GroEL triggers NLRP3 inflammasome activation through the TLR/NF-κB p-p65 axis in human periodontal ligament stem cells

Abstract

The interaction between bacteria and the host plays a vital role in the initiation and progression of systemic diseases, including gastrointestinal and oral diseases, due to the secretion of various virulence factors from these pathogens. GroEL, a potent virulence factor secreted by multiple oral pathogenic bacteria, is implicated in the damage of gingival epithelium, periodontal ligament, alveolar bone and other peripheral tissues. However, the underlying biomechanism is still largely unknown. In the present study, we verify that GroEL can trigger the activation of NLRP3 inflammasome and its downstream effector molecules, IL-1β and IL-18, in human periodontal ligament stem cells (hPDLSCs) and resultantly induce high activation of gelatinases (MMP-2 and MMP-9) to promote the degradation of extracellular matrix (ECM). GroEL-mediated activation of the NLRP3 inflammasome requires the participation of Toll-like receptors (TLR2 and TLR4). High upregulation of TLR2 and TLR4 induces the enhancement of NF-κB (p-p65) signaling and promotes its nuclear accumulation, thus activating the NLRP3 inflammasome. These results are verified in a rat model with direct injection of GroEL. Collectively, this study provides insight into the role of virulence factors in bacteria-induced host immune response and may also provide a new clue for the prevention of periodontitis.

1. Introduction

Periodontitis, a microbiome-driven chronic inflammatory disease, occurs in periodontal tissue and can lead to persistent destruction of periodontal tissue and eventually tooth loss [1,2]. Severe periodontitis is the sixth most prevalent disease worldwide and affects approximately 10% of adults globally [3]. In addition, periodontitis is closely related to the occurrence and development of systemic diseases, such as diabetes, cardiovascular disease, Alzheimer's disease and rheumatoid joints [1,4,5]. Pathogenic bacteria in subgingival plaques are a precondition for periodontitis [6,7]. Pathogenic bacteria and their microbial virulence factors, such as lipopolysaccharide (LPS), proteases, GroEL and lipoteichoic acid, stimulate host cells to produce inflammatory factors and enzymes, which in turn contribute to the destruction of periodontal tissues [8].

Bacterial GroEL, a member of the heat shock protein (HSP) family, is highly homologous to human HSP60 and plays an important role in the proper folding and translocation of proteins during protein synthesis, as well as in the recombination of denatured proteins [9,10]. Nonetheless, GroEL has also been reported to play a pathogenic role in a variety of bacterial infectious diseases and autoimmune diseases [11‒13]. Our previous study revealed that GroEL levels in gingival crevicular fluid and saliva were higher in patients with apical periodontitis than in healthy individuals [14]. A previous study based on clinical investigation reported that salivary immunoglobulin A (IgA) has immunoreactivities with Porphyromonas gingivalis (P. gingivalis) GroEL and Campylobacter rectus (C. rectus) GroEL, and its level is correlated with the severity of periodontal disease [15]. Another study reported that elevated levels of anti-HSP60 antibodies were detected in the serum of cardiovascular patients with severe periodontitis [16]. Meanwhile, periodontal therapy could significantly reduce the GroEL expression level in serum derived from P. gingivalis [17]. Local injection of bacterial GroEL induces absorption of the skull in mice and alveolar bone in rats [11,18]. In addition, GroEL from Tannerella forsythia (T. forsythia) and C. rectus promotes the production of interleukin 6 and 8 in human gingival fibroblasts and periodontal ligament fibroblasts (hPDLFs) [11,19]. Lin et al. [20] reported that P. gingivalis GroEL induced the production of IL-6 and IL-8 in osteoblasts through p38/MAPK or JNK/MAPK signaling. Fusobacterium nucleatum (F. nucleatum) GroEL significantly upregulated the expressions of adhesion molecules such as intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) and E-selectin, as well as chemokines such as IL-8 and monocyte chemoattractant protein-1 (MCP-1), in human microvascular endothelial cells and enhanced the adhesion and migration of monocytes [21]. Previous evidence strongly suggested that GroEL may be a potential stimulator of periodontal disease. However, its virulent effect on periodontitis is not fully understood.