The contributions of inflammatory cocktail and stromal cell origin on a scaffold-free 3D biofabricated SSuPer tissue of endometriosis and normal endometrium

子宫内膜异位症是一种使人衰弱的疾病，影响高达10%的美国妇女。虽然有治疗方法，但 50%的子宫内膜异位症患者会有疾病复发。缺乏有用的模式继续阻碍 发现病理生理学的各个方面，这些方面适合于新的干预措施。因此，有一个 迫切需要开发新的子宫内膜异位症模型，以显着推进该领域。本申请 建议创建3D生物制造的SSuPer（自支撑灌注）组织， 子宫内膜异位症或正常子宫内膜使用Kenzan方法。Kenzan方法使用Regenova Bio 3D打印机将多细胞球体放置在微针阵列上，称为Kenzan。在Kenzan星球上， 融合通过细胞间的相互作用和分泌他们的天然细胞外基质，使更多的内在 细胞间通讯上皮细胞和间质细胞之间的细胞间通讯是至关重要的， 子宫内膜功能初步数据显示，Kenzan方法有效地生物制造了第一个 无支架的3D生物打印构建体，包括子宫内膜上皮和子宫内膜 基质细胞此外，SSuPer组织含有微通道，可以在生物惰性环境中灌注， FABRICA生物反应器。由于人子宫内膜间质成纤维细胞对子宫功能至关重要， 来自患有子宫内膜异位症的妇女的成纤维细胞可用于模拟异常的细胞功能、异常的 分子信号传导和促炎转录组学特征。此外，亲- 炎症介质在子宫内膜异位症患者的腹腔液中升高， 孕酮抵抗该应用程序的目的是生物制造子宫内膜异位症和子宫内膜异位症的3D模型。 正常子宫内膜，其再现炎症、转录组和类固醇激素反应 忠实地中心假设是炎性细胞因子和基质细胞来源（即，异位症 或不）独立影响每个SSuPer组织的子宫内膜异位或子宫内膜表型。超级组织， 包括来自子宫内膜异位症或正常子宫内膜的上皮细胞和基质细胞， 并用培养基或具有炎性混合物的培养基灌注。将确定SUPER组织为 无论是子宫内膜异位症或正常子宫内膜炎性标志物分泌，表达子宫内膜异位症 生物标志物和整体转录组学变化。二次验证研究将重点关注类固醇激素 反应在这项申请中提出的研究是创新的，因为它将产生新的无支架3D 与子宫内膜异位症或正常子宫内膜保持高度生物保真度的生物制造SSuper组织。 严格验证的体外模型将为研究肿瘤的病理生理学提供有用的模型。 子宫内膜异位症和其他子宫功能障碍疾病，如不孕症和反复流产。高度 可重复的模型是测试新疗法所必需的。

Endometriosis is a debilitating disease that affects up to 10% of U.S. women. While treatments exist, more than 50% of women with endometriosis will have disease recurrence. The lack of useful models continues to impede the discovery of aspects of pathophysiology that are amenable to novel interventions. Therefore, there is an urgent need to develop novel models of endometriosis to advance the field significantly. This application proposes to create 3D biofabricated SSuPer (Self-Supporting Perfused) tissues that recapitulate either endometriosis or normal endometrium using the Kenzan method. The Kenzan method uses the Regenova Bio 3D Printer to place multicellular spheroids onto a microneedle array, called a Kenzan. On Kenzan, the spheroids fuse through cell-cell interaction and secretion of their native extracellular matrix, allowing more intrinsic intercellular communication. Intercellular communication between epithelial and stromal cells is paramount in endometrial function. Preliminary data shows that the Kenzan method effectively biofabricates the first scaffold-free 3D bioprinted constructs comprised of both endometriotic epithelial and endometrial stromal cells. Further, SSuPer tissues contain microchannels that can be perfused in a biologically inert FABRICA bioreactor. Because human endometrial stromal fibroblasts are critical to uterine function, stromal fibroblasts from women with endometriosis can be used to model the abnormal cellular function, aberrant molecular signaling, and pro-inflammatory transcriptomic profiles that occur in endometriosis. Additionally, pro- inflammatory mediators are elevated in the peritoneal fluid of women with endometriosis and likely play a role in progesterone resistance. This application's objective is to biofabricate 3D models of both endometriosis and normal endometrium, which recapitulate the inflammatory, transcriptomic, and steroid hormone response faithfully. The central hypothesis is that both inflammatory cytokines and stromal cell origin (i.e., endometriosis or not) independently affect the endometriotic or endometrial phenotype of each SSuPer tissue. SSuPer tissues, comprised of epithelial and stromal cells from either endometriosis or normal endometrium, will be biofabricated and perfused with either media or media with an inflammatory cocktail. SSuPer tissues will be determined to be either endometriosis or normal endometrium by inflammatory marker secretion, expression of endometriosis biomarkers, and global transcriptomic changes. Secondary validation studies will focus on steroid hormone response. The research proposed in this application is innovative because it will generate novel scaffold-free 3D biofabricated SSuPer tissues that maintain high biological fidelity to endometriosis or normal endometrium. Rigorously validated in vitro models will provide useful models for investigating the pathophysiology of endometriosis and other uterine dysfunction diseases such as infertility and recurrent pregnancy loss. Highly reproducible models are necessary for testing novel therapies.