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

在过去几年中，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。自我更新的干细胞已经在各种成人组织中得到证实，它们显示出沿着意想不到的途径分化的潜力，不仅产生相同发育谱系的细胞，也产生干细胞来源的不同谱系的细胞。典型的例子是骨髓来源的干细胞，它可以在体外分化成成骨细胞、软骨细胞、脂肪细胞和成肌细胞，并且可以在体内并入骨骼、骨骼肌、肺、肠道和中枢神经系统。成体干细胞的另一个例子是脂肪源性干细胞（ASC），它可以沿间充质谱系分化产生脂肪细胞、软骨细胞、肌细胞和成骨细胞，也可以沿神经元谱系分化。在本研究中，我们发现从脂肪组织中分离出来的基质血管部分（SVF）中未分化的干细胞群体只有在含有成纤维细胞生长因子(FGF)-2的低血清培养基中培养才能扩增。在血清中含有的分化因子减少的培养条件下，未分化的干细胞群体表现出几乎无限的增殖潜力，分化成脂肪细胞、成骨细胞和软骨细胞的频率很高。因此，我们认为脂肪组织中的干细胞在低血清培养基中维持其自我更新周期，保留了多重分化的潜力。近年来科学家对ASC的研究发现，脂肪组织中含有大量间充质干细胞和成血管细胞的共同祖细胞。荧光活化细胞分选分析显示，这些干细胞（CD34+）占脂肪组织分散的间质血管部分细胞的主要群体。在脂肪组织中有大量的多能干细胞，下一个要解决的问题是ASCs如何保持静止而没有自发增殖和分化。我们已经证明层粘连蛋白α4链基因表达的增加依赖于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模块对分缩酶具有很强的亲和力。我们在体内发现α4 LG3-5是从层粘连蛋白-8 （α4β1γ1）中分离出来的，并探索其作为阻断FGF-2信号传导的配体的功能。在注射Matrigel + FGF-2部位的小鼠新生脂肪形成中，在20 nM处添加α4 LG4-5完全抑制脂肪形成。lg5 -5的活性在整个G结构域内是隐性的，因为LG1-5的α4只表现出微弱的作用。α4 LG4的抑制作用在其突变体中丧失，肝素结合活性降低。针对syndecan-2和4的细胞外（而不是细胞质）结构域的抗体抑制脂肪形成。因此，通过syndecans的fgf信号传导似乎被α 4g结构域片段阻断，因为它们与syndecans具有竞争性结合。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