Micropatterned Cell-Cell Interactions Enable Functional Encapsulation of Primary Hepatocytes in Hydrogel Microtissues
Micropatterned Cell-Cell Interactions Enable Functional Encapsulation of Primary Hepatocytes in Hydrogel Microtissues
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DOI：
10.1089/ten.tea.2013.0667
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发表时间：
2014-08-01
影响因子：
4.1
通讯作者：
Bhatia, Sangeeta N.
中科院分区：
文献类型：
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作者：
Li, Cheri Y.;Stevens, Kelly R.;Bhatia, Sangeeta N.
药物性肝损伤是药物开发失败和上市后退出的主要原因。结合原代肝细胞的体外模型已被证明比依赖肝微粒体或肝细胞癌细胞系的模型系统更具预测性。在体外稳定原代肝细胞表型的方法通常依赖于模拟肝脏微环境线索，如细胞-细胞相互作用和细胞-基质相互作用。在这项工作中，我们试图将表型稳定的肝细胞整合到三维（3D）显微组织中，这反过来又可以部署在药物筛选平台中，如多孔板和各种器官芯片设备。我们首先利用胶原蛋白I的微图图化来确定二维的细胞-细胞相互作用，然后通过胶原酶消化产生良好控制的聚集体，用于聚乙二醇（PEG）二丙烯酸酯的3D封装。使用这种方法，我们检测了同型肝细胞相互作用和包封水凝胶组成的影响，并实现了肝脏特异性功能维持超过50天。随后使用微流体液滴发生器封装最佳预聚集结构以产生三维微组织。通过流式细胞术表征了工程肝微组织与药物的相互作用，并诱导P450酶响应原型小分子和引起肝毒性的药物-药物相互作用。总的来说，这项研究建立了一个制造3D肝脏微组织的管道，这些组织具有稳定的肝脏特异性功能，可以被纳入广泛的新兴药物开发平台。
Drug-induced liver injury is a major cause of drug development failures and postmarket withdrawals. In vitro models that incorporate primary hepatocytes have been shown to be more predictive than model systems which rely on liver microsomes or hepatocellular carcinoma cell lines. Methods to phenotypically stabilize primary hepatocytes ex vivo often rely on mimicry of hepatic microenvironmental cues such as cell-cell interactions and cell-matrix interactions. In this work, we sought to incorporate phenotypically stable hepatocytes into three-dimensional (3D) microtissues, which, in turn, could be deployed in drug-screening platforms such as multiwell plates and diverse organ-on-a-chip devices. We first utilize micropatterning on collagen I to specify cell-cell interactions in two-dimensions, followed by collagenase digestion to produce well-controlled aggregates for 3D encapsulation in polyethylene glycol (PEG) diacrylate. Using this approach, we examined the influence of homotypic hepatocyte interactions and composition of the encapsulating hydrogel, and achieved the maintenance of liver-specific function for over 50 days. Optimally preaggregated structures were subsequently encapsulated using a microfluidic droplet-generator to produce 3D microtissues. Interactions of engineered hepatic microtissues with drugs was characterized by flow cytometry, and yielded both induction of P450 enzymes in response to prototypic small molecules and drug-drug interactions that give rise to hepatotoxicity. Collectively, this study establishes a pipeline for the manufacturing of 3D hepatic microtissues that exhibit stabilized liver-specific functions and can be incorporated into a wide array of emerging drug development platforms.