Engineering a biomimetic matrix to promote development of human ovarian follicles in vitro

工程仿生基质促进人类卵泡体外发育

• 批准号: 10311304
• 负责人: Andrea Jones
• 金额: $ 4.05万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-19 至 2024-08-18
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10311304
• 关键词: Age; Angiopoietins; Apoptotic; Atlases; Autologous Transplantation; Automobile Driving; Basement membrane; Bioinformatics; Biological; Biomimetics; Caliber; Cancer Survivor; Cell Proliferation; Cells; Characteristics; Chemistry; Clinic; Clinical; Complex; Computational Biology; Cortex of ovary; Coupled; Cryopreservation; Data Set; Deposition; Development; Dissociation; Embryo; Endocrine; Engineering; Environment; Extracellular Matrix; Extracellular Matrix Proteins; Fertility; Fertilization; Flow Cytometry; Fluorescence; Future; Gene Expression Profile; Gene Expression Profiling; Germ Cells; Goals; Growth; Growth Factor; Hormonal; Hormone Responsive; Hormones; Human; Hydrogels; In Vitro; Individual; Infertility; Knowledge; Life; Ligand Binding; Malignant Neoplasms; Mechanics; Methods; Molecular Biology; Mus; Nature; Oocytes; Ovarian; Ovarian Follicle; Ovarian Tissue; Ovarian tissue cryopreservation; Ovary; PGF gene; Paracrine Communication; Pathway interactions; Patients; Peptide Hydrolases; Peptides; Platelet-Derived Growth Factor; Population; Pregnancy; Primordial Follicle; Procedures; Protocols documentation; Reproducibility; Research; Risk; Role; Savings; Signal Transduction; Somatic Cell; Sorting - Cell Movement; Stains; Standardization; Stimulus; Stromal Cells; Structure; Supplementation; Support System; Suspensions; System; Tissue Engineering; Translating; Translations; Vascular Endothelial Growth Factors; Woman; Work; aged; angiogenesis; anticancer treatment; autocrine; base; cancer cell; cancer recurrence; cancer risk; cancer therapy; comparative; cytokine; cytotoxic; design; egg; ethylene glycol; fertility improvement; fertility preservation; folliculogenesis; girls; human tissue; improved; in vivo; innovation; next generation sequencing; novel; oocyte cryopreservation; oocyte retrieval; paracrine; prepuberty; primary ovarian insufficiency; prospective; reproductive; scaffold; self assembly; single cell sequencing; single-cell RNA sequencing; standard of care; success; three dimensional structure; tissue culture; transcription factor; transcriptome; transcriptomics

The long-term goal of the proposed research is to establish a broad fertility preservation option for women undergoing gonadotoxic anticancer treatments and facing infertility. The overall objective of this proposal in working towards the presented goal and mitigating the risks associated with autotransplantation is to create a biomimetic environment that promotes human follicle development from the primordial stage in vitro. The low success rates of small follicle culture are largely attributed to the complex and poorly understood paracrine, autocrine and endocrine signaling between follicular cells, neighboring follicles, and stromal cells. The central hypothesis is that transcriptional profiling of human follicles will reveal mechanisms driving development and recreating the ovarian microenvironment through design of a biomimetic hydrogel which retains cell-secreted extracellular matrix (ECM) will support human follicle development in vitro. The rationale for the proposed work is that by deciphering the mechanisms driving follicle activation and early development, and recapitulating the natural ovarian microenvironment in an ECM-sequestering hydrogel, future culture systems can be translated to the clinic for maturation of cryopreserved ovarian follicles and subsequent fertilization and pregnancy. In the first aim, single cell RNA sequencing will be used to profile human ovarian follicles and supportive stromal cells. In the second aim, ECM-sequestering peptides will be incorporated in a biomimetic poly (ethylene glycol) (PEG) hydrogel system using Michael-type addition chemistry to promote deposition of ECM components and mimic the native ovarian tissue. The follicle’s basement membrane is composed of ECM proteins and it functions as structural support for follicular cells, a selective barrier for molecules entering the follicle, and a scaffold for retaining soluble growth factors and cytokines. It is continuously remodeled during follicle development, but cell- secreted ECM molecules are unable to adhere to unmodified PEG for self-assembly. By integrating ECM- sequestering peptides in the PEG hydrogels, the structural and biological roles of ECM can be restored for in vitro follicle development. The contribution of this work will be a single cell atlas of the reproductive-age ovary highlighting mechanisms driving follicle development and stromal cells’ supportive roles in folliculogenesis and a novel in vitro follicle culture system that supports human follicle development. The contribution of this work will be significant because it will guide the development of a standardized in vitro culture for maturation of human follicles and a safe fertility preservation option for patients unable to produce mature eggs as a result of gonadotoxic treatments. The proposed work is innovative in that it will be the first single cell dataset from healthy reproductive aged women and the first instance of human follicle culture in a synthetic ECM-sequestering matrix.

拟议研究的长期目标是为接受性腺毒性抗癌治疗并面临不孕症的女性建立广泛的生育力保留选择。该提案致力于实现所提出的目标并减轻与自体移植相关的风险，其总体目标是创建一个仿生环境，促进人类卵泡从体外原始阶段发育。小卵泡培养的低成功率很大程度上归因于卵泡细胞、邻近卵泡和基质细胞之间复杂且知之甚少的旁分泌、自分泌和内分泌信号传导。中心假设是，人类卵泡的转录谱将揭示驱动发育的机制，并通过设计保留细胞分泌的细胞外基质（ECM）的仿生水凝胶来重建卵巢微环境，从而支持人类卵泡的体外发育。这项工作的基本原理是，通过破译驱动卵泡激活和早期发育的机制，并在 ECM 隔离水凝胶中重现自然卵巢微环境，未来的培养系统可以应用于临床，用于冷冻保存的卵泡的成熟以及随后的受精和妊娠。第一个目标是利用单细胞 RNA 测序来分析人类卵泡和支持性基质细胞。第二个目标是，利用迈克尔型加成化学将 ECM 螯合肽纳入仿生聚乙二醇 (PEG) 水凝胶系统中，以促进 ECM 成分的沉积并模拟天然卵巢组织。毛囊的基底膜由 ECM 蛋白组成，其功能是毛囊细胞的结构支撑、分子进入毛囊的选择性屏障以及保留可溶性生长因子和细胞因子的支架。它在卵泡发育过程中不断重塑，但细胞分泌的 ECM 分子无法粘附到未修饰的 PEG 上进行自组装。通过将 ECM 螯合肽整合到 PEG 水凝胶中，可以恢复 ECM 的结构和生物学作用，以促进体外卵泡发育。这项工作的贡献将是育龄卵巢的单细胞图谱，突出显示驱动卵泡发育的机制和基质细胞在卵泡发生中的支持作用，以及支持人类卵泡发育的新型体外卵泡培养系统。这项工作的贡献将是巨大的，因为它将指导人类卵泡成熟的标准化体外培养物的开发，并为因性腺毒性治疗而无法产生成熟卵子的患者提供安全的生育力保存选择。这项工作的创新之处在于，它将是第一个来自健康生育年龄女性的单细胞数据集，也是第一个在合成 ECM 隔离基质中进行人类卵泡培养的实例。