Adenosine and the adenosine A2A receptor agonist, CGS21680, upregulate CD39 and CD73 expression through E2F-1 and CREB in regulatory T cells isolated from septic mice

Abstract

The number of regulatory T cells (Treg cells) and the expression of ectonucleoside triphosphate diphosphohydrolase 1 (ENTPD1; also known as CD39) and 5'-ectonucleotidase (NT5E; also known as CD73) on the Treg cell surface are increased during sepsis. In this study, to determine the factors leading to the high expression of CD39 and CD73, and the regulation of the CD39/CD73/adenosine pathway in Treg cells under septic conditions, we constructed a mouse model of sepsis and separated the Treg cells using a flow cytometer. The Treg cells isolated from the peritoneal lavage and splenocytes of the mice were treated with adenosine or the specific adenosine A2A receptor agonist, CGS21680, and were transfected with specific siRNA targeting E2F transcription factor 1 (E2F-1) or cyclic adenosine monophosphate (cAMP) response element-binding protein (CREB), which are predicted transcription regulatory factors of CD39 or CD73. The regulatory relationships among these factors were then determined by western blot analysis and dual-luciferase reporter assay. In addition, changes in adenosine metabolism were measured in the treated cells. The results revealed that adenosine and CGS21680 significantly upregulated CD39 and CD73 expression (P<0.01). E2F-1 and CREB induced CD39 and CD73 expression, and were upregulated by adenosine and CGS21680. Adenosine triphosphate (ATP) hydrolysis and adenosine generation were inhibited by the knockdown of E2F-1 or CREB, and were accelerated in the presence of CGS21680. Based on these results, it can be inferred that adenosine, the adenosine A2A receptor agonist, E2F-1 and CREB are the possible factors contributing to the high expression of CD39 and CD73 on the Treg cell surface during sepsis. Adenosine and its A2A receptor agonist served as the signal transducer factors of the CD39/CD73/adenosine pathway, accelerating adenosine generation. Our study may benefit further research on adenosine metabolism for the treatment of sepsis.

Identification of key pathogenic genes of sepsis based on the Gene Expression Omnibus database

“…To verify the results of the bioinformatics analyses, the expression levels of selected DEGs were quantified by RT-qPCR in three sepsis and three healthy blood samples. A total of 6 DEGs, including IRAK3, adrenomedullin (ADM), arachidonate 5-lipoxygenase (ALOX5), matrix metallopeptidase 9 (MMP9), S100A8 and ectonucleoside triphosphate diphosphohydrolase 1 (ENTPD1) were selected as candidate genes based on the previous literature reviews ( 1 , 17 – 21 ). IRAK3 (P<0.01), ADM (P<0.05), ALOX5 (P<0.001), MMP9 (P<0.05) and S100A8 (P<0.05) were significantly upregulated in the blood samples of patients with sepsis compared with those of healthy controls ( Fig.…”

“…Sepsis is a life-threatening condition that is caused by the entry of bacteria, fungi, viruses or parasites to the bloodstream, which subsequently triggers a systemic uncontrolled inflammatory response ( 1 ). Sepsis is be divided into three classifications with increasing severity: Sepsis, severe sepsis and septic shock ( 2 ).…”

Effects of Caffeine on Behavioural and Cognitive Deficits in Rats

“…Adenosine has strong immunosuppressive effects, many of which are mediated by A2A receptors expressed on immune cells. The activation of adenosine A2A receptor inhibits the proliferation of T cells and production of proinflammatory cytokines, which contributes to the activation of the synthesis of anti‐inflammatory cytokines, thereby suppressing the systemic response .…”

Purinergic signalling in liver diseases: Pathological functions and therapeutic opportunities

“…30 With regard to Tregs, adenosine promotes the generation and expansion of Tregs, induces immunosuppressive functions, and upregulates CD39 and CD73 expression. [31][32][33][34] Adenosine terminates macrophage activation via A2A/NF-jBinduced reduction of tumour necrosis factor (TNF)-a expression 35 and inhibits macrophage colony-stimulating factor-dependent proliferation of macrophages by inducing A2B/cAMP/p27 kip-1 expression. 36 Moreover, adenosine also inhibits the cytotoxic activity and cytokine production of NK cells 37 and exerts immunosuppressive effects on DCs, including reducing interleukin (IL)-12, TNF-a, C-X-C motif chemokine ligand (CXCL)-10, C-C motif chemokine ligand (CCL)-2, and CCL-12 secretion, as well as decreasing major histocompatibility complex (MHC) class II expression, and impairing allogenic T cell proliferation via the cAMP/protein kinase A (PKA) signalling pathway.…”