Meier L, Hay E D
J Cell Biol. 1975 Aug;66(2):275-91. doi: 10.1083/jcb.66.2.275.
The present study was undertaken to determine whether or not physical contact with the substratum is essential for the stimulatory effect of extracellular matrix (ECM) on corneal epithelial collagen synthesis. Previous studies showed that collagenous substrata stimulate isolated epithelia to produce three times as much collagen as they produce on noncollagenous substrate; killed collagenous substrata (e.g., lens capsule) are just as effective as living substrata (e.g., living lens) in promoting the production of new corneal stroma in vitro. In the experiments to be reported here, corneal epithelia were placed on one side of Nucleopore filters of different pore sizes and killed lens capsule on the other, with the expectation that contact of the reacting cells with the lens ECM should be limited by the number and size of the cell processes that can tranverse the pores. Transfilter cultures were grown for 24 h in [3H]proline-containing median and incorporation of isotope into hot trichloroacetic acid-soluble protein was used to measure corneal epithelial collagen production. Epithelial collagen synthesis increases directly as the size of the pores in the interposed filter increases and decreases as the thickness of the filter layer increases. Cell processes within Nucleopore filters were identified with the transmission electron microscope with difficulty; with the scanning electron microscope, however, the processes could easily be seen emerging from the undersurface of even 0.1-mum pore size filters. Morphometric techniques were used to show that cell surface area thus exposed to the underlying ECM is linearly correlated with enhancement of collagen synthesis. Epithelial cell processes did not pass through ultrathin (25-mum thick) 0.45-mum pore size Millipore filters nor did "induction" occur across them. The results are discussed in relation to current theories of embryonic tissue interaction.
本研究旨在确定与基质的物理接触对于细胞外基质（ECM）对角膜上皮胶原合成的刺激作用是否至关重要。先前的研究表明，胶原基质刺激分离的上皮细胞产生的胶原量是它们在非胶原基质上产生量的三倍；在体外促进新角膜基质的产生方面，灭活的胶原基质（如晶状体囊）与活基质（如活晶状体）一样有效。在本报告的实验中，将角膜上皮置于不同孔径的核孔滤膜的一侧，另一侧放置灭活的晶状体囊，预期反应细胞与晶状体ECM的接触应受能够穿过孔的细胞突起的数量和大小的限制。跨滤膜培养物在含[3H]脯氨酸的培养基中培养24小时，通过将同位素掺入热的三氯乙酸可溶性蛋白中来测量角膜上皮胶原的产生。上皮胶原合成随着插入滤膜孔径的增大而直接增加，随着滤膜层厚度的增加而减少。用透射电子显微镜很难识别核孔滤膜内的细胞突起；然而，用扫描电子显微镜，即使是孔径为0.1μm的滤膜下表面伸出的突起也很容易看到。形态计量学技术用于表明这样暴露于下层ECM的细胞表面积与胶原合成的增强呈线性相关。上皮细胞突起未穿过超薄（25μm厚）、孔径为0.45μm的微孔滤膜，且在其两侧均未发生“诱导”。结合当前胚胎组织相互作用的理论对结果进行了讨论。
