Rossette formation between stromal and hemopoietic cells: a simple assay for the supportive activity of stromal cells

Hemopoietic stromal cells play an important role in the proliferation and differentiation of hemopoietic cells in vitro. Recently, it has been emphasized that the attachment of hemopoietic cells to stromal cells is the first step in this process and that this is necessary in hemopoiesis in vitro. A rosette formation technique was developed to quantitatively measure this attachment. The murine leukemic cell line ELM-D, which grows only in close contact with stromal cells, was used to represent hemopoietic cells. Rosettes were formed with about 75% of the normal murine bone marrow derived hemopoietic supportive stromal cells (MS-1). However, ELM-D formed rosettes with only 15% of the hemopoietic non-supportive stromal cells (MS-K). The maximum rosette formation was obtained at an ELM-D to MS-1 ratio of 20:1. When MS-1 and MS-K were mixed and seeded as stromal cells, MS-K did not alter the attachments of ELM-D to MS-1; the percentage of rosette formation increased in parallel with the increase in the number of MS-1 cells in the stromal layer. These findings suggest that MS-K does not have an inhibitory effect. Rosette formation is a simple, quantitative technique for assaying the hemopoietic supportive activity of stromal cells in vitro.