A novel 'sandwich' assay for quantifying chemo-regulated cell migration within 3-dimensional matrices: wound healing cytokines exhibit distinct motogenic activities compared to the transmembrane assay.
A novel 'sandwich' assay for quantifying chemo-regulated cell migration within 3-dimensional matrices: wound healing cytokines exhibit distinct motogenic activities compared to the transmembrane assay.
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DOI：
10.1002/cm.20123
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发表时间：
2006-05
影响因子：
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通讯作者：
Seth L. Schor;Ian R. Ellis;Koji Harada;Katsumi Motegi;Alexander R.A. Anderson;M. Chaplain;R. Keatch;A. Schor
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文献类型：
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作者：
Seth L. Schor;Ian R. Ellis;Koji Harada;Katsumi Motegi;Alexander R.A. Anderson;M. Chaplain;R. Keatch;A. Schor
细胞外基质深刻地影响细胞对可溶性运动原的反应。鉴于基质功能的这一关键方面，我们开发了一种新的测定方法来定量生物相关的三维基质内化学调节的细胞迁移。在该“夹心”测定中，在存在各向同性（均匀）或各向异性（梯度）空间分布的测试原的情况下，将靶细胞接种在上基质隔室和下基质隔室之间的界面处。响应于不同条件的细胞迁移通过量化它们在上基质隔室和下基质隔室中的后续分布来确定。本研究的目的是比较血小板衍生生长因子（PDGF-AB）和转化生长因子-β亚型（TGF-β 1，-β 2和-β 3）在夹心试验和常用的跨膜试验中的运动活性。如前所述，真皮成纤维细胞在跨膜试验中对所有测试的细胞因子的各向同性和各向异性分布表现出运动反应。相比之下，只有PDGF-AB和TGF-β 3在夹心试验中是有活性的，各自引起响应于各向同性细胞因子分布和响应于各向异性分布的定向偏置响应的向上和向下I型胶原基质的定向无偏（对称）迁移。TGF-β 1和-β 2完全没有运动活性。这些结果是一致的报告差异生物活性的PDGF和TGF-β 3相比，TGF-β 1和-β 2在动物模型的伤口愈合，并表明，三明治试验提供了一种手段，获得生理相关的数据化学调节细胞迁移。
The extracellular matrix profoundly affects cellular response to soluble motogens. In view of this critical aspect of matrix functionality, we have developed a novel assay to quantify chemo-regulated cell migration within biologically relevant 3-dimensional matrices. In this "sandwich" assay, target cells are plated at the interface between an upper and lower matrix compartment, either in the presence of an isotropic (uniform) or anisotropic (gradient) spatial distribution of test motogen. Cell migration in response to the different conditions is ascertained by quantifying their subsequent disposition within the upper and lower matrix compartments. The objective of this study has been to compare the motogenic activities of platelet-derived growth factor (PDGF-AB) and transforming growth factor-beta isoforms (TGF-beta1, -beta2 and -beta3) in the sandwich assay and the commonly employed transmembrane assay. As previously reported, dermal fibroblasts exhibited a motogenic response to isotropic and anisotropic distributions of all tested cytokines in the transmembrane assay. In contrast, only PDGF-AB and TGF-beta3 were active in the sandwich assay, each eliciting directionally unbiased (symmetrical) migration into the upper and lower type I collagen matrices in response to an isotropic cytokine distribution and a directionally biased response to an anisotropic distribution. TGF-beta1 and -beta2 were completely devoid of motogenic activity. These results are consistent with the reported differential bioactivities of PDGF and TGF-beta3 compared to TGF-beta1 and -beta2 in animal models of wound healing and suggest that the sandwich assay provides a means of obtaining physiologically relevant data regarding chemo-regulated cell migration.