Gene Expression Profiling of Adenosine Signaling in the Human Monocytic THP-1 Cells.

Abstract Inflammation, a host response to infection or injury, is usually beneficial to the host. However, uncontrolled or prolonged inflammation can cause tissue and organ damage, and possibly lead to diseases like atherosclerosis, rheumatoid arthritis, asthma, and cancers. Much attention has focused on pro-inflammatory signaling but little is known about the mechanisms that resolve inflammation (anti-inflammation). Adenosine, released from leukocytes and endothelial cells, is an endogenous anti-inflammatory mediator that down-regulates acute inflammation in part by modulating macrophage cytokine production. LPS (lipopolysaccharide) is a potent pro-inflammatory reagent of macrophages that acts primarily through Toll-like receptors (TLRs). Studies using murine cells have showed that co-stimulation of both adenosine receptors and TLRs greatly increases the production of vascular endothelial growth factor (VEGF). The mechanisms for this synergistic interaction are currently not known. It is also unknown whether the adenosine-LPS interaction occurs in human macrophages. To gain insights and to generate new hypotheses, we have utilized oligonucleotide microarrays to examine how adenosine alters global gene expression in the human monocytic THP-1 cell line. We analyzed differentially expressed genes induced by adenosine alone, by LPS alone, and by adenosine in the presence of LPS. The genes activated by adenosine are enriched in processes transcription, RNA modification and cellular defense response, but are lacking processes involved in lipid biosynthesis and protein catabolism. Our result shows that at the mRNA level adenosine alone induces a nine-fold increase of VEGF, and adenosine in the presence of LPS elevates VEGF expression three fold when compared to controls. Many genes with the greatest induction by adenosine in the presence of LPS have unknown functions in macrophages. The genes with known functions include transcription regulation (EGR1, HEY1, and CTBP1), nuclear receptors (NR4A2), immune response modulators (IL1A, STAT4, and INHBA), apoptosis facilitator (BCL2L11), signal transducers (PRPF4B, LNK, and ARHGEF7), and glutamate transporter (SLC1A2). These results suggest that the mechanisms by which adenosine mediates its anti-inflammatory signals are far more complex than simply regulating cytokine production. In addition, we found that three TLR signaling genes are uniquely suppressed by adenosine in the presence of LPS: CREB1, TLR1, and IRF2. CREB1 modulates transcription of cAMP signaling; TLR1 is part of the TLR2/TLR1 complex and can regulate TLR2 mediated pathways; IRF2 is a mediator of interferon signaling. We are in the process of verifying these observations using real-time PCR and independent RNA samples.