Transketolase and vitamin B1 influence on ROS-dependent neutrophil extracellular traps (NETs) formation

Abstract

Neutrophil extracellular traps (NETs) are a recently identified, web-like, extracellular structure composed of decondensed nuclear DNA and associated antimicrobial granules. NETs are extruded into the extracellular environment via the reactive oxygen species (ROS)-dependent cell death pathway participating in inflammation and autoimmune diseases. Transketolase (TKT) is a thiamine pyrophosphate (vitamin B1)-dependent enzyme that links the pentose phosphate pathway with the glycolytic pathway by feeding excess sugar phosphates into the main carbohydrate metabolic pathways to generate biosynthetic reducing capacity in the form of NADPH as a substrate for ROS generation. In this work, TKT was selected as a lead candidate from 24 NET-associated proteins obtained by literature screening and knowledge gap assessment. Consequently, we determined whether TKT influenced NET formation in vitro . We firstly established that the release of ROS-dependent NETs was significantly decreased after purified human PMNs were pretreated with oxythiamine, a TKT inhibitor, and in a concentration dependent manner. As a cofactor for TKT reaction, we evaluated the release of NET formation either in vitamin B1 treatment or in combined use of oxythiamine and vitamin B1, and found that those treatments also exerted a significant suppressive effect on the amount of NET-DNA and ROS production. The regulation of TKT by oxythiamine and/or vitamin B1 may therefore be associated with response to the modulation of NET formation by preventing generation of excessive NETs in inflammatory diseases.

The Non-canonical Role of Metabolic Enzymes in Immune Cells and Its Impact on Diseases

“…Also, in Staphylococcus aureus-infected PMNs, the inhibition of TKT led to a reduction of ROS production and NETosis [186]. Altogether, this evidence indicates that inhibition of TKT activity impairs the production of NADPH and, by consequence, diminishes NET formation in PMNs [186,190].…”

“…On the other hand, when TKT was blocked, in a dose-dependent manner, a reduction in the levels of ROS and of NET formation in PMA-stimulated PMNs was observed [186]. Also, in Staphylococcus aureus-infected PMNs, the inhibition of TKT led to a reduction of ROS production and NETosis [186]. Altogether, this evidence indicates that inhibition of TKT activity impairs the production of NADPH and, by consequence, diminishes NET formation in PMNs [186,190].…”

“…TKT is widely expressed in most tissues, and some studies revealed that this enzyme can be located in the cytoskeleton of polymorphonuclear (PMN) neutrophils [185]. Interestingly, Riyapa et al [186] demonstrated that the expression of TKT transcripts is elevated in PMNs and monocytes when compared with lymphocytes and natural killer cells. Besides phagocytosis and degranulation, PMNs present a third antimicrobial mechanism that consists of the release of nuclear material associated with proteins from the granules in the form of neutrophil extracellular traps (NETs) [187].…”

“…Some studies demonstrated that NETosis, the process for producing NETs, is stimulated by the production of ROS due to the generation of free radicals and that PPP contributes to the NET formation by the generation of NADPH by G6PD [55,188,189]. Indeed, human's PMNs treated with phorbol myristate acetate (PMA), a positive control for NET formation, the protein level of TKT, were increased [186]. On the other hand, when TKT was blocked, in a dose-dependent manner, a reduction in the levels of ROS and of NET formation in PMA-stimulated PMNs was observed [186].…”

“…Indeed, human's PMNs treated with phorbol myristate acetate (PMA), a positive control for NET formation, the protein level of TKT, were increased [186]. On the other hand, when TKT was blocked, in a dose-dependent manner, a reduction in the levels of ROS and of NET formation in PMA-stimulated PMNs was observed [186]. Also, in Staphylococcus aureus-infected PMNs, the inhibition of TKT led to a reduction of ROS production and NETosis [186].…”