Department for Trauma Surgery, University of Regensburg Medical Center, Regensburg, Germany.
Int Orthop. 2013 Jan;37(1):153-8. doi: 10.1007/s00264-012-1726-z. Epub 2012 Dec 11.
Insulin is a commonly used additive in chondrogenic media for differentiating mesenchymal stem cells (MSCs). The indispensability of other bioactive factors like TGF-β or dexamethasone in these medium formulations has been shown, but the role of insulin is unclear. The purpose of this study was to investigate whether insulin is essential for MSC chondrogenesis and if there is a dose-dependent effect of insulin on MSC chondrogenesis.
We cultivated human MSCs in pellet culture in serum-free chondrogenic medium with insulin concentrations between 0 and 50 μg/ml and assessed the grade of chondrogenic differentiation by histological evaluation and determination of glycosaminoglycan (GAG), total collagen and DNA content. We further tested whether insulin can be delivered in an amount sufficient for MSC chondrogenesis via a drug delivery system in insulin-free medium.
Chondrogenesis was not induced by standard chondrogenic medium without insulin and the expression of cartilage differentiation markers was dose-dependent at insulin concentrations between 0 and 10 μg/ml. An insulin concentration of 50 μg/ml had no additional effect compared with 10 μg/ml. Insulin was delivered by a release system into the cell culture under insulin-free conditions in an amount sufficient to induce chondrogenesis.
Insulin is essential for MSC chondrogenesis in this system and chondrogenic differentiation is influenced by insulin in a dose-dependent manner. Insulin can be provided in a sufficient amount by a drug delivery system. Therefore, insulin is a suitable and inexpensive indicator substance for testing drug release systems in vitro.
胰岛素是用于分化间充质干细胞（MSCs）的软骨形成培养基中常用的添加剂。已经表明，在这些培养基配方中，TGF-β或地塞米松等其他生物活性因子是不可或缺的，但胰岛素的作用尚不清楚。本研究的目的是研究胰岛素是否对 MSC 软骨形成是必需的，以及胰岛素对 MSC 软骨形成是否存在剂量依赖性作用。
我们在无血清软骨形成培养基中以 0 至 50μg/ml 的胰岛素浓度进行微球体培养，培养人 MSCs，并通过组织学评估和糖胺聚糖（GAG）、总胶原和 DNA 含量的测定来评估软骨分化程度。我们进一步测试了胰岛素是否可以通过无胰岛素的药物输送系统以足够的量输送到 MSC 软骨形成中。
标准无胰岛素软骨形成培养基不能诱导软骨形成，并且在 0 至 10μg/ml 的胰岛素浓度范围内，软骨分化标志物的表达呈剂量依赖性。与 10μg/ml 相比，50μg/ml 的胰岛素浓度没有额外的效果。胰岛素在无胰岛素的条件下通过释放系统输送到细胞培养中，其量足以诱导软骨形成。
在该系统中，胰岛素是 MSC 软骨形成所必需的，并且胰岛素以剂量依赖性方式影响软骨分化。胰岛素可以通过药物输送系统以足够的量提供。因此，胰岛素是体外测试药物释放系统的合适且廉价的指示剂。
