Tavassoli M, Takahashi K
Am J Anat. 1982 Jun;164(2):91-111. doi: 10.1002/aja.1001640202.
In long-term cultures of bone marrow, the adherent stromal cells provide support for the proliferation and maintenance of hemopoietic stem cells. These stromal cells and their interactions were characterized by means of scanning (SEM) and transmission (TEM) electron microscopy in correlation with functional studies. Cultures were initiated by establishing the adherent stromal layer as a "soil" which was then "seeded" after 3 weeks by the addition of another marrow-cell suspension. Clonal assay of the supernatant demonstrated the continuous proliferation of the hemopoietic stem cell. The stroma essentially consisted of two cell types, macrophages and epithelioid cells. Macrophages were smaller, 10-15 microns, phagocytosed latex and carbon particles, and contained lysosomes. Their surface did not stain with polycationic ferritin (PCF). Epithelioid cells were much larger, more than 100 microns; contained numerous thin, elongated mitochondria; did not phagocytose latex particles; but did display strong surface staining with PCF. The appearance of epithelioid cells in TEM depended on their state of development and whether the section was parallel or perpendicular to the substratum. Epithelioid cells displayed a maturational spectrum, at two ends of which were synthetic and storage phases. In the synthetic phase, the cell contained numerous profiles of rough endoplasmic reticulum, and in the storage phase, numerous storage granules. These two phases were best appreciated in sections perpendicular to the substratum, demonstrating synthetic cells on top settling over the substratum upon maturation into the storage cells. Both macrophages and epithelioid cells contained fat globules which increased in number and size with the addition of hydrocortisone to the culture medium. A distinct fat-cell type, as has been claimed, was not found in this study. Granulopoiesis was observed in the culture system in the absence of colony-stimulating activity in the supernatant, suggesting direct cellular interaction or short-range factors in the induction of granulopoiesis. Widespread cellular interactions were noted between macrophages and epithelioid cells, the latter often completely embracing the former and both extending cytoplasmic processes toward each other. This is reminiscent of the cooperative interaction of endoderm and mesoderm in chick embryo hemopoiesis and may be necessary for the maintenance of stem cells in these cultures.
在骨髓的长期培养中，贴壁基质细胞为造血干细胞的增殖和维持提供支持。通过扫描电子显微镜（SEM）和透射电子显微镜（TEM）对这些基质细胞及其相互作用进行了表征，并与功能研究相关联。培养通过建立贴壁基质层作为“土壤”开始，然后在3周后通过添加另一种骨髓细胞悬液进行“播种”。上清液的克隆分析证明了造血干细胞的持续增殖。基质主要由两种细胞类型组成，即巨噬细胞和上皮样细胞。巨噬细胞较小，10 - 15微米，吞噬乳胶和碳颗粒，并含有溶酶体。它们的表面不会被聚阳离子铁蛋白（PCF）染色。上皮样细胞要大得多，超过100微米；含有许多细长的线粒体；不吞噬乳胶颗粒；但确实显示出强烈的PCF表面染色。上皮样细胞在TEM中的外观取决于它们的发育状态以及切片是平行还是垂直于基质。上皮样细胞呈现出成熟谱，其两端分别是合成期和储存期。在合成期，细胞含有大量粗面内质网轮廓，而在储存期，含有大量储存颗粒。这两个阶段在垂直于基质的切片中最易观察到，显示出顶部的合成细胞在成熟为储存细胞时沉降在基质上。巨噬细胞和上皮样细胞都含有脂肪球，随着向培养基中添加氢化可的松，脂肪球的数量和大小都会增加。在本研究中未发现如所声称的独特脂肪细胞类型。在培养系统中，即使上清液中没有集落刺激活性，也观察到了粒细胞生成，这表明在粒细胞生成的诱导中存在直接的细胞间相互作用或短程因子。注意到巨噬细胞和上皮样细胞之间存在广泛的细胞间相互作用，后者常常完全包围前者，并且两者都向彼此伸出细胞质突起。这让人想起鸡胚造血过程中内胚层和中胚层的协同相互作用，并且可能对于维持这些培养物中的干细胞是必要的。
