Neuroinflammatory and Oncogenic Effects of Extracellular Vesicles Derived from a Periodontal Pathogen : 치주 병원체 유래 세포밖소포체에 의한 신경염증 및 종양유발 기전

다국어 초록 (Multilingual Abstract)

Gram‑negative bacterial extracellular vesicles (bEVs), also known as outer‑membrane vesicles, have emerged as key mediators of intercellular signaling that can cross tissue barriers. This study investigates how bEVs released by the periodontopathogen Aggregatibacter actinomycetemcomitans (Aa) influence both the oral–brain axis and head and neck squamous cell carcinoma (HNSCC). In a ligature‑induced mouse model of periodontitis, Aa‑derived bEVs administered by intragingival injection or bEV‑soaked hydrogel reached the brain and trigeminal ganglion (TG), where interleukin‑6 (IL‑6) and tumor necrosis factor‑α (TNF‑α) levels were significantly elevated. Human Embryonic Kidney (HEK) ‑Blue reporter assays and analyses in Myeloid differentiation primary response 88 (MyD88) ‑deficient mice showed that lipopolysaccharide (LPS) on the bEV surface activates Toll-like receptor 4 (TLR4), whereas RNA cargo within the bEV stimulates endosomal Toll-like receptor (TLR8), both converging on the MyD88–Nuclear factor kappa-light-chain-enhancer of activated B cells (NF‑κB) axis. Following trans‑epidermal or gingival administration, Aa‑derived bEVs were transported retrogradely along axons to TG neuronal somata, and electrophysiological recordings demonstrated reduced action‑potential thresholds and increased firing frequencies in small‑diameter nociceptive neurons, indicating enhanced neuroinflammatory sensitization. In parallel, two human HNSCC cell lines (UMSCC1 and FaDu) exposed to Aa‑derived bEVs exhibited accelerated cell‑cycle progression through the S and G2 phases, increased proliferative activity in Ki‑67 and Carboxyfluorescein succinimidyl ester (CFSE) assays, and enhanced invasiveness in Matrigel‑based invasion assays. Transcriptome profiling revealed broad reprogramming of gene expression, while microRNA (miRNA) sequencing identified selective upregulation of miR‑146a, a regulatory molecule linking immune modulation and tumorigenesis. Functionally, miR‑146a elevation correlated with increased invasion, highlighting a convergence between inflammatory and oncogenic pathways. Collectively, Aa‑derived bEVs exert distinct biological effects through two mechanistically linked routes: in neural tissues, they amplify IL‑6 and TNF‑α production via TLR4/8–MyD88–NF‑κB signaling and exRNA‑dependent activation, whereas in malignant epithelial cells, they remodel transcriptional and miRNA networks to promote proliferation and invasion. These findings position bEVs as pivotal molecular messengers that clarify how oral infection contributes to neuroinflammation and cancer progression and suggest clinical opportunities for targeting bEV biogenesis, circulation, cellular uptake, or TLR8‑dependent RNA signaling, as well as for developing bEV‑based liquid biopsy strategies.

목차 (Table of Contents)

• ABSTRACT 1
• INTRODUCTION 3
• MATERIAL AND METHODS 7
• RESULTS 19
• ⚫ Part 1. Aggregatibacter actinomycetemcomitans Extracellular Vesicles Promote Neuroinflammation in the Brain and Trigeminal Ganglion 19
•  Basic characterization of Aa EVs 19
•  Effect of Aa EVs on TNF‑α and IL‑6 Expression in the Brain of Ligature‑Induced Periodontitis Mice 21
•  Effect of Aa EVs on TLR4 and TLR8 Signaling Pathways 31
•  Effect of Aa EVs on Retrograde Axonal Transport to the Trigeminal Ganglion 35
•  Effect of Aa EVs on the excitability of nociceptive sensory neurons 43
• ⚫ Part 2. Aggregatibacter actinomycetemcomitans Extracellular Vesicles Induce Transcriptomic and Oncogenic Changes in Head and Neck Squamous Cell Carcinoma Cells 46
•  Analysis of purified Aa EVs 46
•  Effect of Aa EVs on Cell Viability in HNSCC Cell Lines 47
•  Effect of Aa EVs on Proliferation and Invasion of HNSCC Cell Lines 48
•  Effect of Aa EVs on RNA‑Seq Transcriptome Profiles of HNSCC Cell Lines 51
•  Effect of Aa EVs on Transcriptome Profiles Validated by qRT-PCR in HNSCC Cell Lines 55
•  Effect of Ectopic miR‑146a Expression on Invasion of HNSCC Cell Lines 56
• DISCUSSION 58
• REFERENCES 70
• 초록 84