Glands; Shapes; Signal Transduction; Sjogren' s Syndrome; Source; Stem cells; Structure; Study models; Syringes; System; Technology; Testing; Therapeutic; Time; Tissue Engineering; Tissues; Travel; Variant; Work; Xerostomia; absorption; base; biomaterial compatibility; bioprinting; cell growth; cell motility; cell type; cost; crosslink; implantable device; implantation; improved; lithography; migration; monolayer; new technology; novel; progenitor; prototype; reconstitution; reduce symptoms; regenerative therapy; response; salivary acinar cell; salivary assay; salivary cell; standard of care; stem; temporal measurement; three dimensional cell culture; time use; tool; two-photon

Project Summary: Xerostomia, or “dry mouth”, is a challenging clinical condition, caused by damage to the cells of the salivary gland. It may result from a variety of tissue insults, including acute damage from radiation therapy for head and neck cancers, progressive auto-immune response in Sjogren’s disease, or other unknown etiology from aging. Current treatments offer only temporary relief of symptoms, and poor resolution of associated oral health decay. The cost of this condition is considerable, both in quality of life and the financial burden of increased dental care. The fields of tissue engineering and regenerative medicine offer many tools for the potential reconstitution of healthy salivary-derived cells within supportive hydrogel matrices, but few of these options provide sufficient spatial and temporal resolution to restore the complex branched structure and precise spatial phenotype map of the major salivary glands. However, new discoveries in laser-based hydrogel degradation (LBHD) can be used to “carve” pathways through intact hydrogel slabs, with pinpoint, subcellular resolution in xyz, and offer a method to guide a growing salivary epithelial bud in 3 dimensions. Our hypothesis for the present proposal is that we can use multiphoton-based LBHD to elongate a multicellular cluster in a given direction, and recreate key elements of the native gland. To do this, we will employ our laboratory’s expertise in isolation of primary human salivary-derived stem/progenitor cells (hS/PCs) from healthy tissues, and encapsulation as responsive 3D multicellular spheroid clusters within customizable, biocompatible hyaluronic acid (HA) hydrogels. Our ongoing work has shown that, by tailoring the porosity of these hydrogels and their concentration of bioactive epitopes, we can impact cluster size, morphology, and interaction with the surrounding extracellular matrix. We will test our hypothesis through the following Specific Aims: Aim 1. Establish parameters to carve “tunnels” through HA hydrogels and promote HS/PC cluster ingrowth. Aim 2. Adapt the system to alternate matrices that enable fibroblast co-culture, or incorporate photolabile crosslinkers for easier fabrication. Aim 3. Assess phenotype of the growing cluster, at its trailing and leading edges and branched termini, for signs of differentiated phenotype. If successful, this system could serve as a useful model for studying mechanisms of human salivary cell organization and differentiation; the system might also be an early prototype for manufacturing tissue engineered gland replacements. The R03 mechanism will provide support for the necessary pilot and feasibility studies, to demonstrate that these proven technologies can be combined to produce a novel platform.

项目概要： 口干症或“口干”是一种具有挑战性的临床病症，由唾液细胞损伤引起 腺。它可能是由多种组织损伤引起的，包括头部和头部放射治疗造成的急性损伤。 颈部癌症、干燥病的进行性自身免疫反应或其他未知的衰老病因。 目前的治疗只能暂时缓解症状，并且无法解决相关的口腔健康问题 衰变。这种情况的代价是相当大的，无论是在生活质量方面还是在增加的经济负担方面 牙科护理。组织工程和再生医学领域为发挥潜力提供了许多工具 在支持性水凝胶基质中重建健康的唾液来源的细胞，但这些选择很少 提供足够的空间和时间分辨率来恢复复杂的分支结构和精确的空间 主要唾液腺的表型图。然而，基于激光的水凝胶降解的新发现 （LBHD）可用于通过完整的水凝胶板“雕刻”路径，并具有精确的亚细胞分辨率 xyz，并提供了一种在 3 个维度上引导唾液上皮芽生长的方法。我们的假设为 目前的建议是，我们可以使用基于多光子的 LBHD 来延长给定的多细胞簇 方向，并重建原生腺体的关键元素。为此，我们将利用我们实验室的专业知识 从健康组织中分离原代人唾液干细胞/祖细胞 (hS/PC)，以及 封装为可定制、生物相容性透明质酸中的响应性 3D 多细胞球体簇 酸（HA）水凝胶。我们正在进行的工作表明，通过定制这些水凝胶的孔隙率及其 生物活性表位的浓度，我们可以影响簇的大小、形态以及与 周围的细胞外基质。我们将通过以下具体目标来检验我们的假设： 目标 1. 建立参数以通过 HA 水凝胶雕刻“隧道”并促进 HS/PC 簇向内生长。 目标 2. 使系统适应替代基质，以实现成纤维细胞共培养，或纳入不稳定性光 交联剂，更容易制造。目标 3. 评估生长簇的表型，包括尾随和前导 边缘和分支末端，用于分化表型的迹象。如果成功的话，该系统可以作为 研究人类唾液细胞组织和分化机制的有用模型；系统可能会 也是制造组织工程腺体替代品的早期原型。 R03机制将 为必要的试点和可行性研究提供支持，以证明这些经过验证的技术 可以组合起来产生一个新颖的平台。