FUNCTIONALLY DISTINCT HUMAN MARROW STROMAL CELL-LINES IMMORTALIZED BYTRANSDUCTION WITH THE HUMAN PAPILLOMA-VIRUS E6 E7 GENES/

A replication-defective recombinant retrovirus containing the human pa pilloma virus E6/E7 genes (LXSN-16 E6E7) was used to immortalize strom al cells from human marrow. The E6/E7 gene products interfere with the function of tumor-suppressor proteins p53 and Rb, respectively, there by preventing cell cycle arrest without causing significant transforma tion. Twenty-seven immortalized clones designated HS-1 to HS-27 were i solated, four of which are characterized in this report. Two cell line s, HS-5 and HS-21, appear to be fibroblastoid and secrete significant levels of granulocyte colony-stimulating factor (G-CSF), granulocyte-m acrophage-CSF (GM-CSF), macrophage-CSF (M-CSF), Kit ligand (KL), macro phage-inhibitory protein-1 alpha, interleukin-6 (IL-6), IL-8, and IL-1 1. However, only HS-5 supports proliferation of hematopoietic progenit or cells when cocultured in serum-deprived media with no exogenous fac tors. Conditioned media (CM) from HS-5 promotes growth of myeloid colo nies to significantly greater extent than a cocktail of recombinant fa ctors containing 10 ng/mL of IL-1, IL-3, IL-6, G-CSF, GM-CSF, and KL a nd 3 U of erythropoietin (Epo). Two additional clones, HS-23 and HS-27 , resemble ''blanket'' cells, with an epithelioid morphology, and are much larger, broader, and flatter when compared with HS-5 and HS-21. T hese lines secrete low levels of growth factors and do not support pro liferation of isolated progenitor cells in cocultures. CM from HS-23 a nd HS-27 also fail to support growth of myeloid colonies. Both HS-23 a nd HS-27 express relatively high levels of VCAM-1, yet HS-27 is the on ly line that supports the formation of ''cobblestone'' areas by isolat ed CD34(+)38(lo) cells. We hypothesize that HS-5, HS-21, HS-23, and HS -27 represent functionally distinct components of the marrow microenvi ronment. (C) 1995 by The American Society of Hematology.