This paper describes the manufacture of geometrically inverted mammary organoids encapsulating primary mammary preadipocytes and adipocytes. Material manipulation in an array of 192 hanging drops induces cells to self-assemble into inside-out organoids where an adipose tissue core is enveloped by a cell-produced basement membrane, indicated by laminin V staining and then a continuous layer of mammary epithelial cells. This inverted tissue structure enables investigation of multiple mammary cancer subtypes, with a significantly higher extent of invasion by triple-negative MDA-MB-231 breast cancer cells compared to MCF7 cells. By seeding cancer cells into co-culture around pre-formed organoids with encapsulated preadipocytes/adipocytes, invasion through the epithelium, then into the adipose core is observable through acquisition of confocal image stacks of whole mount specimens. Furthermore, in regions of the connective tissue core where invasion occurs, there is an accumulation of collagen in the microenvironment. Suggesting that this collagen may be conducive to increased invasiveness, the anti-fibrotic drug pirfenidone shows efficacy in this model by slowing invasion. Comparison of adipose tissue derived from three different donors shows method consistency as well as the potential to evaluate donor cell-based biological variability. Insight box Geometrically inverted mammary organoids encapsulating primary preadipocytes/adipocytes (P/As) are bioengineered using a minimal amount of Matrigel scaffolding. Use of this eversion-free method is key to production of adipose mammary organoids (AMOs) where not only the epithelial polarity but also the entire self-organizing arrangement, including adipose position, is inside-out. While an epithelial-only structure can analyze cancer cell invasion, P/As are required for invasion-associated collagen deposition and efficacy of pirfenidone to counteract collagen deposition and associated invasion. The methods described strike a balance between repeatability and preservation of biological variability: AMOs form consistently across multiple adipose cell donors while revealing cancer cell invasion differences.

中文翻译：

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三阴性乳腺癌细胞侵入脂肪细胞/前脂肪细胞包裹的几何倒置的乳腺类器官。

本文介绍了封装原发性乳腺前脂肪细胞和脂肪细胞的几何倒置乳腺类器官的制造。192 个悬滴阵列中的材料操作诱导细胞自组装成由内而外的类器官，其中脂肪组织核心被细胞产生的基底膜包裹，层粘连蛋白 V 染色表明，然后是连续的乳腺上皮细胞层。这种倒置的组织结构能够研究多种乳腺癌亚型，与 MCF7 细胞相比，三阴性 MDA-MB-231 乳腺癌细胞的侵袭程度明显更高。通过将癌细胞播种到预先形成的类器官周围与封装的前脂肪细胞/脂肪细胞共培养，通过上皮细胞侵入，然后通过获取整个安装标本的共焦图像堆栈可以观察到脂肪核心。此外，在发生入侵的结缔组织核心区域，微环境中存在胶原蛋白的积累。表明这种胶原蛋白可能有助于增加侵袭性，抗纤维化药物吡非尼酮通过减缓侵袭在该模型中显示出疗效。来自三个不同供体的脂肪组织的比较显示了方法的一致性以及评估基于供体细胞的生物变异性的潜力。Insight box 封装初级前脂肪细胞/脂肪细胞 (P/As) 的几何倒置乳腺类器官是使用最少量的 Matrigel 脚手架进行生物工程改造的。使用这种无外翻方法是生产脂肪乳腺类器官 (AMO) 的关键，其中不仅上皮极性而且整个自组织排列（包括脂肪位置）都是由内而外的。虽然仅上皮结构可以分析癌细胞侵袭，但与侵袭相关的胶原沉积和吡非尼酮抵消胶原沉积和相关侵袭的功效需要 P/As。所描述的方法在可重复性和保持生物变异性之间取得平衡：AMO 在多个脂肪细胞供体中一致形成，同时揭示癌细胞侵袭差异。P/A 是侵袭相关的胶原蛋白沉积所必需的，吡非尼酮抵消胶原蛋白沉积和相关侵袭的功效也是必需的。所描述的方法在可重复性和保持生物变异性之间取得平衡：AMO 在多个脂肪细胞供体中一致形成，同时揭示癌细胞侵袭差异。P/A 是侵袭相关的胶原蛋白沉积所必需的，吡非尼酮抵消胶原蛋白沉积和相关侵袭的功效也是必需的。所描述的方法在可重复性和保持生物变异性之间取得平衡：AMO 在多个脂肪细胞供体中一致形成，同时揭示癌细胞侵袭差异。