Regulation of Hematopoietic Growth Factor Production by Genetically Modified Human Bone Marrow Stromal Cells Expressing Interleukin-1 β Antisense RNA

Abstract

Interleukin-1 (IL-1) plays a major role in the regulation of bone marrow stromal cell function and hematopoiesis. It is known to induce secretion of the hematopoietic growth factors granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage CSF (GM-CSF), IL-6, and IL-8 as well as IL-1 itself in stromal cells. We investigated the role of IL-1β-mediated growth factor production in the human stromal cell line L88/5. Using liposome-mediated DNA transfer, two stromal cell transfectants that constitutively express IL-1β antisense (AS) RNA were generated. Expression of IL-1β AS RNA and IL-1β RNA was determined by RT-PCR. The stromal cell transfectants were strongly impaired in their endogenous IL-1β production, and this effect was present even when strong IL-1β inducers, such as IL-1α and tumor necrosis factor-α (TNF-α), were used. Reduced endogenous IL-1β levels had no effect on the constitutive production of IL-6, IL-8, and GM-CSF measured by ELISA. In contrast to lipopolysaccharide (LPS) stimulation, IL-1α-mediated stimulation of GM-CSF production was significantly reduced in AS transfectants. TNF-α induced GM-CSF production was also reduced. IL-6 and IL-8 production was increased in transfectants, suggesting a negative regulatory role of IL-1β in L88/5. This new approach using AS technology to specifically target constitutive RNA expression will allow further characterization of the bone marrow cytokine network in normal and malignant hematopoiesis.