Hemopoiesis in long-term stroma-dependent cultures from lymphoid tissue: production of cells with myeloid/dendritic characteristics

Abstract

A long-term stroma-dependent culture system (LTC) has been developed which continuously produces hemopoietic cells providing an in vitro system for the study of cell differentiation. These nonadherent cell populations contain a large subpopulation of dendritic cells (DC). LTC producing DC were easily generated from spleen, but could also be established from bone marrow (BM) and lymph node with less success. It was difficult to establish DC-producing LTC from thymus. The properties of splenic and thymic stroma have been compared. Spleen stroma developed more complicated networks of fibroblasts, endothelial cells, macrophages, and DC. Thymic stromal monolayers were dominated by epithelial cells and fibroblasts, with a lower proportion of macrophages and endothelial cells. They had a relatively sparse structure of cell networks compared with spleen stroma. Cells with dendritiform morphology first appeared in cultures by 2-3 wk. The majority of cells produced were large cells which expressed DC-specific cell surface markers, major histocompatibility complex (MHC) Class II molecules, and the CD80/CD86(B7) costimulator. A high proportion of cells also expressed myeloid cell markers. No T or B lymphoid cells or granulocytes were present in the cultures. LTC continued to produce nonadherent cells resembling myeloid/DC for long periods, even after passage of stromal cells and stem cells at about 3-4 mo. after culture establishment. The LTC system offers potential to study the in vitro differentiation of myeloid/DC.