Development of plastic and reconstructive operation using mesenchymal stem cells derived from peripheral tissues

在过去的几年里，人们对成体干细胞的普遍性和可塑性的认识发生了巨大的变化。在各种成体组织中已经证实了自我更新干细胞，它们显示出沿着意想不到的途径分化的潜力，不仅产生了相同发育谱系的细胞，而且产生了不同于干细胞来源的谱系。最典型的例子是骨髓来源的干细胞，它在体外可以分化为成骨细胞、软骨细胞、脂肪细胞和成肌细胞，在体内可以并入骨、骨骼肌、肺和肠道以及中枢神经系统。成体干细胞的另一个例子是脂肪干细胞，它可以沿着间充质层分化为脂肪细胞、软骨细胞、肌细胞和成骨细胞，也可以沿着神经元连接分化。在本研究中，我们发现脂肪组织分散的间质血管部分中的未分化干细胞群可以被…只有在含有成纤维细胞生长因子-2的低血清培养液中培养，才能扩增更多的细胞。在随着血清中含有的分化因子的减少培养条件下，未分化的干细胞群体表现出几乎无限的增殖潜力，所产生的细胞表现出向脂肪细胞、成骨细胞和软骨细胞的高分化频率。因此，我们认为脂肪组织中的干细胞在低血清介质中维持其自我更新周期，保留了多向分化的潜能。最近研究ASC的科学家发现，脂肪组织中含有大量间充质干细胞和血管母细胞(中血管母细胞)的共同祖细胞。荧光激活细胞分选仪分析表明，在脂肪组织散在的间质血管部分中，这类干细胞(CD34+)占细胞的主要数量。在脂肪组织中有大量的多能干细胞，下一个需要解决的问题是如何在不自发增殖和分化的情况下保持ASCs的静止。我们曾经证明，层粘连蛋白α4链的基因表达依赖于3T3-L1成纤维细胞的脂肪转化。A4链的层粘连蛋白G样结构域(LG)4模块与Syndecans有很强的亲和力。我们在体内发现α4 LG3-5是从层粘连蛋白-8(α4β1γ1)中切割出来的，并探讨了其作为阻断FGF2信号转导的配体的功能。在注射Matrigel+FGF2处的小鼠新生成脂实验中，在20 nm处加入α_4LG4-5可完全抑制成脂。由于α4LG1-5的作用较弱，因此LG4-5的活性在G结构域的整体结构中是隐蔽的。α-4LG4在其肝素结合活性降低的突变体中失去抑制作用。抗Syndecan-2和Syndecan-4胞外(但不是胞浆)结构域的抗体抑制脂肪生成。因此，由于α4 G结构域片段与Syndecans的竞争性结合，通过Syndecans传递的信号似乎被阻断。较少

Appreciation of ubiquity and plasticity of adult stem cells has changed dramatically in the last several years. Self-renewing stem cells have been demonstrated in various adult tissues and they show potential of differentiation along unexpected pathways, giving rise to the cells of not only the same developmental linage but also the linage different from which the stem cells were derived. Well-characterized example is bone marrow-derived stem cells that can differentiate in vitro into osteoblasts, chondrocytes, adipocytes and myoblasts, and can be incorporated in vivo into bone, skeletal muscle, lung and gut and the central nervous system. Another example of adult stem cell is adipose-derived stem cell(ASC) that can differentiate along mesenchymal linage to produce adipocytes, chondrocytes, myocytes and osteoblasts and also along neuronal linage.In this research, we fond that population of undifferentiated stem cells in stromal vascular fraction(SVF) dispersed from adipose tissue can b … More e expanded only by culturing in a low-serum medium containing fibroblast growth factor(FGF)-2. Under the culture condition decreased with differentiation factors contained in a serum, undifferentiated stem cell population exhibited almost unlimited potential of proliferation and resulting cells showed high frequency of differentiation into adipocytes, osteoblasts and chondrocytes. We thus suggest that the stem cells in adipose tissues maintain their self-renewing cycle in low-serum medium retaining the potential of multiple differentiations. Recent impact among scientist studying ASC is that adipose tissues contain large amount of common progenitor for mesenchymal stem cell and angioblast (meso-angioblast). Fluorescence-activated cell sorter analysis showed such stem cells (CD34+) account for the major population of the cells in stromal vascular fraction dispersed from adipose tissues. Having large number of multi-potent stem cells in adipose tissues, the next question to be addressed is how ASCs remain quiescent without spontaneous proliferation and differentiation.We have ever shown that gene expression of laminin α4 chain increases depending on adipose conversion of 3T3-L1 fibroblasts. Laminin G-like domain (LG) 4 module of a4 chain has strong affinity for syndecans. Finding that α4 LG3-5 is cleaved from laminin-8 (α4β1γ1) in vivo, we explored its function as a ligand blocking FGF-2 signaling. In mouse de novo adipogenesis at the site of injection of Matrigel plus FGF-2,addition of α4 LG4-5 at 20 nM suppressed the adipogenesis completely. Activity of LG4-5 was cryptic within the whole structure of G domain since α4 LG1-5 showed only weak effect. Suppressive effect of α4 LG4 was lost in its mutants with reduced heparin-binding activity. Antibodies against extracellular (but not cytoplasmic) domain of syndecan-2 and 4 suppressed the adipogenesis. Thus, FGF-signaling via syndecans appeared to be blocked by α4 G domain fragments due to their competitive binding to syndecans. Less