Laboratory for Developmental and Stem Cell Biology, Skeletal Biology and Engineering Research Center, Prometheus, Division of Skeletal Tissue Engineering, and Department and Laboratory of Intensive Care Medicine, KU Leuven, Leuven, Belgium; Institute of Orthopaedics and Musculoskeletal Science, Division of Surgery & Interventional Science, University College London, The Royal National Orthopaedic Hospital, Stanmore, Middlesex, United Kingdom.
Stem Cells Transl Med. 2014 Feb;3(2):218-28. doi: 10.5966/sctm.2012-0137. Epub 2013 Dec 27.
The translation of stem cell-based regenerative solutions from the laboratory to the clinic is often hindered by the culture conditions used to expand cell populations. Although fetal bovine serum (FBS) is widely used, regulatory bodies and safety concerns encourage alternative, xeno-free culturing practices. In an attempt to apply this approach to a bone-forming combination product of human periosteal progenitors (human periosteum derived cells) on a clinically used calcium phosphate carrier, FBS was substituted for human allogeneic serum (hAS) during cell expansion. It was found that cell proliferation was increased in hAS along with an apparent commitment to the osteogenic lineage, indicated by enhanced Runx2 expression, as well as alkaline phosphatase activity and matrix mineralization. Following analysis of signaling pathways, it was found that interferon-mediated signaling was downregulated, whereas JAK-STAT signaling was upregulated. STAT3 phosphorylation was enhanced in hAS-cultured human periosteum derived cells, inhibition of which ablated the proliferative effect of hAS. Furthermore, following in vivo implantation of hAS-cultured cells on NuOss scaffolds, enhanced bone formation was observed compared with FBS (71% increase, p < .001). Interestingly, the de novo-formed bone appeared to have a higher ratio of immature regions to mature regions, indicating that after 8 weeks implantation, tissue-formation processes were continuing. Integration of the implant with the environment appeared to be altered, with a decrease in calcium phosphate grain size and surface area, indicative of accelerated resorption. This study highlights the advantages of using humanized culture conditions for the expansion of human periosteal progenitors intended for bone regeneration.
从实验室到临床，干细胞再生解决方案的转化常常受到用于扩大细胞群体的培养条件的限制。尽管胎牛血清（FBS）被广泛使用，但监管机构和安全问题鼓励使用替代的、无动物源的培养方法。为了将这种方法应用于一种基于人骨膜祖细胞的成骨组合产品（人骨膜来源细胞）上，该产品在临床上使用磷酸钙载体，在细胞扩增过程中用异体人血清（hAS）替代 FBS。结果发现，hAS 中细胞增殖增加，同时向成骨谱系明显分化，表现为 Runx2 表达增强，碱性磷酸酶活性和基质矿化增强。对信号通路进行分析后发现，干扰素介导的信号通路被下调，而 JAK-STAT 信号通路被上调。hAS 培养的人骨膜来源细胞中的 STAT3 磷酸化增强，抑制该磷酸化可消除 hAS 的增殖作用。此外，与 FBS 相比，在 NuOss 支架上植入 hAS 培养的细胞后，观察到骨形成增强（增加 71%，p <.001）。有趣的是，新形成的骨似乎具有更高比例的不成熟区域和成熟区域，表明在植入 8 周后，组织形成过程仍在继续。植入物与环境的整合似乎发生了改变，磷酸钙颗粒尺寸和表面积减小，表明吸收加速。本研究强调了使用人源化培养条件来扩增用于骨再生的人骨膜祖细胞的优势。
