Influence of insulin immobilization to thermoresponsive culture surfaces on cell proliferation and thermally induced cell detachment.
Influence of insulin immobilization to thermoresponsive culture surfaces on cell proliferation and thermally induced cell detachment.
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DOI：
10.1016/j.biomaterials.2004.11.061
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发表时间：
2005-09
期刊：
影响因子：
14
通讯作者：
Hideyuki Hatakeyama;A. Kikuchi;M. Yamato;T. Okano
中科院分区：
文献类型：
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作者：
Hideyuki Hatakeyama;A. Kikuchi;M. Yamato;T. Okano
已经制造并研究了固定有胰岛素的温度响应培养皿，以通过促进更快的细胞增殖来缩短细胞培养周期。简单地通过温度变化将细胞恢复为连续的细胞片。功能化的培养皿制备通过先前报道的电子束接枝共聚N-异丙基丙烯酰胺（IPAAm）与其羧酸衍生的类似物，2-羧基异丙基丙烯酰胺（CIPAAm），具有类似的分子结构IPAAm，但羧酸侧链的组织培养聚苯乙烯皿。然后通过与CIPAAm羧酸酯基团形成标准酰胺键将胰岛素固定在培养皿上。在这些胰岛素固定的培养皿上检测牛颈动脉内皮细胞（ECs）的粘附和增殖。胰岛素固定化显示出在补充血清的培养基中促进细胞增殖。增加组织培养表面上的接枝CIPAAm含量降低了细胞粘附和增殖，即使这些表面含有增加量的固定化胰岛素。该结果意味着在不含CIPAAm的羧酸酯基团的量和用于最佳细胞增殖刺激的固定化胰岛素之间存在离散平衡。在胰岛素固定化表面上生长的细胞可以简单地通过降低培养温度以连续细胞单层的形式回收，而不需要添加外源酶或钙螯合剂。总之，胰岛素修饰的温敏培养皿可能被证明对高级细胞培养和组织工程应用是有用的，因为它们促进细胞增殖，并且培养的细胞可以仅通过降低培养温度而恢复为可行的连续单层。
Temperature-responsive culture dishes immobilized with insulin have been fabricated and studied to shorten cell culture periods by facilitating more rapid cell proliferation. Cells are recovered as contiguous cell sheets simply by temperature changes. Functionalized culture dishes were prepared by previously reported electron beam grafting copolymerization of N-isopropylacrylamide (IPAAm) with its carboxylate-derivatized analog, 2-carboxyisopropylacrylamide (CIPAAm), having similar molecular structure to IPAAm but with carboxylate side chains to tissue culture polystyrene dishes. Insulin was then immobilized onto culture dishes through standard amide bond formation with CIPAAm carboxylate groups. Adhesion and proliferation of bovine carotid artery endothelial cells (ECs) were examined on these insulin-immobilized dishes. Insulin immobilization was shown to promote cell proliferation in serum-supplemented medium. Increasing the grafted CIPAAm content on the tissue culture surfaces reduces cell adhesion and proliferation, even though these surfaces contained increased amounts of immobilized insulin. This result implies that a discrete balance exists between the amount of CIPAAm-free carboxylate groups and immobilized insulin for optimum cell proliferative stimulation. Cells grown on the insulin-immobilized surfaces can be recovered as contiguous cell monolayers simply by lowering culture temperature, without need for exogenous enzyme or calcium chelator additions. In conclusion, insulin-modified thermoresponsive culture dishes may prove useful for advanced cell culture and tissue engineering applications since they facilitate cell proliferation, and cultured cells can be recovered as viable contiguous monolayers by merely reducing culture temperature.