An ovary-on-a-chip for understanding early folliculogenesis and reproductive toxicology in a large mammalian model

用于了解大型哺乳动物模型中早期卵泡发生和生殖毒理学的卵巢芯片

• 批准号: 10267664
• 负责人: Jennifer Nagashima
• 金额: $ 4.06万
• 依托单位: SMITHSONIAN INSTITUTION
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-11-01 至 2021-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10267664
• 关键词: 3-Dimensional; Alginates; Anatomy; Apoptosis; Atomic Force Microscopy; Biocompatible Materials; Biological; Biological Assay; Biomimetics; Cancer Patient; Cells; Characteristics; Chemicals; Clinic; Clinical; Coculture Techniques; Communication; Complex; Cortex of ovary; Cyclophosphamide; Development; Ensure; Environment; Epigenetic Process; Ethics; Exposure to; Family Felidae; Felis catus; Female; Fertility; Future; Genes; Goals; Growing Follicle; Growth; Health; Hormonal; Household; Human; Hydrogels; In Situ; In Vitro; Incubated; Infertility; Laboratories; Maintenance; Mammals; Mentors; Metabolic; Methods; Microfluidic Microchips; Microfluidics; Modeling; Mus; Nuclear; Nutrient; Oocytes; Ovarian; Ovarian Follicle; Ovary; Pattern; Perfosfamide; Pharmaceutical Preparations; Primordial Follicle; Proteins; Proteome; Protocols documentation; Recovery; Reproduction; Reproductive Health; Research; Risk; Secure; Somatic Cell; Structure; Study models; System; Technology; Tissues; Toxicology; Toxin; Translating; chemotherapy; density; dynamic system; epigenetic marker; experience; female fertility; fertility improvement; fertility preservation; folliculogenesis; histone methylation; imprint; improved; in vitro testing; in vivo; innovation; interest; methylation pattern; microfluidic technology; mimicry; nonhuman primate; novel; offspring; organ on a chip; ovotoxicity; paracrine; preservation; reproductive; tandem mass spectrometry; toxicant; transcriptome; transcriptome sequencing; young woman

PROJECT SUMMARY/ABSTRACT Recapitulation of folliculogenesis in vitro is of interest for both preserving human fertility and assessing the influence of chemical contaminants on ovarian health. Although live offspring can be produced from oocytes recovered from murine follicles grown in vitro, the larger size and protracted duration of folliculogenesis of large mammals likely necessitates more complex culture requirements. The domestic cat will be used as a study model because this species shares a number of anatomical and reproductive characteristics including follicle size and oocyte nuclear configuration with the human. The proposed project will take advantage of ‘organ-on-chip’ microfluidic technology combined with somatic cell co-culture and my experience with 3-dimensional hydrogels to recapitulate, as possible, in vivo conditions in an “ovary-on-a-chip”. Specifically, I propose to quantify the rigidity of the domestic cat ovary via micropipette aspiration and atomic force microscopy and identify biomimetic densities of alginate hydrogel that provide optimal structural support to primordial follicles. I will determine the influence of rigidity and medium flow rate on in vitro folliculogenesis, and identify via tandem mass spectrometry and RNA sequencing technologies, proteins and genes involved in follicle activation in culture. Comparison of the dynamic system with traditional, static culture will allow me to improve understanding of the contributions of structural rigidity (hydrogel density), medium flow (microfluidic chip), and the paracrine environment (produced via co-culture) on follicle survival and activation in vitro (Aim 1). I then propose to evaluate the epigenetic impacts (methylation of histones, key imprinted genes) of long-term culture in this system compared with traditional culture, as well as determine the utility of the “ovary-on-a-chip” in ovarian toxicology studies, using the active form of the chemotherapeutic cyclophosphamide in proof-of-concept studies (Aim 2). Together, these studies advance understanding of contributions of the microenvironment on follicle development toward the goal of an in vitro system for enhancing fertility preservation and reproductive toxicology.

项目总结/摘要 体外卵泡发生的重演对于保持人类生育力和评估卵泡发育的能力都是有意义的。 化学污染物对卵巢健康的影响虽然卵母细胞可以产生活的后代 从体外培养的小鼠卵泡中回收，大的卵泡体积更大， 哺乳动物可能需要更复杂的培养要求。家猫将被用作研究模型 因为这一物种具有许多解剖学和生殖特征，包括卵泡大小， 卵母细胞核构型与人类相似。拟议项目将利用“器官芯片” 微流控技术结合体细胞共培养和我的三维水凝胶经验 以尽可能概括“卵巢芯片”中的体内条件。具体来说，我建议量化 通过微量吸管抽吸和原子力显微镜观察家猫卵巢的硬度并识别仿生 密度的藻酸盐水凝胶，为原始卵泡提供最佳的结构支持。我将决定 刚性和培养液流速对体外卵泡形成影响及串联质谱鉴定 和RNA测序技术，蛋白质和基因参与卵泡激活的文化。比较 动态系统与传统的静态文化将使我能够更好地理解 结构刚性（水凝胶密度）、介质流动（微流控芯片）和旁分泌环境（产生的 通过共培养）对体外卵泡存活和活化的影响（目的1）。然后，我建议评估表观遗传的影响， （组蛋白甲基化，关键印迹基因）的长期培养，在这个系统与传统的相比， 培养，以及确定在卵巢毒理学研究中的“卵巢芯片”的效用，使用活性 概念验证研究中化疗环磷酰胺的形式（目标2）。这些研究一起 促进对卵泡发育的微环境的贡献的理解， 用于增强生育力保存和生殖毒理学的体外系统。

项目成果

In vitro development of mechanically and enzymatically isolated cat ovarian follicles.

• DOI: 10.1530/raf-20-0067
• 发表时间: 2021
• 期刊: Reproduction & fertility
• 影响因子: 2.1
• 作者: Nagashima JB;Hill AM;Songsasen N
• 通讯作者: Songsasen N

Evaluation of an ovary-on-a-chip in large mammalian models: Species specificity and influence of follicle isolation status.

大型哺乳动物模型中卵巢芯片的评估：物种特异性和卵泡分离状态的影响。

• DOI: 10.1002/term.2623
• 发表时间: 2018
• 期刊: Journal of tissue engineering and regenerative medicine
• 影响因子: 3.3
• 作者: Nagashima,JenniferB;ElAssal,Rami;Songsasen,Nucharin;Demirci,Utkan
• 通讯作者: Demirci,Utkan

Follicular extracellular vesicles enhance meiotic resumption of domestic cat vitrified oocytes

• DOI: 10.1038/s41598-020-65497-w
• 发表时间: 2020-05-25
• 期刊: SCIENTIFIC REPORTS
• 影响因子: 4.6
• 作者: Ferraz, Marcia de Almeida Monteiro Melo;Fujihara, Mayako;Songsasen, Nucharin
• 通讯作者: Songsasen, Nucharin

Comparative Tensile Properties and Collagen Patterns in Domestic Cat (Felis catus) and Dog (Canis lupus familiaris) Ovarian Cortical Tissues.

• DOI: 10.3390/bioengineering10111285
• 发表时间: 2023-11-04
• 期刊: BIOENGINEERING-BASEL
• 影响因子: 4.6
• 作者: Nagashima, Jennifer B.;Zenilman, Shoshana;Raab, April;Aranda-Espinoza, Helim;Songsasen, Nucharin
• 通讯作者: Songsasen, Nucharin