Surface engineering of living myoblasts via selective periodate oxidation
Surface engineering of living myoblasts via selective periodate oxidation
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DOI：
10.1002/bit.10525
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发表时间：
2003-03-30
影响因子：
3.8
通讯作者：
Shakesheff, KM
中科院分区：
文献类型：
--
作者：
De Bank, PA;Kellam, B;Shakesheff, KM
细胞表面分子对于正常细胞活动至关重要。为了研究这些分子的功能或操纵细胞行为，用我们选择的生物活性分子装饰细胞表面的能力是一种潜在的强大技术。在这里，我们描述了通过选择性高碘酸氧化唾液酸残基对活 L6 成肌细胞单层进行的分子工程，以及这种表面修饰在细胞人工聚集中的应用。该反应产生的醛基用于选择性地将模型分子生物素酰肼连接到细胞表面。用荧光缀合亲和素染色后的流式细胞术分析显示，与未处理的细胞相比，荧光强度呈浓度依赖性增加，最大位移为 345.1 +/- 27.4 倍，EC50 为 17.4 +/- 1.1 muM。这种温和的氧化反应不会影响细胞数量、活力或形态。然后，我们将这种化学技术与使用 N-乙酰丙酰甘露糖胺 (ManLev) 的反应性细胞表面酮基的代谢掺入进行比较。在该细胞系中，当成肌细胞与高浓度的 ManLev 一起孵育 48 小时时，与未处理的细胞相比，仅观察到 22.3 倍的荧光位移。然后使用高碘酸盐氧化来修饰成肌细胞表面以诱导细胞聚集。生物素化成肌细胞在培养物中不会自发聚集，与抗生物素蛋白交联导致毫米大小的多细胞结构的快速形成。这些数据表明，高碘酸钠处理是一种有效的、无细胞毒性的活细胞表面分子工程方法，具有细胞生物学和组织工程应用的潜力。 (C) 2003 年 Wiley 期刊公司。
Cell surface molecules are vital for normal cell activity. To study the functions of these molecules or manipulate cell behavior, the ability to decorate cell surfaces with bioactive molecules of our choosing is a potentially powerful technique. Here, we describe the molecular engineering of living L6 myoblast monolayers via selective periodate oxidation of sialic acid residues and the application of this surface modification in the artificial aggregation of cells. The aldehyde groups generated by this reaction were used to selectively ligate a model molecule, biotin hydrazide, to the cell surfaces. Flow cytometry analysis after staining with fluorescently conjugated avidin revealed a concentration-dependent increase in fluorescence compared to untreated cells with a maximal shift of 345.1 +/- 27.4-fold and an EC50 of 17.4 +/- 1.1 muM. This mild oxidation reaction did not affect cell number, viability, or morphology. We then compared this chemical technique with the metabolic incorporation of reactive cell surface ketone groups using N-levulinoylmannosamine (ManLev). In this cell line, only a 22.3-fold fluorescence shift was observed compared to untreated cells when myoblasts were incubated with a high concentration of ManLev for 48 hours. Periodate oxidation was then used to modify myoblast surfaces to induce cell aggregation. Crosslinking biotinylated myoblasts, which do not spontaneously aggregate in culture, with avidin resulted in the rapid formation of millimeter-sized, multicellular structures. These data indicate that sodium periodate treatment is an effective, noncytotoxic method for the in vitro molecular engineering of living cell surfaces with the potential for cell biology and tissue engineering applications. (C) 2003 Wiley Periodicals, Inc.