Induction of thrombospondin-1 by all-trans retinoic acid modulates growth and differentiation of HL-60 myeloid leukemia cells

Thrombospondin-1 (TSP), a multifunctional extracellular matrix protein, modulates human hematopoietic stem cell adherence and thus may play a role in blood cell proliferation and/or differentiation. The expression of TSP was studied in the human myeloid leukemia cell line, HL-60, upon differentiation into monocytes by phorbol-13-monoacetate (PMA) or into granulocytes by all-trans retinoic acid (RA). HL-60 cells cultured under serum-free conditions constitutively secreted low amounts of TSP into the cultured medium, approximately 13 ng/10(6) cells/24 h. PMA used at 4 x 10(-8) M did not significantly modulate TSP secretion over a 24 h period. In contrast, RA at 10(-7) M induced a 5- to 10-fold increase in TSP secreted by HL-60 cells during their differentiation into granulocytes over a 5 day period. The role of secreted TSP in RA-dependent cessation of growth and differentiation was examined using blocking anti-TSP antibodies. In the presence of the polyclonal anti-TSP antibody R5 (25 microg/ml), growth of RA-treated HL-60 cells was maintained at control levels for up to 3 days and a concomitant delay in granulocytic differentiation was observed. Moreover, the addition of soluble TSP (0.5-5 microg/ml) to untreated HL-60 cells decreased their growth and promoted their differentiation in a dose-dependent manner. Using a neutralizing antibody to transforming growth factor beta (TGF-beta) or purified TGF-beta1 we further demonstrated that the effects of TSP were not mediated through activation of latent TGF-beta. These studies indicate that TSP decreases the proliferation and promotes the differentiation of HL-60 cells.