The Regulatory Roles of Ezh2 in Response to Lipopolysaccharide (LPS) in Macrophages and Mice with Conditional Ezh2 Deletion with LysM-Cre System

Abstract

The responses of macrophages to lipopolysaccharide (LPS) might determine the direction of clinical manifestations of sepsis, which is the immune response against severe infection. Meanwhile, the enhancer of zeste homologue 2 (<i>Ezh2</i>), a histone lysine methyltransferase of epigenetic regulation, might interfere with LPS response. Transcriptomic analysis on LPS-activated wild-type macrophages demonstrated an alteration of several epigenetic enzymes. Although the <i>Ezh2</i>-silencing macrophages (RAW264.7), using small interfering RNA (siRNA), indicated a non-different response to the control cells after a single LPS stimulation, the <i>Ezh2</i>-reducing cells demonstrated a less severe LPS tolerance, after two LPS stimulations, as determined by the higher supernatant TNF-α. With a single LPS stimulation, <i>Ezh2</i> null (<i>Ezh2</i>^flox/flox; LysM-Cre^cre/-) macrophages demonstrated lower supernatant TNF-α than <i>Ezh2</i> control (<i>Ezh2</i>^fl/fl; LysM-Cre<i>^-/-</i>), perhaps due to an upregulation of <i>Socs3</i>, which is a suppressor of cytokine signaling 3, due to the loss of the <i>Ezh2</i> gene. In LPS tolerance, <i>Ezh2</i> null macrophages indicated higher supernatant TNF-α and IL-6 than the control, supporting an impact of the loss of the <i>Ezh2</i> inhibitory gene. In parallel, <i>Ezh2</i> null mice demonstrated lower serum TNF-α and IL-6 than the control mice after an LPS injection, indicating a less severe LPS-induced hyper-inflammation in <i>Ezh2</i> null mice. On the other hand, there were similar serum cytokines after LPS tolerance and the non-reduction of serum cytokines after the second dose of LPS, indicating less severe LPS tolerance in <i>Ezh2</i> null mice compared with control mice. In conclusion, an absence of <i>Ezh2</i> in macrophages resulted in less severe LPS-induced inflammation, as indicated by low serum cytokines, with less severe LPS tolerance, as demonstrated by higher cytokine production, partly through the upregulated <i>Socs3</i>.