Bara Jennifer J, Herrmann Marietta, Menzel Ursula, Benneker Lorin, Alini Mauro, Stoddart Martin J
AO Research Institute Davos, Davos Platz, Switzerland.
Department of Orthopaedic Surgery, Inselspital, Bern University Hospital and University of Bern, Bern, Switzerland.
Cytotherapy. 2015 Apr;17(4):458-72. doi: 10.1016/j.jcyt.2014.12.011. Epub 2015 Feb 10.
The diverse phenotypic changes and clinical and economic disadvantages associated with the monolayer expansion of bone marrow-derived mesenchymal stromal cells (MSCs) have focused attention on the development of one-step intraoperative cells therapies and homing strategies. The mononuclear cell fraction of bone marrow, inclusive of discrete stem cell populations, is not well characterized, and we currently lack suitable cell culture systems in which to culture and investigate the behavior of these cells.
Human bone marrow-derived mononuclear cells were cultured within fibrin for 2 weeks with or without fibroblast growth factor-2 supplementation. DNA content and cell viability of enzymatically retrieved cells were determined at days 7 and 14. Cell surface marker profiling and cell cycle analysis were performed by means of multi-color flow cytometry and a 5-ethynyl-2'-deoxyuridine incorporation assay, respectively.
Total mononuclear cell fractions, isolated from whole human bone marrow, was successfully cultured in fibrin gels for up to 14 days under static conditions. Discrete niche cell populations including MSCs, pericytes and hematopoietic stem cells were maintained in relative quiescence for 7 days in proportions similar to that in freshly isolated cells. Colony-forming unit efficiency of enzymatically retrieved MSCs was significantly higher at day 14 compared to day 0; and in accordance with previously published works, it was fibroblast growth factor-2-dependant.
Fibrin gels provide a simple, novel system in which to culture and study the complete fraction of bone marrow-derived mononuclear cells and may support the development of improved bone marrow cell-based therapies.
骨髓间充质基质细胞（MSCs）单层扩增所伴随的多种表型变化以及临床和经济方面的劣势，已使人们将注意力集中在一步法术中细胞治疗和归巢策略的开发上。骨髓中的单核细胞部分，包括离散的干细胞群体，其特征尚不明确，并且我们目前缺乏合适的细胞培养系统来培养和研究这些细胞的行为。
将人骨髓来源的单核细胞在有或无成纤维细胞生长因子-2补充的情况下于纤维蛋白中培养2周。在第7天和第14天测定酶解回收细胞的DNA含量和细胞活力。分别通过多色流式细胞术和5-乙炔基-2'-脱氧尿苷掺入试验进行细胞表面标志物分析和细胞周期分析。
从全人骨髓中分离出的总单核细胞部分在静态条件下成功地在纤维蛋白凝胶中培养了长达14天。包括MSCs、周细胞和造血干细胞在内的离散龛位细胞群体在7天内保持相对静止，其比例与新鲜分离细胞中的比例相似。与第0天相比，酶解回收的MSCs在第14天的集落形成单位效率显著更高；并且与先前发表的研究一致，这是成纤维细胞生长因子-2依赖性的。
纤维蛋白凝胶提供了一个简单、新颖的系统，可用于培养和研究骨髓来源单核细胞的完整部分，并可能支持改进的基于骨髓细胞的治疗方法的开发。
