Trevisan M, Iscove N N
Ontario Cancer Institute, University of Toronto, Canada.
J Exp Med. 1995 Jan 1;181(1):93-103. doi: 10.1084/jem.181.1.93.
Early hemopoietic precursors have been extensively studied using short-term assays based on colony formation or in vivo reconstitution that do not run beyond a few weeks. However, little information is available on the phenotype of the stem cells that are detectable in 6-12-mo transplantation assays, and their relationship to cells detected in short-term assays is not known. In this study, we investigated the phenotype and separability by cell sorting of a spectrum of hemopoietic precursor cells in normal adult mouse marrow, including cells quantitated in a 1 yr competitive transplantation assay in vivo as well as in short-term colony assays in vitro and in vivo. Two principal findings emerged. The first was that cells detected in a variety of short-term assays--CFU-S12 (spleen colony-forming cells), CFCmulti (multilineage colony-forming cells), pre-CFCmulti (precursors of CFCmulti), CFC-E/Mg (erythroid/megakaryocyte CFC) and CFC-G/M (granulocyte/macrophage CFC)--were phenotypically similar and could not be separated from one another using a panel of markers useful in segregating them from more differentiated cells, including buoyant density, sedimentation velocity, adhesiveness to plastic, light scatter, high rhodamine-123 retention, and expression of surface wheat-germ agglutinin (WGA)-binding carbohydrate, H-2K, CD45, AA4.1, heat stable antigen (HSA), CD71, and Ly6A/Sca-1 antigens. Long-term reconstituting (LTR) cells quantitated in vivo differed little from the other precursors in expression of many of the above markers. However, they differed somewhat in lower sedimentation velocity and lower expression of WGA-binding surface carbohydrate, and most strikingly in their conditional adhesiveness to plastic, very low retention of Rh123 and high level expression of Ly6A/Sca-1, to a degree that would permit the quantitative separation of the two precursor classes from each other. The results provide a comprehensive characterization of LTR cells measured to 12 mo in vivo and a direct and quantitative analysis of their separation from cells detected in colony assays.
早期造血前体细胞已通过基于集落形成或体内重建的短期检测方法进行了广泛研究，这些检测方法持续时间不超过几周。然而，关于在6至12个月移植检测中可检测到的干细胞表型，以及它们与短期检测中检测到的细胞之间的关系，所知甚少。在本研究中，我们通过细胞分选研究了正常成年小鼠骨髓中一系列造血前体细胞的表型和可分离性，包括在1年体内竞争性移植检测中定量的细胞，以及体外和体内短期集落检测中的细胞。出现了两个主要发现。第一个发现是，在各种短期检测中检测到的细胞——CFU-S12（脾集落形成细胞）、CFCmulti（多谱系集落形成细胞）、pre-CFCmulti（CFCmulti的前体细胞）、CFC-E/Mg（红系/巨核系CFC）和CFC-G/M（粒系/巨噬系CFC）——在表型上相似，并且使用一组有助于将它们与更分化细胞分离的标志物无法彼此分离，这些标志物包括浮力密度、沉降速度、对塑料的粘附性、光散射、高罗丹明-123保留率，以及表面小麦胚凝集素（WGA）结合碳水化合物、H-2K、CD45、AA4.1、热稳定抗原（HSA）、CD71和Ly6A/Sca-1抗原的表达。在体内定量的长期重建（LTR）细胞在上述许多标志物的表达上与其他前体细胞差异不大。然而，它们在沉降速度较低和WGA结合表面碳水化合物表达较低方面有所不同，最显著的是它们对塑料的条件性粘附性、Rh123保留率极低以及Ly6A/Sca-1高水平表达，在一定程度上允许将这两类前体细胞彼此定量分离。这些结果提供了在体内测量至12个月的LTR细胞的全面特征，以及对它们与集落检测中检测到的细胞分离的直接和定量分析。
