Distinct niches within the extracellular matrix dictate fibroblast function in (cell free) 3D lung tissue cultures
Distinct niches within the extracellular matrix dictate fibroblast function in (cell free) 3D lung tissue cultures
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DOI：
10.1152/ajplung.00408.2017
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发表时间：
2018-05-01
影响因子：
4.9
通讯作者：
Eickelberg, Oliver
中科院分区：
文献类型：
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作者：
Burgstaller, Gerald;Sengupta, Arunima;Eickelberg, Oliver
细胞外基质(ECM)的信号及其与细胞的功能相互作用在发育、组织修复和疾病中发挥着关键作用。然而，这种相互作用的确切性质仍然难以捉摸。我们使用了一种创新的三维细胞培养ECM模型，通过脱细胞的300亩m厚的体外肺组织支架(d3D-LTCs)来自疾病和健康的小鼠肺组织，它广泛模拟了自然(病理)生理在体ECM微环境。我们成功地用原代人和小鼠成纤维细胞重新填充了所有d3D-LTCs，而且，我们还证明了这些细胞也以真正的3D方式填充了d3D-LTCs的最内侧核心区。植入的成纤维细胞表现出惊人的功能可塑性，这取决于它们在d3D-LTCs不同的ECM生态位中的定位，影响细胞的组织植入、细胞迁移速度、细胞形态、蛋白质表达和磷酸化水平。令人惊讶的是，我们还观察到了归巢于肺支架间质的成纤维细胞以及侵袭纤维化区域的成纤维细胞。到目前为止，3D细胞基质黏附的功能性质以及在体内和3D培养模型中的存在仍然是不清楚和有争议的。在这里，我们显示成纤维细胞与d3D-LTCs的附着明显是通过局灶性粘连发生的，因此提倡在体内发挥相关的功能作用。此外，我们发现，与附着在2D塑料培养皿上相比，Talin、Paxlin和Zysin的蛋白水平和Paxlin Y118的磷酸化水平以及迁移相关的小GTP酶RhoA、Rac和CDC42显著降低。综上所述，我们的结果惊人地表明，ECM固有的物理或成分特征可以作为改变移植细胞功能行为的指导性线索。因此，d3D-LTCs可能有助于在体外获得更真实的数据，对药物发现和机制研究都具有很高的相关性。
Cues from the extracellular matrix (ECM) and their functional interplay with cells play pivotal roles for development, tissue repair, and disease. However, the precise nature of this interplay remains elusive. We used an innovative 3D cell culture ECM model by decellularizing 300-mu m-thick ex vivo lung tissue scaffolds (d3D-LTCs) derived from diseased and healthy mouse lungs, which widely mimics the native (patho) physiological in vivo ECM microenvironment. We successfully repopulated all d3D-LTCs with primary human and murine fibroblasts, and moreover, we demonstrated that the cells also populated the innermost core regions of the d3D-LTCs in a real 3D fashion. The engrafted fibroblasts revealed a striking functional plasticity, depending on their localization in distinct ECM niches of the d3D-LTCs, affecting the cells' tissue engraftment, cellular migration rates, cell morphologies, and protein expression and phosphorylation levels. Surprisingly, we also observed fibroblasts that were homing to the lung scaffold's interstitium as well as fibroblasts that were invading fibrotic areas. To date, the functional nature and even the existence of 3D cell matrix adhesions in vivo as well as in 3D culture models is still unclear and controversial. Here, we show that attachment of fibroblasts to the d3D-LTCs evidently occurred via focal adhesions, thus advocating for a relevant functional role in vivo. Furthermore, we found that protein levels of talin, paxillin, and zyxin and phosphorylation levels of paxillin Y118, as well as the migration-relevant small GTPases RhoA, Rac, and CDC42, were significantly reduced compared with their attachment to 2D plastic dishes. In summary, our results strikingly indicate that inherent physical or compositional characteristics of the ECM act as instructive cues altering the functional behavior of engrafted cells. Thus, d3D-LTCs might aid to obtain more realistic data in vitro, with a high relevance for drug discovery and mechanistic studies alike.