虽然3D细胞模型对研究生物过程很有用,凝胶包埋的类器官具有很大的变异性。本文介绍了大型(〜1mm直径)的高产生产,无脚手架,使用MCF10A细胞的一滴一类器官格式的高度球形类器官,一种非致瘤性乳腺细胞系.这些类器官显示出中空管腔和次生腺泡,并表达乳腺特异性和祖细胞标记,类似于正常人的乳房腺泡。当接受TGF-β治疗时,缺氧模拟试剂CoCl2,或与间充质干细胞/基质细胞(MSC)共培养,类器官增加胶原蛋白I的产生,并经历肿瘤进展的大的表型和形态变化,它们是可重复和可量化的。这种无脚手架的优点,三维乳房类器官模型具有较高的一致性和可重复性,能够测量细胞胶原蛋白I的产生,而没有外源性胶原蛋白的噪音,以及使类器官经受来自微环境和外源治疗的各种刺激的能力,并且具有精确的时间,而无需考虑基质结合。使用这个系统,我们从MMTV-Cre的原发性化生乳腺癌中产生了类器官;Ccn6fl/fl小鼠,保留了原发肿瘤的高级梭形细胞形态。该平台可用作标准化的3D细胞模型,用于研究微环境因素如何影响乳腺肿瘤发生。以及潜在的治疗方法。
While 3D cellular models are useful to
study biological processes, gel-embedded organoids have large variability. This paper describes high-yield production of large (~1 mm diameter), scaffold-free, highly-spherical organoids in a one drop-one organoid format using MCF10A cells, a non-tumorigenic breast cell line. These organoids display a hollow lumen and secondary acini, and express mammary gland-specific and progenitor markers, resembling normal human breast acini. When subjected to treatment with TGF-β, the hypoxia-mimetic reagent CoCl2, or co-culture with mesenchymal stem/stromal cells (MSC), the organoids increase collagen I production and undergo large phenotypic and morphological changes of neoplastic progression, which were reproducible and quantifiable. Advantages of this scaffold-free, 3D breast organoid model include high consistency and reproducibility, ability to measure cellular collagen I production without noise from exogenous collagen, and capacity to subject the organoid to various stimuli from the microenvironment and exogenous treatments with precise timing without concern of matrix binding. Using this system, we generated organoids from primary metaplastic mammary carcinomas of MMTV-Cre;Ccn6fl/fl mice, which retained the high grade spindle cell morphology of the primary tumors. The platform is envisioned to be useful as a standardized 3D cellular model to
study how microenvironmental factors influence breast tumorigenesis, and to potential therapeutics.