Toll-like receptor 4-interacting SPA4 peptide suppresses the NLRP3 inflammasome in response to LPS and ATP stimuli

Abstract

Inflammation is induced because of interplay among multiple signaling pathways and molecules during infectious and noninfectious tissue injuries. Crosstalk between Toll-like receptor-4 signaling and the neuronal apoptosis inhibitor protein, major histocompatibility class 2 transcription activator, incompatibility locus protein from Podospora anserina, and telomerase-associated protein (NACHT), leucine-rich repeat (LRR), and pyrin domain-containing protein 3 (NLRP3) inflammasome against pathogen- or damage-associated molecular patterns can cause exaggerated inflammation. We previously established that the Toll-like receptor-4-interacting SPA4 peptide suppresses gram-negative bacterial lipopolysaccharide (Toll-like receptor-4 ligand)-induced nuclear factor-κB and inflammatory response. In the present study, we hypothesized that the SPA4 peptide exerts its anti-inflammatory effects by suppressing the crosstalk between Toll-like receptor-4 signaling and the NLRP3 inflammasome. We evaluated binding of the lipopolysaccharide-ligand to cell-surface Toll-like receptor-4 in the presence or absence of adenosine triphosphate (an NLRP3 inflammasome inducer) by flow cytometry. The expression and activity of NLRP3 inflammasome-related parameters were studied in cells challenged with lipopolysaccharide and adenosine triphosphate using molecular and immunologic methods. The cells were challenged with lipopolysaccharide and treated with SPA4 peptide before (pre-adenosine triphosphate) or after (post-adenosine triphosphate) secondary challenge with adenosine triphosphate. Our data demonstrate that the Toll-like receptor-4-interacting SPA4 peptide does not affect the binding of lipopolysaccharide to Toll-like receptor-4 in the presence or absence of adenosine triphosphate. We also found that the SPA4 peptide inhibits mRNA and cellular protein levels of pro-interleukin-1β and NLRP3, formation of the NLRP3 inflammasome, caspase activity, and release of interleukin-1β. Furthermore, the SPA4 peptide treatment reduced the secreted levels of interleukin-1β from cells overexpressing Toll-like receptor-4 compared with cells expressing the dominant-negative form of Toll-like receptor-4. Together our results suggest that the SPA4 peptide exerts its anti-inflammatory activity by suppressing Toll-like receptor-4-priming of the NLRP3 inflammasome.

Lung and general health effects of Toll-like receptor-4 (TLR4)-interacting SPA4 peptide

“…Initial results about the efficacy of SPA4 peptide are promising for its therapeutic effects in mouse models of lung inflammation and infection. The SPA4 peptide on its own does not induce an inflammatory response [2,4,6,12]. Thus, the present work focused on assessing the health effects, toxicity profile, and biological activity of SPA4 peptide in mice.…”

“…The CD-1 mice included in this investigation have been used extensively for studies related to safety and toxicity assessment of new drugs and chemical entities [8][9][10][11]. Previously published results have demonstrated that the SPA4 peptide treatment on its own does not induce any cytokine response in dendritic cells and macrophages [2,4,12]. However, the toxicity of SPA4 peptide remained unknown.…”

Emerging Mechanisms of Innate Immunity and Their Translational Potential in Inflammatory Bowel Disease

“…To this end, inhibitors of the NLRP3 inflammasome such as Astragalus polysaccharide ( 183 , 184 ), INF39 (an acrylate derivative) ( 185 ), and levornidazole ( 186 ) have been shown in experimental models to improve colitis. The peptide SPA4 has anti-inflammatory actions through indirect suppression of the NLRP3 inflammasome by interacting with TLR4 ( 187 ) although this has not yet been used in colitis models. However, the plant flavonoid alpinetin does improve DSS colitis through a similar mechanism of modulation of the NLRP3 and TLR4 pathways ( 188 ).…”

“…In addition to the studies on SPA4 ( 187 ), alpinetin ( 188 ), and BML-111 ( 200 ) detailed above, other lines of enquiry suggest that modulation of TLR4 signaling may yield benefit in the management of intestinal inflammation. Unlike studies on TLR3 where enhanced signaling is effective the evidence with TLR4 is predominantly in blocking signaling.…”

Prognostic predictive value of TLR4 polymorphisms in Han Chinese population with hypertrophic cardiomyopathy

“…Recently, increasing evidence has demonstrated that the immune system and multiple pro‐inflammatory factors play a fundamental role in the pathogenesis of HCM [6]. Toll‐like receptor‐4 ( TLR4 ) is a family member of pattern recognition receptors that play important roles in mediating the inflammatory response [7–9]. TLR4 is mapped to 9q32‐q33 region and contains three exons, which participates in immuno‐regulation by suppressing T lymphocyte proliferation and regulating macrophage function [10–12].…”