Lavagnino Michael, Arnoczky Steven P
Laboratory for Comparative Orthopaedic Research, College of Veterinary Medicine, Michigan State University, East Lansing, MI 48824, USA.
J Orthop Res. 2005 Sep;23(5):1211-8. doi: 10.1016/j.orthres.2005.04.001.
An in vitro collagen gel system was used to determine the effect of alterations in cytoskeletal tensional homeostasis on gene expression in tendon cells. Collagen gel matrices, seeded with rat tail tendon cells, underwent cytochalasin D and gel contraction treatments designed to alter the internal cytoskeletal homeostasis of the cells. Gels were examined for cytoskeletal organization using a rhodamine phalloidin stain for actin. The effect of altered cytoskeletal organization on mRNA expression of a catabolic (interstitial collagenase) and anabolic (alpha1(I) collagen) gene was examined using northern blot analysis. Tendon cells in adhered gels demonstrated a highly organized cytoskeleton and showed evidence of alpha1(I) collagen mRNA expression but no evidence of collagenase mRNA expression. Treatment of the attached gel with cytochalasin D disrupted the cytoskeletal organization and resulted in the up-regulation of collagenase mRNA and the inhibition of alpha1(I) collagen mRNA expression. Release of the gels resulted in a cell mediated gel contraction, an immediate loss of cytoskeletal organization, and an mRNA expression pattern similar to that seen with cytochalasin D treatment. Isometric contraction of the gel on itself or around a 3-point traction device resulted in an mRNA expression pattern similar to the adhered gel. Gene expression in the contracted gels could be reversed through chemical cytoskeletal disruption or removal of the traction device which permitted further gel contraction. The results of the study suggest that tendon cells can establish an internal cytoskeletal tension through interactions with their local extracellular environment. Alterations in this tension appear to control the expression of both catabolic and anabolic genes in a reciprocal manner.
采用体外胶原凝胶系统来确定细胞骨架张力稳态改变对肌腱细胞基因表达的影响。接种大鼠尾腱细胞的胶原凝胶基质，接受了细胞松弛素D和凝胶收缩处理，旨在改变细胞内部的细胞骨架稳态。使用罗丹明鬼笔环肽对肌动蛋白进行染色，检查凝胶的细胞骨架组织。采用Northern印迹分析，研究细胞骨架组织改变对分解代谢（间质胶原酶）和合成代谢（α1（I）型胶原）基因mRNA表达的影响。附着在凝胶上的肌腱细胞显示出高度有序的细胞骨架，有α1（I）型胶原mRNA表达的证据，但没有胶原酶mRNA表达的证据。用细胞松弛素D处理附着的凝胶会破坏细胞骨架组织，导致胶原酶mRNA上调，α1（I）型胶原mRNA表达受到抑制。凝胶的释放导致细胞介导的凝胶收缩、细胞骨架组织立即丧失，以及与细胞松弛素D处理相似的mRNA表达模式。凝胶自身或围绕三点牵引装置进行等长收缩，会产生与附着凝胶相似的mRNA表达模式。收缩凝胶中的基因表达可通过化学性细胞骨架破坏或移除牵引装置来逆转，这会使凝胶进一步收缩。该研究结果表明，肌腱细胞可通过与其局部细胞外环境的相互作用建立内部细胞骨架张力。这种张力的改变似乎以相互的方式控制分解代谢和合成代谢基因的表达。
