Effect of Serum and Insulin Modulation on the Organization and Morphology of Matrix Synthesized by Bovine Corneal Stromal Cells
Effect of Serum and Insulin Modulation on the Organization and Morphology of Matrix Synthesized by Bovine Corneal Stromal Cells
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DOI：
10.1089/ten.tea.2008.0404
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发表时间：
2009-11-01
影响因子：
4.1
通讯作者：
Ruberti, Jeffrey W.
中科院分区：
文献类型：
--
作者：
Bueno, Ericka M.;Saeidi, Nima;Ruberti, Jeffrey W.
角膜角质细胞在三维无支架培养系统中体外生产高度组织化的胶原原纤维，为直接观察有组织的基质形成提供了独特的机会。本研究的目的是在玻璃基质中开发这样一种培养系统（为了光学可及性），并直接检查减少血清和/或增加胰岛素对第四至第五代牛角膜基质角质细胞分层和分泌排列基质的影响。研究了0%、1%或10%胎牛血清和0%或1%胰岛素-转铁蛋白-硒的中浓度。采用高分辨率差干涉对比显微镜、速冻/深刻蚀和常规透射电镜来监测细胞-基质结构的演变、形态和超微结构。在含有血清和胰岛素-转铁蛋白-硒各1%的培养基中，基质细胞分层并分泌丰富的局部排列基质，产生最厚的细胞-基质结构（允许用镊子处理）。这项研究的结果有可能通过(i)阐明调节有组织基质的体外细胞分泌的线索和（ii）建立一个光学可及的细胞培养系统来研究排列的胶原原纤维的细胞分泌机制，从而显著推进承重组织的发育功能工程领域。
The in vitro production of highly organized collagen fibrils by corneal keratocytes in a three-dimensional scaffold-free culture system presents a unique opportunity for the direct observation of organized matrix formation. The objective of this investigation was to develop such a culture system in a glass substrate (for optical accessibility) and to directly examine the effect of reducing serum and/or increasing insulin on the stratification and secretion of aligned matrix by fourth- to fifth-passage bovine corneal stromal keratocytes. Medium concentrations of 0%, 1%, or 10% fetal bovine serum and 0% or 1% insulin-transferrin-selenium were investigated. High-resolution differential interference contrast microscopy, quick-freeze/deep-etch, and conventional transmission electron microscopy were used to monitor the evolution, morphology, and ultrastructure of the cell-matrix constructs. In a medium containing 1% each of serum and insulin-transferrin-selenium, stromal cells stratified and secreted abundant and locally aligned matrix, generating the thickest cell-matrix constructs (allowing handling with forceps). The results of this study have the potential to significantly advance the field of developmental functional engineering of load-bearing tissues by (i) elucidating cues that modulate in vitro cell secretion of organized matrix and (ii) establishing an optically accessible cell culture system for investigating the mechanism of cell secretion of aligned collagen fibrils.