Stepwise mechanical stretching inhibits chondrogenesis through cell-matrix adhesion mediated by integrins in embryonic rat limb-bud mesenchymal cells
Stepwise mechanical stretching inhibits chondrogenesis through cell-matrix adhesion mediated by integrins in embryonic rat limb-bud mesenchymal cells
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DOI：
10.1016/j.ejcb.2004.09.004
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发表时间：
2005-01-01
影响因子：
6.6
通讯作者：
Mitani, H
中科院分区：
文献类型：
--
作者：
Onodera, K;Takahashi, I;Mitani, H
生物力学力是决定骨骼组织形态和分化的主要表观遗传因素，并可通过细胞粘附转导至细胞内生化信号通路。为了验证逐步拉伸在早期软骨形成过程中转化为分子信号的假设，我们开发了一种培养系统来研究软骨细胞的增殖和分化。将大鼠胚胎第12天的肢芽显微解剖并解离成细胞，然后将其在硅树脂膜上微团培养并维持长达7天。将逐步增加的拉伸施加到硅树脂膜，其在软骨形成开始后第4天对培养物施加剪切应力。在拉伸条件下，II型胶原蛋白的表达被显着抑制44%的第1天和67%的第2天，和这种差异在II型胶原蛋白的培养3天后达到80%。II型胶原蛋白的积累和软骨形成结节的大小在第3天减少了50%。另一方面，非软骨形成标志物纤连蛋白的表达在第3天显著上调1.8倍，而I型胶原的上调是最小的，甚至到第3天。当细胞-细胞外基质附着被Gly-Arg-Gly-Asp-Ser-Pro-Lys肽或通过应用α 2、α 5或β 1整联蛋白的阻断抗体抑制时，软骨形成标记物表达的下调完全恢复。我们的结论是，剪切应力产生的逐步拉伸抑制软骨形成通过整合素，并提出，从生物力学刺激的信号转导可能介导的细胞-细胞外基质粘附。(C)2004年Elsevier GmbH。All rights reserved.
Biomechanical forces are major epigenetic factors that determine the form and differentiation of skeletal tissues, and may be transduced through cell adhesion to the intracellular biochemical signaling pathway. To test the hypothesis that stepwise stretching is translated to molecular signals during early chondrogenesis, we developed a culture system to study the proliferation and differentiation of chondrocytes. Rat embryonic day-12 limb buds were microdissected and dissociated into cells, which were then micromass cultured on a silicone membrane and maintained for up to 7 days. Stepwise-increased stretching was applied to the silicone membrane, which exerted shearing stress on the cultures on day 4 after the initiation of chondrogenesis. Under stretched conditions, type II collagen expression was significantly inhibited by 44% on day 1 and by 67% on day 2, and this difference in type II collagen reached 80% after 3 days of culture. Accumulation of type II collagen protein and the size of the chondrogenic nodules had decreased by 50% on day 3. On the other hand, expression of the non-chondrogenic marker fibronectin was significantly up-regulated by 1.8-fold on day 3, while the up-regulation of type I collagen was minimal, even by day 3. The down-regulation in the expression of chondrogenic markers was completely recovered when cell-extracellular matrix attachment was inhibited by Gly-Arg-Gly-Asp-Ser-Pro-Lys peptide or by the application of blocking antibodies for alpha2, alpha5 or beta1 integrins. We conclude that shearing stress generated by stepwise stretching inhibits chondrogenesis through integrins, and propose that signal transduction from biomechanical stimuli may be mediated by cell-extracellular matrix adhesion. (C) 2004 Elsevier GmbH. All rights reserved.