The effects of ethyl pyruvate on NLRP3 inflammasome activation in microglial cells

Inflammasome activation is a process that begins and develops differently from classical inflammatory responses. For inflammasome activation, a danger signal must first reach the macrophage or microglia cell. In order to induce inflammasome activation, there must be a danger signal detected by the macrophage or microglia cell. This signal either can be named damage-associated molecular pattern (DAMP) or pathogen-associated molecular pattern (PAMP). When inflammasome mediators are active, cytokines such as interleukin-1β and interleukin-18 (IL-18) are released via caspase-1. Regulation and suppression of inflammasome activation is important because cell death can occur when inflammatory activity is present. Ethyl pyruvate (EP) is a molecule with anti-inflammatory and pro metabolic effects. EP has been shown to improve clinical presentation of neurodegenerative diseases such as Alzheimer's and Parkinson's. It was also found that BV2 inhibits inflammasome activation in the microglial cell line. In our study, it was aimed to investigate the effect of EP on N9 microglial cell line for inflammasome activation. In our study, inflammasome activation of protein 3 (NLRP3) containing NAD-like receptor family pyrine domain by Lipopolysaccharide (LPS) and ATP administration to N9 microglial cells was established. First, it will play an active role in the inflammasome activation of EP and most importantly it has been started by determining the dose not toxic to the cells. Subsequently, outputs of inflammasome activation were examined by real-time PCR, protein level ELISA and immunoblot methods at mRNA level. In addition, mitochondrial reactive oxygen species (ROS) and mitochondrial membrane potential were determined by fluorometric methods. According to our results, EP significantly reduced the amount of IL-1β and IL-18 inflammatory cytokines in mRNA level with LPS+ATP model which activates NLPR3 inflammasome in N9 microglial cells. In addition, it suppressed the amount of mitochondrial ROS and prevented the mitochondrial membrane potential from deteriorating. This study is the first to demonstrate that EP is effective to suppress the NLRP3 inflammasome activation in microglial cells. Keywords: Ethyl Pyruvate, Microglia, NLRP3 Inflammasome, Pyroptosis, LPS, ATP