Ethyl Pyruvate Modulates Murine Dendritic Cell Activation and Survival Through Their Immunometabolism

Abstract

Attenuating the innate immunity activation could ameliorate inflammation and disease in settings such as transplant rejection or autoimmunity. Recently, a pivotal role for metabolic re-programming in TLR-induced dendritic cell (DC) activation has emerged. Ethyl pyruvate (EP), a pyruvate derivative, possesses anti-inflammatory properties in vitro and in animal models of disease. However, its effects on DCs remain elusive. We found that EP attenuated LPS-induced activation of murine GM-CSF bone marrow-derived dendritic cells (DCs) in vitro, reducing pro-inflammatory cytokine and IL-10 production, costimulatory molecule and MHC expression, the type I Interferon (IFN-I) response, the LPS-induced cell death, and the ability of DCs to stimulate allogeneic T cells. DC activation induced by TLR7 and TLR9 ligands was also suppressed by EP in vitro. Finally, EP decreased TLR-induced activation stimulated in vivo in conventional DCs and inflammatory monocytes. Investigating EP mechanisms, we found that EP decreased glycolysis and mitochondrial respiration, upon and in absence of TLR stimulation, by reducing ERK, AKT, and nitric oxide (NO) activation. These results indicate that EP inhibits most of the DC biological responses to TLR triggering, altering the metabolic reprogramming necessary for DC activation.

Generating Bovine Monocyte-Derived Dendritic Cells for Experimental and Clinical Applications Using Commercially Available Serum-Free Medium

“…This is in contrast to LPS-stimulated DC models showing a decrease in basal respiration, but is in agreement with other studies utilizing "cocktails" to stimulate DC maturation, suggesting the response may be stimulation-method dependent (73)(74)(75)(76). PMA+I stimulated bovine MoDC also showed a decrease in coupling efficiency but an increase in proton leak and mitochondrial ATP production compared to unstimulated MoDC, and is consistent with one study in which DC were stimulated with LPS (74). Phorbol esters (including PMA) are distinguished from cytokine-and TLR-induced DC activation in their significant formation of reactive oxygen species (ROS), which may have implications in complex mitochondrial metabolic pathways (67).…”

“…Notably, stimulated bovine MoDC exhibited a significant increase in basal mitochondrial respiration compared to unstimulated MoDC and was also observed for serum-free and serum-supplemented cultures. This is in contrast to LPS-stimulated DC models showing a decrease in basal respiration, but is in agreement with other studies utilizing "cocktails" to stimulate DC maturation, suggesting the response may be stimulation-method dependent (73)(74)(75)(76). PMA+I stimulated bovine MoDC also showed a decrease in coupling efficiency but an increase in proton leak and mitochondrial ATP production compared to unstimulated MoDC, and is consistent with one study in which DC were stimulated with LPS (74).…”

“…Essentially, mitochondrial reprogramming involves a rapid switch of ATP production from mitochondrial oxidative phosphorylation to aerobic glycolysis and is necessary to support the generation of new bioenergetic and metabolic resources and for DC survival (73). Previous studies have demonstrated transient increases in mitochondrial activity followed by subsequent mitochondrial collapse after DC stimulation, including TLR-stimulation (73)(74)(75)(76). Consistent with these studies, bovine MoDC stimulated for 16 h with PMA+I and cultured in either AV-SF or RP-S medium, displayed an increased glycolytic rate compared to unstimulated MoDC, evidenced by a consistent increase in extracellular acidification rate (ECAR).…”

Ethyl Pyruvate Promotes Proliferation of Regulatory T Cells by Increasing Glycolysis

“…Additionally, recent data suggest that in vitro differentiation of Treg depends on both glycolysis and FAO [ 12 ]. Our results with EP contradict the findings made by Chakhtoura et al, 2019 [ 27 ], where EP application down-regulated glycolysis in dendritic cells. In spite of this, it is firmly established that pyruvate conversion to lactate transforms NADH into NAD+, thereby maintaining the large NAD+/NADH ratio necessary for driving glycolysis [ 28 ].…”

“…In addition, the proliferating capacity of dendritic cells and Treg is different, and EP effects may depend upon the activation status of the particular cell. For example, according to reference [ 27 ], EP inhibited glycolysis when applied to differentiated dendritic cells that are mainly quiescent. In contrast, in this study EP stimulated glycolysis but was applied to proliferating Treg suggesting that EP may engage different pathways depending on the cell type and their division status.…”