Ligustrazine disrupts lipopolysaccharide-activated NLRP3 inflammasome pathway associated with inhibition of Toll-like receptor 4 in hepatocytes

Abstract

Intestine microbial products may translocate into the liver via portal vein and trigger or exacerbate hepatocyte inflammatory responses during liver injury. The NLRP3 inflammasome pathway plays a key role in regulation of inflammatory cytokines in response to bacterial products. The present study was aimed to investigate the effects of ligustrazine, a natural alkaloid compound, on the NLRP3 inflammasome pathway activation and interleukin-1β (IL-1β) generation in hepatocytes. We cultured human LO2 hepatocytes and treated them with lipopolysaccharide (LPS), a membrane component of Gram-negative bacteria, for mimicking hepatic exposure to microbial products in vitro. The results demonstrated that LPS upregulated NLRP3 and cleaved-caspase-1, and promoted the expression and secretion of IL-1β in LO2 cells. Ligustrazine was found to reduce NLRP3 and cleaved-caspase-1, prevented IL-1β cleavage, and decreased IL-1β secretion into extracellular environment. Further examinations showed that LPS upregulated the expression of Toll-like receptor 4 (TLR4), but ligustrazine repressed TLR4 expression in LPS-treated hepatocytes. Moreover, pharmacological inhibition of TLR4 by its specific inhibitor TAK-242 downregulated NLRP3 and cleaved-caspase-1, and combination treatment with TAK-242 and ligustrazine led to more significant inhibitory effects on the NLRP3 pathway. TAK-242 also reduced cleaved-IL-1β, and this reducing effect was enhanced by ligustrazine. Collectively, the current results revealed that ligustrazine interrupted LPS-activated NLRP3 inflammasome signaling and reduced generation of IL-1β in hepatocytes, which was associated with inhibition of TLR4. This study uncovered a novel mechanism for ligustrazine as a potential hepatoprotective agent.

Keywords: Hepatocyte; Ligustrazine; Lipopolysaccharide; Liver injury; NLRP3 inflammasome; Toll-like receptor 4.