In vitro two-dimensional (2D) and three-dimensional (3D) cultivation of mammalian cells requires supplementation with serum. Mesenchymal stem cells (MSCs) are widely used in clinical trials for bioregenerative medicine and in most cases, in vitro expansion and differentiation of these cells are required before application. Optimized expansion and differentiation protocols play a key role in the treatment outcome. 3D cell cultivation systems are more comparable to in vivo conditions and can provide both, more physiological MSC expansion and a better understanding of intercellular and cell-matrix interactions. Xeno-free cultivation conditions minimize risks of immune response after implantation. Human platelet lysate (hPL) appears to be a valuable alternative to widely used fetal calf serum (FCS) since no ethical issues are associated with its harvest, it contains a high concentration of growth factors and cytokines and it can be produced from expired platelet concentrate. In this study, we analyzed and compared proliferation, as well as osteogenic and chondrogenic differentiation of human adipose tissue-derived MSCs (hAD-MSC) using three different supplements: FCS, human serum (HS), and hPL in 2D. Furthermore, online monitoring of osteogenic differentiation under the influence of different supplements was performed in 2D. hPL-cultivated MSCs exhibited a higher proliferation and differentiation rate compared to HS- or FCS-cultivated cells. We demonstrated a fast and successful chondrogenic differentiation in the 2D system with the addition of hPL. Additionally, FCS, HS, and hPL were used to formulate Gelatin-methacryloyl (GelMA) hydrogels in order to evaluate the influence of the different supplements on the cell spreading and proliferation of cells growing in 3D culture. In addition, the hydrogel constructs were cultivated in media supplemented with three different supplements. In comparison to FCS and HS, the addition of hPL to GelMA hydrogels during the encapsulation of hAD-MSCs resulted in enhanced cell spreading and proliferation. This effect was promoted even further by cultivating the hydrogel constructs in hPL-supplemented media.

中文翻译：

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2D 和 3D 系统中胎牛血清、人血清和血小板裂解物中间充质干细胞培养的比较分析

哺乳动物细胞的体外二维 (2D) 和三维 (3D) 培养需要补充血清。间充质干细胞 (MSCs) 广泛用于生物再生医学的临床试验，在大多数情况下，这些细胞在应用前需要进行体外扩增和分化。优化的扩增和分化方案在治疗结果中起着关键作用。3D 细胞培养系统与体内条件更具可比性，可以提供更生理性的 MSC 扩增和更好地了解细胞间和细胞-基质相互作用。无异种培养条件可最大限度地降低植入后免疫反应的风险。人血小板裂解物 (hPL) 似乎是广泛使用的胎牛血清 (FCS) 的有价值的替代品，因为其收获不涉及伦理问题，它含有高浓度的生长因子和细胞因子，并且可以从过期的血小板浓缩物中生产. 在这项研究中，我们使用三种不同的补充剂分析和比较了人脂肪组织来源的 MSC (hAD-MSC) 的增殖以及成骨和软骨分化：FCS、人血清 (HS) 和 2D 中的 hPL。此外，在线监测不同补充剂影响下的成骨分化是在 2D 中进行的。与 HS 或 FCS 培养的细胞相比，hPL 培养的 MSC 表现出更高的增殖和分化率。我们在添加 hPL 的 2D 系统中展示了快速且成功的软骨分化。此外，FCS、HS 和 hPL 用于配制明胶-甲基丙烯酰 (GelMA) 水凝胶，以评估不同补充剂对 3D 培养中细胞扩散和增殖的影响。此外，水凝胶构建体在补充有三种不同补充剂的培养基中培养。与 FCS 和 HS 相比，在 hAD-MSC 封装期间向 GelMA 水凝胶添加 hPL 导致细胞扩散和增殖增强。通过在补充 hPL 的培养基中培养水凝胶构建体，可以进一步促进这种效果。和 hPL 用于配制明胶-甲基丙烯酰 (GelMA) 水凝胶，以评估不同补充剂对 3D 培养中细胞扩散和增殖的影响。此外，水凝胶构建体在补充有三种不同补充剂的培养基中培养。与 FCS 和 HS 相比，在 hAD-MSC 封装期间向 GelMA 水凝胶添加 hPL 导致细胞扩散和增殖增强。通过在补充 hPL 的培养基中培养水凝胶构建体，可以进一步促进这种效果。和 hPL 用于配制明胶-甲基丙烯酰 (GelMA) 水凝胶，以评估不同补充剂对 3D 培养中细胞扩散和增殖的影响。此外，水凝胶构建体在补充有三种不同补充剂的培养基中培养。与 FCS 和 HS 相比，在 hAD-MSCs 封装期间向 GelMA 水凝胶添加 hPL 导致细胞扩散和增殖增强。通过在补充 hPL 的培养基中培养水凝胶构建体，可以进一步促进这种效果。在 hAD-MSCs 封装期间向 GelMA 水凝胶添加 hPL 导致细胞扩散和增殖增强。通过在补充 hPL 的培养基中培养水凝胶构建体，可以进一步促进这种效果。在 hAD-MSCs 封装期间向 GelMA 水凝胶添加 hPL 导致细胞扩散和增殖增强。通过在补充 hPL 的培养基中培养水凝胶构建体，可以进一步促进这种效果。