Regulation of interleukin-11 protein and mRNA expression in neonatal and adult fibroblasts and endothelial cells

Abstract

Interleukin-11 (IL-11), a newly-identified cytokine produced by stromal cells, elevates platelet counts in neonatal rats in vivo and synergizes in vitro with IL-3 in supporting murine megakaryocyte colony formation and stimulating hematopoietic stem cells. Megakaryocytopoiesis is also enhanced by other colony-stimulating factors (CSFs), including IL-3, IL- 6, and Steel factor (SLF). Dysregulation of neonatal thrombopoiesis predisposes newborns to develop thrombocytopenia during sepsis, despite increased circulating pools of committed thrombopoietic progenitors in newborn cord blood compared with adult. We previously reported reduced expression of granulocyte-macrophage colony-stimulating factor (GM- CSF), granulocyte-colony-stimulating factor (G-CSF), and IL-3 from stimulated cord mononuclear cells, but increased expression of SLF in human umbilical vein endothelial cells (HUVEC). Therefore, we hypothesized that IL-3, IL-6, and SLF might modulate megakaryocytopoiesis by inducing IL-11 expression, and newborns might express altered levels of IL-11 mRNA expression during activated conditions, contributing to the difference in circulating colony- forming unit-megakaryocyte (CFU-Meg) cord and adult blood. Phorbol myristate acetate (PMA) induced a twofold greater increase in IL-11 mRNA expression in neonatal fibroblasts (NFb) compared with adult fibroblasts (AFb), and a 3.6-fold greater increase in HUVEC than human adult aorta endothelial cells (HAEC) by Northern blot analysis. PMA also induced a threefold greater increase in IL-11 protein production in NFb than AFb. Physiologic agonists IL-1 alpha, transforming growth factor-beta 1 (TGF-beta 1), and TGF-beta 2 triggered upregulation of IL- 11 mRNA expression in both NFb and AFb. However, IL-3, IL-6, PIXY321 (a GM-CSF-IL-3 fusion protein), and SLF failed to upregulate IL-11 mRNA expression from the basal level, while macrophage-colony stimulating factor (M-CSF) mRNA was significantly induced. These data suggest that the hematopoietic effect of IL-6, SLF, and IL-3 on megakaryocytopoiesis is probably not mediated by secondary IL-11 mRNA expression. Similarly, inflammatory agonists IL-1 beta, lipopolysaccharide (LPS), and tumor necrosis factor-alpha (TNF-alpha) alone did not upregulate IL-11 expression from the basal level in endothelial cells, whereas intracellular adhesion molecule-1 (ICAM-1) and endothelial leukocyte adhesion molecule-1 were strongly induced. Minimal basal IL-11 expression was detected by reverse transcriptase-polymerase chain reaction (RT-PCR) in NFb, AFb, HUVEC and HAEC. The quantitative RT-PCR assay also verified that IL-1 beta and TNF-alpha-stimulated HUVEC and HAEC, and IL-3- and IL-6-stimulated NFb and AFb only expressed minimal levels of IL-11 mRNA.(ABSTRACT TRUNCATED AT 400 WORDS)