Altering the cellular mechanical force balance results in integrated changes in cell, cytoskeletal and nuclear shape.
Altering the cellular mechanical force balance results in integrated changes in cell, cytoskeletal and nuclear shape.
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发表时间：
1992-12
影响因子：
4
通讯作者：
John R. Sims;Seth J. Karp;Donald E. Ingber
中科院分区：
文献类型：
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作者：
John R. Sims;Seth J. Karp;Donald E. Ingber
采用纤维连接蛋白（FN）包被培养皿培养毛细血管内皮细胞，探讨细胞外基质（ECM）调控细胞和细胞核形状的机制。为了研究细胞骨架在形状决定中独立于跨膜渗透压变化的作用，我们使用缓冲液将贴壁细胞的膜用皂素（25微克/毫升）渗透，以保持收缩微丝的功能完整性。实时视频显微镜研究显示，添加250微米ATP导致渗透性细胞和细胞核的时间依赖性收缩和变圆，其方式类似于使用胰蛋白酶- edta分离后在完整活细胞中观察到的情况。计算机图像分析证实，在没有ATP的情况下，通透化的细胞基本上保持刚性，并且当ATP浓度从10微米增加到250微米时，以剂量依赖的方式刺激收缩。最大舍入发生在30分钟，投影的细胞和核区域分别减少了69%和41%。atp诱导的圆角也伴随着微丝的重新分配，导致在收缩的细胞核周围形成密集的f -肌动蛋白网。重要的是，在渗透细胞中，使用合成肌球蛋白肽（IRICRKG）可以防止atp刺激的细胞、细胞骨架和核形式的变化，该肽先前已被证明可以抑制肌动球蛋白丝在肌肉中的滑动。相比之下，当使用可溶性FN肽GRGDSP将固定的FN从细胞表面整合素受体上移除时，暴露于ATP的通透化细胞的细胞和细胞核圆切的速度和程度都增加了。（摘要删节250字）
Studies were carried out with capillary endothelial cells cultured on fibronectin (FN)-coated dishes in order to analyze the mechanism of cell and nuclear shape control by extracellular matrix (ECM). To examine the role of the cytoskeleton in shape determination independent of changes in transmembrane osmotic pressure, membranes of adherent cells were permeabilized with saponin (25 micrograms/ml) using a buffer that maintains the functional integrity of contractile microfilaments. Real-time videomicroscopic studies revealed that addition of 250 microM ATP resulted in time-dependent retraction and rounding of permeabilized cells and nuclei in a manner similar to that observed in intact living cells following detachment using trypsin-EDTA. Computerized image analysis confirmed that permeabilized cells remained essentially rigid in the absence of ATP and that retraction was stimulated in a dose-dependent manner as the concentration of ATP was raised from 10 to 250 microM. Maximal rounding occurred by 30 min with projected cell and nuclear areas being reduced by 69 and 41%, respectively. ATP-induced rounding was also accompanied by a redistribution of microfilaments resulting in formation of a dense net of F-actin surrounding retracted nuclei. Importantly, ATP-stimulated changes in cell, cytoskeletal, and nuclear form were prevented in permeabilized cells using a synthetic myosin peptide (IRICRKG) that has been previously shown to inhibit actomyosin filament sliding in muscle. In contrast, both the rate and extent of cell and nuclear rounding were increased in permeabilized cells exposed to ATP when the soluble FN peptide, GRGDSP, was used to dislodge immobilized FN from cell surface integrin receptors.(ABSTRACT TRUNCATED AT 250 WORDS)