Flow Perfusion Culture of Marrow Stromal Cells Seeded on Porous Biphasic Calcium Phosphate Ceramics

Calcium phosphate ceramics have been widely used for filling bone defects to aid in the regeneration of new bone tissue. Addition of osteogenic cells to porous ceramic scaffolds may accelerate the bone repair process. This study demonstrates the feasibility of culturing marrow stromal cells (MSCs) on porous biphasic calcium phosphate ceramic scaffolds in a flow perfusion bioreactor. The flow of medium through the scaffold porosity benefits cell differentiation by enhancing nutrient transport to the scaffold interior and by providing mechanical stimulation to cells in the form of fluid shear. Primary rat MSCs were seeded onto porous ceramic (60% hydroxyapatite, 40% β-tricalcium phosphate) scaffolds, cultured for up to 16 days in static or flow perfusion conditions, and assessed for osteoblastic differentiation. Cells were distributed throughout the entire scaffold by 16 days of flow perfusion culture whereas they were located only along the scaffold perimeter in static culture. At all culture times, flow perfused constructs demonstrated greater osteoblastic differentiation than statically cultured constructs as evidenced by alkaline phosphatase activity, osteopontin secretion into the culture medium, and histological evaluation. These results demonstrate the feasibility and benefit of culturing cell/ceramic constructs in a flow perfusion bioreactor for bone tissue engineering applications.     

成人骨髓间充质干细胞对造血干细胞体外增殖的影响

目的研究骨髓间充质干细胞（MSC）对脐带血（CB）CD34^＋细胞体外增殖和造血重建能力的影响。方法取人骨髓单个核细胞贴壁培养．梭形细胞完全融合后传代，用流式细胞仪检测免疫表型；将CBCD34^＋细胞接种到MSC或其他培养液中．比较不同培养条件对造血干细胞扩增能力、集落形成能力及黏附分子表达的影响。结果在加入IL-3的培养体系中．在MSC和细胞因子作用下，CD34^＋细胞扩增7d和14d后，有核细胞（NC）、CD34^＋细胞和CDl33^＋细胞数，实验组均显著多于对照组。CD34＋细胞在未加入IL-3的培养体系中培养8d后，实验组NC、CD34^＋细胞、CD34^＋CD38-细胞和造血祖细胞集落扩增倍数均显著高于对照组。扩增后CD34^＋细胞的ALCAM、VLA-α4、VLA-α5、VLA-β1、HCAM、PECAM和LFA-1表达较扩增前无显著变化。结论MSC可为造血干细胞（HSC）体外扩增提供适宜的微环境，有助于CD34^＋细胞体外增殖并抑制HSC分化，保持其造血重建潜能和归巢能力。     

珍珠层水溶性提取物对人骨髓基质细胞成骨性分化的诱导作用

目的:探讨珍珠层水溶性提取物(WSM)对人骨髓基质细胞的诱导作用。方法:原代培养人骨髓基质细胞,条件培养基和WSM分别作用于第3代细胞,对照组不加处理因素。倒置显微镜观察细胞在施加处理因素后的生长状态;采用钙钴法染色检测AKP表达;应用RT-PCR方法检测BMP-2等生长因子表达;应用茜素红染色检测骨髓基质细胞的矿化结节形成情况。结果:施加处理因素第7d,WSM组、条件培养基组的BMP-2表达水平较对照组明显增加;WSM组及条件培养基组TGF-β1表达量较对照组相比未见差异。施加处理因素第7d,WSM组及条件培养基组骨髓基质细胞AKP染色明显,与对照组相比差异显著。每高倍视野阳性细胞数,条件培养基组、WSM明显多于对照组,差异具有显著性(P<0.01);加入处理因素18d,条件培养基组可见典型红色钙化结节形成,WSM组也可见到钙结节形成,但没有条件培养基组典型,对照组偶有零星钙化结节形成。结论:WSM可以促进体外培养的人骨髓基质细胞成骨性分化进程,具有一定的骨诱导作用。     

The Secret Life of Exosomes: What Bees Can Teach Us About Next-Generation Therapeutics

Mechanistic exploration has pinpointed nanosized extracellular vesicles, known as exosomes, as key mediators of the benefits of cell therapy. Exosomes appear to recapitulate the benefits of cells and more. As durable azoic entities, exosomes have numerous practical and conceptual advantages over cells. Will cells end up just being used to manufacture exosomes, or will they find lasting value as primary therapeutic agents? Here, a venerable natural process—the generation of honey—serves as an instructive parable. Flowers make nectar, which bees collect and process into honey. Cells make conditioned medium, which laboratory workers collect and process into exosomes. Unlike flowers, honey is durable, compact, and nutritious, but these facts do not negate the value of flowers themselves. The parallels suggest new ways of thinking about next-generation therapeutics.     

MRI tracking of transplanted iron‐labeled mesenchymal stromal cells in an immune‐compromised mouse model of critical limb ischemia

Peripheral arterial disease is a clinical problem in which mesenchymal stromal cell (MSC) transplantation may offer substantial benefit by promoting the generation of new blood vessels and improving limb ischemia and wound healing via their potent paracrine activities. MRI allows for the noninvasive tracking of cells over time using iron oxide contrast agents to label cells before they are injected or transplanted. However, a major limitation of the tracking of iron oxide‐labeled cells with MRI is the possibility that dead or dying cells will transfer the iron oxide label to local bystander macrophages, making it very difficult to distinguish between viable transplanted cells and endogenous macrophages in the images. In this study, a severely immune‐compromised mouse, with limited macrophage activity, was investigated to examine cell tracking in a system in which bystander cell uptake of dead, iron‐labeled cells or free iron particles was minimized. MRI was used to track the fate of MSCs over 21 days after their intramuscular transplantation in mice with a femoral artery ligation. In all mice, a region of signal loss was observed at the injection site and the volume of signal hypointensity diminished over time. Fluorescence and light microscopy showed that iron‐positive MSCs persisted at the transplant site and often appeared to be integrated in perivascular niches. This was compared with MSC transplantation in immune‐competent mice with femoral artery ligation. In these mice, the regions of signal loss caused by iron‐labeled MSC cleared more slowly, and histology revealed iron particles trapped at the site of cell transplantation and associated with areas of inflammation. Copyright © 2012 John Wiley & Sons, Ltd.