Influence of cholesterol/caveolin-1/caveolae homeostasis on membrane properties and substrate adhesion characteristics of adult human mesenchymal stem cells.
Influence of cholesterol/caveolin-1/caveolae homeostasis on membrane properties and substrate adhesion characteristics of adult human mesenchymal stem cells.
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DOI：
10.1186/s13287-018-0830-4
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发表时间：
2018-04-03
影响因子：
7.5
通讯作者：
Tuan RS
中科院分区：
文献类型：
--
作者：
Sohn J;Lin H;Fritch MR;Tuan RS
成体间充质干细胞（MSC）是组织生长、修复和再生的重要资源。为了更有效地利用MSC，清楚地了解它们对环境线索的反应是至关重要的。目前，关于质膜的组成如何影响干细胞的表型和性质的了解相对较少。脂质分子，特别是胆固醇，在质膜中的存在在调节细胞中的各种生理过程中起着至关重要的作用。在这项研究中，我们研究了胆固醇/小窝蛋白-1（CAV-1）/小窝稳态的扰动对MSC的膜特性和粘附特性的影响。这项研究的结果将有助于了解胆固醇/CAV-1/小窝如何调节细胞膜的各个方面，这些方面对细胞粘附，底物传感和微环境相互作用至关重要。我们产生了五个实验MSC组：1）未处理的MSC; 2）胆固醇耗尽的MSC; 3）胆固醇补充的MSC; 4）用对照、非特异性小干扰（si）RNA转染的MSC;和5）用CAV-1 siRNA转染的MSC。通过Amplex Red胆固醇测定、菲律宾荧光染色和实时聚合酶链反应（PCR）分析每个细胞组的胆固醇状态和CAV-1表达的扰动。采用芘荧光探针染色和流式细胞仪（FACS）分析测定5个实验细胞组的膜流动性。检测细胞与胶原和纤连蛋白的粘附以及细胞表面整合素的表达。向MSC补充胆固醇增加了膜胆固醇，并导致膜流动性降低以及细胞膜上的小窝和CAV-1数量增加。这些细胞表现出α1、α4和β1整合素表达增加，并表现出对纤连蛋白和胶原的较高粘附率。相反，敲低CAV-1表达或MSC上的胆固醇消耗导致小窝含量的平行减少和膜流动性的增加，这是由于胆固醇向细胞膜的递送减少。CAV-1表达缺失的细胞显示细胞表面整合素表达降低，与不同底物的粘附较慢。我们的研究结果表明，胆固醇/CAV-1水平的扰动显着影响MSC的膜特性。这些发现表明，膜胆固醇和/或CAV-1和小窝的修饰可用于操纵MSC的生物活性。本文的在线版本（10.1186/s13287-018-0830-4）包含补充材料，可供授权用户使用。
Adult mesenchymal stem cells (MSCs) are an important resource for tissue growth, repair, and regeneration. To utilize MSCs more effectively, a clear understanding of how they react to environmental cues is essential. Currently, relatively little is known about how the composition of the plasma membranes affects stem cell phenotype and properties. The presence of lipid molecules, including cholesterol in particular, in the plasma membrane plays a crucial role in regulating a variety of physiological processes in cells. In this study, we examined the effects of perturbations in cholesterol/caveolin-1 (CAV-1)/caveolae homeostasis on the membrane properties and adhesive characteristics of MSCs. Findings from this study will contribute to the understanding of how cholesterol/CAV-1/caveolae regulates aspects of the cell membrane important to cell adhesion, substrate sensing, and microenvironment interaction. We generated five experimental MSC groups: 1) untreated MSCs; 2) cholesterol-depleted MSCs; 3) cholesterol-supplemented MSCs; 4) MSCs transfected with control, nonspecific small interfering (si)RNA; and 5) MSCs transfected with CAV-1 siRNA. Each cell group was analyzed for perturbation of cholesterol status and CAV-1 expression by performing Amplex Red cholesterol assay, filipin fluorescence staining, and real-time polymerase chain reaction (PCR). The membrane fluidity in the five experimental cell groups were measured using pyrene fluorescence probe staining followed by FACS analysis. Cell adhesion to collagen and fibronectin as well as cell surface integrin expression were examined. Cholesterol supplementation to MSCs increased membrane cholesterol, and resulted in decreased membrane fluidity and localization of elevated numbers of caveolae and CAV-1 to the cell membrane. These cells showed increased expression of α1, α4, and β1 integrins, and exhibited higher adhesion rates to fibronectin and collagen. Conversely, knockdown of CAV-1 expression or cholesterol depletion on MSCs caused a parallel decrease in caveolae content and an increase in membrane fluidity due to decreased delivery of cholesterol to the cell membrane. Cells with depleted CAV-1 expression showed decreased cell surface integrin expression and slower adhesion to different substrates. Our results demonstrate that perturbations in cholesterol/CAV-1 levels significantly affect the membrane properties of MSCs. These findings suggest that modification of membrane cholesterol and/or CAV-1 and caveolae may be used to manipulate the biological activities of MSCs. The online version of this article (10.1186/s13287-018-0830-4) contains supplementary material, which is available to authorized users.
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5.3
作者：
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通讯作者：
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DOI：
10.1016/j.bbalip.2010.04.001
发表时间：
2010-07-01
影响因子：
4.8
作者：
Fu, Chenglai;He, Jinlong;Zhu, Yi
通讯作者：
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