Gurevich Olga, Vexler Akiva, Marx Gerard, Prigozhina Tatyana, Levdansky Lila, Slavin Shimon, Shimeliovich Irina, Gorodetsky Raphael
BMT Department, Hadassah University Hospital, Jerusalem, Israel.
Tissue Eng. 2002 Aug;8(4):661-72. doi: 10.1089/107632702760240571.
It has been demonstrated that bone marrow (BM)-derived pluripotent stem cells can be incorporated into muscle, bone, nerve, lung, stomach, intestine, and skin. Fibrin-based biodegradable microbeads (FMB) were developed for culturing, in suspension, a high density of cells, mostly of mesenchymal origin. In the current study, FMB were used to isolate and expand mesenchymal progenitor cells from BM of mice and rats. Cells from BM isolated on FMB (FMB-BM cells) were visualized by fluorescent confocal microscopy and quantified by a modified MTS colorimetric assay. Downloading the BM cells from FMB onto plastic induced their differentiation into islets of cells with osteogenic phenotype that secreted mineralized extracellular matrix. This was augmented by inducers of osteogenesis, such as ascorbic acid, beta-glycerophosphate, and dexamethasone, or osteoblast-growth peptides (OGP). Implanting FMB-BM cells under the kidney capsule in mouse tested the osteogenic potential of these cells in vivo. Thirty days after implantation, bone structures with typical BM elements were seen in 8/53 kidneys in 6-Gy-irradiated mice and in 1/10 kidneys in nonirradiated recipients; bone formation was verified by soft x-ray imaging and elemental analysis that showed elevated Ca and Fe in the implant region. FMB-BM cells - downloaded onto plastic flasks, cultured for 2 weeks, mechanically harvested and then implanted - induced 100% bone formation in both irradiated (6/6) and nonirradiated (3/3) mice. Histology revealed well-organized bone structures under the kidney capsule, including osteoblasts and typical elements of BM. Our findings demonstrate that FMB are capable of isolating and expanding progenitor cells from BM for osteogenesis and possibly for regenerating other mesenchymal tissues.
业已证明，骨髓来源的多能干细胞能够整合到肌肉、骨骼、神经、肺、胃、肠和皮肤中。基于纤维蛋白的可生物降解微珠（FMB）被开发用于在悬浮状态下培养高密度的细胞，这些细胞大多起源于间充质。在本研究中，FMB被用于从小鼠和大鼠的骨髓中分离并扩增间充质祖细胞。在FMB上分离的骨髓细胞（FMB-BM细胞）通过荧光共聚焦显微镜进行可视化，并通过改良的MTS比色法进行定量。将FMB上的骨髓细胞接种到塑料培养皿上可诱导其分化为具有成骨表型的细胞胰岛，这些细胞能分泌矿化的细胞外基质。这可通过成骨诱导剂（如抗坏血酸、β-甘油磷酸和地塞米松）或成骨细胞生长肽（OGP）增强。将FMB-BM细胞植入小鼠肾包膜下，以在体内测试这些细胞的成骨潜力。植入后30天，在6-Gy照射的小鼠中，53个肾脏中有8个出现了具有典型骨髓成分的骨结构，在未照射的受体小鼠中，10个肾脏中有1个出现了这种情况；通过软X射线成像和元素分析证实了骨形成，结果显示植入区域的钙和铁含量升高。将FMB-BM细胞接种到塑料培养瓶上，培养2周，机械收获后再植入，在照射（6/6）和未照射（3/3）的小鼠中均诱导了100%的骨形成。组织学检查显示肾包膜下有组织良好的骨结构，包括成骨细胞和典型的骨髓成分。我们的研究结果表明，FMB能够从骨髓中分离并扩增祖细胞用于成骨，也可能用于再生其他间充质组织。
