Effect of differentiation on endocytic profiles of endothelial and epithelial cell culture models
Effect of differentiation on endocytic profiles of endothelial and epithelial cell culture models
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DOI：
10.1016/j.yexcr.2015.01.002
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发表时间：
2015-03-01
影响因子：
3.7
通讯作者：
Yliperttula, Marjo
中科院分区：
文献类型：
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作者：
Ilina, Polina;Partti, Susanna;Yliperttula, Marjo
了解纳米颗粒的内吞作用和通过内皮和上皮屏障运输纳米颗粒的机制可能会提高纳米颗粒药物输送系统的效率。关于胞内途径表达和活性的细胞模型的详细特征(胞内侧写)应有助于数据解释。我们对Caco-2和hCMEC/D3细胞进行了内吞图谱分析，这两种细胞分别被广泛用作人类肠道和血脑屏障通透性模型，在细胞分化过程中。此外，我们比较了细胞系和原代细胞的内吞图谱。实时定量聚合酶链式反应分析特定内吞途径相关基因在mRNA水平的表达。在可能的情况下，用Western blotting分析各自的蛋白水平，用流式细胞仪分析细胞的内吞活性。我们发现，分化的Caco-2细胞形成紧密的、极化良好的单层，内吞活性降低，同时大多数内吞相关基因的mRNA表达减少。相反，hCMEC/D3细胞形成了一个漏电的、极化较少的屏障，体外分化对这些细胞的内吞相关基因的表达或内吞活性几乎没有影响。体外模型的内吞图谱和与原代细胞的比较是避免纳米粒渗透研究中误导性结论的重要措施。(C)2015 Elsevier Inc.保留所有权利。
Understanding mechanisms of endocytosis and trafficking of nanoparticles through endothelial and epithelial barriers leads potentially to improved efficacy of nanoparticulate drug delivery systems. Detailed characterizations of cell models with respect to endocytic pathway expression and activity (endocytic profiling) should facilitate data interpretation. We performed endocytic profiling of CaCo-2 and hCMEC/D3 cell lines, widely used as human intestinal and blood-brain barrier permeability models, respectively, during cell differentiation. Furthermore, we compared endocytic profiles of cell lines with those of primary cells. Expression of genes involved in specific endocytic pathways was analyzed at mRNA levels by quantitative real time PCR. Where possible, the respective protein levels were analyzed by Western blotting, and endocytic activities of cells were analyzed by flow cytometry. We showed that differentiated CaCo-2 cells formed tight, well polarized monolayers with reduced endocytic activity accompanied by reduced mRNA expression of most of the endocytosis-related genes. In contrast, hCMEC/D3 cells formed a leaky, less polarized barrier, and in vitro differentiation had little effect on either the expression of endocytosis-related genes or endocytic activity of these cells. Endocytic profiling of in vitro models and comparison with primary cells is an important measure to avoid misleading conclusions in nanoparticle permeation studies. (C) 2015 Elsevier Inc. All rights reserved.