Department of Dental Pathology, Faculty of Medicine, University of East Sarajevo, 73300 Foča, Bosnia and Herzegovina.
Clinic of Operative Dentistry, Periodontology and Preventive Dentistry, Saarland University, 664241 Homburg, Germany.
Int J Mol Sci. 2024 Mar 14;25(6):3314. doi: 10.3390/ijms25063314.
The encounter between dental biofilm and neutrophils in periodontitis remains elusive, although it apparently plays a crucial role in the periodontal pathology and constitutes a key topic of periodontology. Dental biofilm and neutrophils were isolated from orally healthy persons and patients with periodontitis. We investigated biofilm and its particle-shedding phenomenon with electron microscopy and nanoparticle tracking analysis (NTA); biofilm shedding-neutrophil interactions were examined ex vivo with epi-fluorescence microscopy. For this purpose, we used acellular dental biofilm shedding, purified lipopolysaccharide (LPS), and phorbol 12-myristate 13-acetate (PMA) as activators, and the interleukin 8 receptor beta (CXCR2) inhibitor and the anti-interleukin 8 receptor alpha (CXCR1) antibody as modulators. The shedding of acellular dental biofilms overwhelmingly consists of bacterial extracellular vesicles (BEVs). The latter induced the moderate formation of neutrophil extracellular traps (NETs) in orally healthy subjects and a strong formation in patients with periodontitis. A CXCR2 inhibitor and an anti-CXCR1 antibody had a minor effect on NET formation. Neutrophils from patients with periodontitis exhibited NET hyper-responsiveness. BEVs were stronger inducers of NET formation than purified LPS and PMA. A plateau of neutrophil responsiveness is reached above the age of 40 years, indicating the abrupt switch of maladaptive trained immunity (TI) into the activated modus. Our results suggest that dental biofilms consist of and disseminate immense amounts of outer membrane vesicles (OMVs), which initiate NET formation via a non-canonical cytosolic LPS/caspase-4/11/Gasdermin D pathway. This modus of NET formation is independent of neutrophil elastase (NE), myeloperoxidase (MPO), peptidylarginine deiminase 4 (PAD4), and toll-like receptors (TLR). In periodontitis, the hyper-responsiveness of neutrophils to BEVs and the increased NET formation appear to be a consequence of TI.
牙周炎中，牙菌斑与中性粒细胞的相互作用仍然难以捉摸，尽管其显然在牙周病理学中起着至关重要的作用，也是牙周病学的一个关键主题。我们从口腔健康者和牙周炎患者中分离出牙菌斑和中性粒细胞。我们使用电子显微镜和纳米颗粒跟踪分析（NTA）研究了生物膜及其颗粒脱落现象；通过荧光显微镜检查了生物膜脱落-中性粒细胞相互作用。为此，我们使用无细胞牙菌斑脱落、纯化脂多糖（LPS）和佛波醇 12-肉豆蔻酸 13-乙酸酯（PMA）作为激活剂，以及白细胞介素 8 受体β（CXCR2）抑制剂和白细胞介素 8 受体α（CXCR1）抗体作为调节剂。无细胞牙菌斑生物膜的脱落主要由细菌细胞外囊泡（BEV）组成。后者在口腔健康者中诱导适度的中性粒细胞细胞外陷阱（NETs）形成，而在牙周炎患者中则诱导强烈的 NETs 形成。CXCR2 抑制剂和抗 CXCR1 抗体对 NET 形成的影响较小。牙周炎患者的中性粒细胞表现出 NET 超反应性。BEVs 比纯化 LPS 和 PMA 更强地诱导 NET 形成。中性粒细胞反应性在 40 岁以上达到平台期，表明适应性训练免疫（TI）突然转变为激活模式。我们的结果表明，牙菌斑由大量外膜囊泡（OMVs）组成，并通过非经典细胞质 LPS/caspase-4/11/Gasdermin D 途径引发 NET 形成。这种 NET 形成模式独立于中性粒细胞弹性蛋白酶（NE）、髓过氧化物酶（MPO）、肽基精氨酸脱亚氨酶 4（PAD4）和 Toll 样受体（TLR）。在牙周炎中，中性粒细胞对 BEVs 的高反应性和增加的 NET 形成似乎是 TI 的结果。
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