Molecular study of hematopoiesis-supporting ability of C3H10T1/2 mouse embryo fibroblasts

C3H10T1/2小鼠胚胎成纤维细胞造血支持能力的分子研究

基本信息

批准号：
06454345
负责人：
OZAWA Keiya
金额：
$ 3.58万
依托单位：
Jichi Medical School
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1994
资助国家：
日本
起止时间：
1994 至 1995
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-06454345/
关键词：
Stromal cell Preadipocyte C3H10T1 2 cell Differentiation induction Hematopoiesis-supporting ability CD34

项目摘要

To investigate the molecular aspects of hematopoiesis-supporting ability of stromal cells, we used a differentiation-inducible mouse embryo fibroblast cell line, C3H10T1/2 (10T1/2). Stably determined preadipocyte and myoblast cell lines were established after a brief exposure of 10T1/2 cells to 5-azacytidine. These cell lines terminally differentiated into adipocytes and myotubes, respectively, under appropriate conditions. The hematopoiesis-supporting ability was significantly elevated at the preadipocyte stage (A54 preadipocyte), and was reduced after terminal adipocytic differentiation. To identify molecules that contribute to the hematopoiesis-supporting ability of preadipocytes, we screened genes that were differentially expressed in A54 preadipocytes and isolated the novel gene by mRNA differential display method. This gene was defined as a gene that was down-regulated during adipocyte differentiation-1 (drad-1). The drad-1 was expressed in other mouse preadipocytes, namely, ST2 and PA6 cells, that also have hematopoiesis-supporting ability. Moreover, the drad-1 was found to be expressed in mouse bone marrow. However, the function of the protein encoded by drad-1 is currently unknown. In addition, we investigated the change in CD34 mRNA expression during stromal cell differentiation. As a result, CD34 mRNA was constitutively expressed by parent 10T1/2 cells but not by adipogenically or myogenically determined cells. This finding supports the concept that CD34 may also be a marker of stromal progenitors and is lost as the cells differentiate into phenotypically distinct stromal elements. The 10T1/2-derived cell lines could provide a valuable tool to aid in the analysis of stromal cell development and differentiation and the search for novel stromal cell-derived factor (s).

为了研究基质细胞造血支持能力的分子方面，我们使用了分化诱导型小鼠胚胎成纤维细胞系 C3H10T1/2 (10T1/2)。将10T1/2细胞短暂暴露于5-氮杂胞苷后，建立了稳定测定的前脂肪细胞和成肌细胞系。在适当的条件下，这些细胞系分别最终分化为脂肪细胞和肌管。造血支持能力在前脂肪细胞阶段(A54前脂肪细胞)显着升高，并在终末脂肪细胞分化后降低。为了鉴定有助于前脂肪细胞造血支持能力的分子，我们筛选了A54前脂肪细胞中差异表达的基因，并通过mRNA差异展示方法分离了新基因。该基因被定义为在脂肪细胞分化-1（drad-1）过程中下调的基因。 drad-1在其他小鼠前脂肪细胞（即ST2和PA6细胞）中表达，这些细胞也具有造血支持能力。此外，发现drad-1在小鼠骨髓中表达。然而，drad-1编码的蛋白质的功能目前尚不清楚。此外，我们研究了基质细胞分化过程中 CD34 mRNA 表达的变化。结果，CD34 mRNA 由亲代 10T1/2 细胞组成型表达，但由脂肪生成或肌生成决定的细胞不表达。这一发现支持这样的概念：CD34 也可能是基质祖细胞的标记，并且随着细胞分化成表型不同的基质元件而丢失。 10T1/2 衍生的细胞系可以提供有价值的工具来帮助分析基质细胞的发育和分化以及寻找新的基质细胞衍生因子。