Porphyromonas gingivalis Induces Chronic Kidney Disease through Crosstalk between the NF-κB/NLRP3 Pathway and Ferroptosis in GMCs.

OBJECTIVE: Porphyromonas gingivalis (P.gingivalis) is a gram-negative bacterium found in the human oral cavity and is a recognized pathogenic bacterium associated with chronic periodontitis and systemic diseases, including chronic kidney disease (CKD), but the roles and molecular mechanism of P.gingivalis in CKD pathogenesis are unclear. METHODS: In this study, an animal model of oral P.gingivalis administration and glomerular mesangial cells (GMCs) cocultured with M1-polarized macrophages and P.gingivalis supernatant were constructed. After seven weeks of P.gingivalis gavaged, peripheral blood was collected to detect the changes in renal function. By collecting the teeth and kidneys of mice, H&E staining and IHC were used to analyze the expression of periodontal inflammatory factors in mice, PAS staining was used to analyze glomerular lesions. The supernatant of macrophages was treated with 5% P.gingivalis supernatant. H&E staining, IHC, Western blot and RT-PCR were applied to analyze renal inflammatory factors, macrophage M1 polarization, NF-κB, NLRP3 and ferroptosis changes in vitro. RESULTS: We found that oral P.gingivalis administration induced CKD in mice. P.gingivalis supernatant induced macrophage polarization and inflammatory factor upregulation, which triggered the activation of the NF-κB/NLRP3 pathway and ferroptosis in GMCs. By inhibiting the NF-κB/NLRP3 pathway and ferroptosis in GMCs, cell viability and the inflammatory response were partially alleviated in vitro. CONCLUSION: We demonstrated that P.gingivalis induced CKD in mice by triggering crosstalk between the NF κB/NLRP3 pathway and ferroptosis in GMCs. Overall, our study suggested that periodontitis can promote the pathogenesis of CKD in mice, which provides evidence of the importance of periodontitis therapy in the prevention and treatment of CKD. P.gingivalis promotes ferroptosis in kidneys and accelerates the progression of CKD through NF-κB/NLRP3 signaling pathway.

The oral pathogen Porphyromonas gingivalis gains tolerance to the antimicrobial peptide DGL13K by synonymous mutations in hagA.

Porphyromonas gingivalis is a keystone pathogen for periodontal disease. The bacteria are black-pigmented and require heme for growth. P. gingivalis exhibit resistance to many antimicrobial peptides, which contributes to their success in the oral cavity. P. gingivalis W50 was resistant to the antimicrobial peptide LGL13K but susceptible to the all-D-amino acid stereoisomer, DGL13K. Upon prolonged exposure to DGL13K, a novel non-pigmented mutant was isolated. Exposure to the L-isomer, LGL13K, did not produce a non-pigmented mutant. The goal of this study was to characterize the genomic and cellular changes that led to the non-pigmented phenotype upon treatment with DGL13K. The non-pigmented mutant showed a low minimum inhibitory concentration and two-fold extended minimum duration for killing by DGL13K, consistent with tolerance to this peptide. The DGL13K-tolerant bacteria exhibited synonymous mutations in the hagA gene. The mutations did not prevent mRNA expression but were predicted to alter mRNA structure. The non-pigmented bacteria were deficient in hemagglutination and hemoglobin binding, suggesting that the HagA protein was not expressed. This was supported by whole cell enzyme-linked immunosorbent assay and gingipain activity assays, which suggested the absence of HagA but not of two closely related gingipains. In vivo virulence was similar for wild type and non-pigmented bacteria in the Galleria mellonella model. The results suggest that, unlike LGL13K, DGL13K can defeat multiple bacterial resistance mechanisms but bacteria can gain tolerance to DGL13K through mutations in the hagA gene.

Intraperitoneal Injection of the Porphyromonas gingivalis Outer Membrane Vesicle (OMV) Stimulated Expressions of Neuroinflammatory Markers and Histopathological Changes in the Brains of Adult Zebrafish.

Porphyromonas gingivalis is the major pathogenic bacteria found in the subgingival plaque of patients with periodontitis, which leads to neuroinflammation. The bacteria destroy periodontal tissue through virulence factors, which are retained in the bacteria's outer membrane vesicles (OMV). This study aimed to determine the real-time effect of an intraperitoneal injection of P. gingivalis OMV on the production and expression of inflammatory markers and histopathological changes in adult zebrafishes' central nervous systems (CNS). Following the LD50 (21 µg of OMV), the zebrafish were injected intraperitoneally with 18 µg of OMVs, and the control group were injected with normal saline at seven different time points. Brains of experimental zebrafish were dissected at desired time points for colorimetric assays, ELISA, and histology. This study discovered that nitric oxide and PGE2 were significantly increased at 45 min, while IL-1ß and IL-6 were expressed at subsequent 12 h and 24 h time points, respectively. Histopathological changes such as blood coagulation, astrocytosis, edema, spongiosis, and necrosis were observed between the 6hour and 24 h time points. The two apoptotic enzymes, caspases 3 and 9, were not expressed at any point. In summary, the OMV-induced neuroinflammatory responses and histopathological changes in adult zebrafish were time-point dependent. This study will enrich our understanding of the mechanism of P. gingivalis OMVs in neuroinflammation in a zebrafish model, most especially the timing of the expression of inflammatory mediators in relation to observable changes in brain tissues.

Oral Pathobiont-Derived Outer Membrane Vesicles in the Oral-Gut Axis.

Oral pathobionts are essential in instigating local inflammation within the oral cavity and contribute to the pathogenesis of diseases in the gastrointestinal tract and other distant organs. Among the Gram-negative pathobionts, Porphyromonas gingivalis and Fusobacterium nucleatum emerge as critical drivers of periodontitis, exerting their influence not only locally but also as inducers of gut dysbiosis, intestinal disturbances, and systemic ailments. This dual impact is facilitated by their ectopic colonization of the intestinal mucosa and the subsequent mediation of distal systemic effects by releasing outer membrane vesicles (OMVs) into circulation. This review elucidates the principal components of oral pathobiont-derived OMVs implicated in disease pathogenesis within the oral-gut axis, detailing virulence factors that OMVs carry and their interactions with host epithelial and immune cells, both in vitro and in vivo. Additionally, we shed light on the less acknowledged interplay between oral pathobionts and the gut commensal Akkermansia muciniphila, which can directly impede oral pathobionts' growth and modulate bacterial gene expression. Notably, OMVs derived from A. muciniphila emerge as promoters of anti-inflammatory effects within the gastrointestinal and distant tissues. Consequently, we explore the potential of A. muciniphila-derived OMVs to interact with oral pathobionts and prevent disease in the oral-gut axis.

Akkermansia muciniphila as a Potential Guardian against Oral Health Diseases: A Narrative Review.

The oral microbiome is a diverse ecosystem containing a community of symbiotic, commensal, and pathogenic microorganisms. One key microorganism linked to periodontal disease (PD) is Porphyromonas gingivalis (P. gingivalis), a Gram-negative anaerobic bacterium known to have several virulence factors that trigger inflammation and immune evasion. On the other hand, Akkermansia muciniphila (A. muciniphila), a symbiotic bacterium, has been recently shown to play an important role in mitigating inflammation and reducing periodontal damage. In vivo and in vitro studies have shown that A. muciniphila decreases inflammatory mediators and improves immune responses, suggesting its role in mitigating PD and related inflammatory systemic conditions such as diabetes, hypertension, and obesity. This review discusses the anti-inflammatory effects of A. muciniphila, its impact on periodontal health, and its potential role in managing systemic diseases. The overall aim is to elucidate how this bacterium might help reduce inflammation, improve oral health, and influence broader health outcomes.

SIRT6 alleviates senescence induced by Porphyromonas gingivalis in human gingival fibroblasts.

OBJECTIVE: Bidirectional influences between senescence and inflammation are newly discovered. This study aimed to clarify the roles and mechanism of Porphyromonas gingivalis (P. gingivalis) in exacerbating senescence in human gingival fibroblasts (HGFs). DESIGN: Subgingival plaque and gingivae were collected from twenty-four periodontitis patients and eighteen periodontally healthy subjects. Quantities of P. gingivalis in subgingival plaque were explored using real-time PCR and the expressions of p53, p21 and SIRT6 in gingivae were detected by IHC. Moreover, senescence in HGFs was induced by P. gingivalis lipopolysaccharide (LPS) and the expressions of senescence-related ß-galactosidase (SA-ß-gal), p53, p21 and senescence-associated secretory phenotype (IL-6 and IL-8) with or without treatment by SIRT6 activator UBCS039 were explored by IHC, western blot and ELISA, respectively. In addition, the levels of SIRT6, Nrf2, HO-1 and reactive oxygen species (ROS) were examined by western blot and flow cytometry. RESULTS: Quantities of P. gingivalis in subgingival plaque and semi-quantitative scores of p53 and p21 in gingivae of periodontitis patients were increased compared with healthy controls (p < 0.05), while SIRT6 score in periodontitis patients was decreased (p < 0.001). Quantities of P. gingivalis were positively correlated with p53 and p21 scores (0.6 < r < 0.9, p < 0.01), and negatively correlated with SIRT6 score (-0.9 < r<-0.6, p < 0.01). Moreover, P. gingivalis LPS increased the levels of SA-ß-gal, p53, p21, IL-6, IL-8 and ROS and decreased the levels of SIRT6, Nrf2 and HO-1 in HGFs, which was rescued by UBCS039 (p < 0.05). CONCLUSIONS: P. gingivalis LPS could induce senescence of HGFs, which could be reversed by SIRT6 via Nrf2-HO-1 signaling pathway.

Bacterial Membrane Vesicles: The Missing Link Between Bacterial Infection and Alzheimer Disease.

Periodontitis is a common chronic inflammatory disease, affecting approximately 19% of the global adult population. A relationship between periodontal disease and Alzheimer disease has long been recognized, and recent evidence has been uncovered to link these 2 diseases mechanistically. Periodontitis is caused by dysbiosis in the subgingival plaque microbiome, with a pronounced shift in the oral microbiota from one consisting primarily of Gram-positive aerobic bacteria to one predominated by Gram-negative anaerobes, such as Porphyromonas gingivalis. A common phenomenon shared by all bacteria is the release of membrane vesicles to facilitate biomolecule delivery across long distances. In particular, the vesicles released by P gingivalis and other oral pathogens have been found to transport bacterial components across the blood-brain barrier, initiating the physiologic changes involved in Alzheimer disease. In this review, we summarize recent data that support the relationship between vesicles secreted by periodontal pathogens to Alzheimer disease pathology.

Epimedium polysaccharides mitigates Porphyromonas gingivalis-exacerbated intestinal inflammation by suppressing the Th17 pathway and modulating the gut microbiota.

This study aimed to investigate the potential alleviating effect of Epimedium polysaccharide (EP) on intestinal inflammation aggravated by Porphyromonas gingivalis (Pg). P. gingivalis, an oral pathogen, may play a role in intestinal inflammation, highlighting the necessity to explore substances capable of inhibiting its pathogenicity. Initially, in vitro screening experiments utilizing co-culturing and quantitative polymerase chain reaction revealed that EP significantly inhibited the growth of P. gingivalis and the levels of virulence genes, including Kgp and RgpA. Subsequent mouse experiments demonstrated that EP notably ameliorated Pg-aggravated weight loss, disease activity index, histopathological lesions, and disruption of intestinal barrier integrity, evidenced by a reduction in tight junction protein levels. Flow cytometry analysis further illustrated that EP attenuated Pg-induced Th17 differentiation and Th17-related cytokines, such as IL-17 and IL-6. Additionally, 16S rRNA amplicon sequencing analysis elucidated that EP significantly mitigated Pg-induced gut microbiota dysbiosis, enriching potentially beneficial microbes, including Akkermansia and Bifidobacterium. The metabolomic analysis provided further insight, indicating that EP intervention altered the accumulation of relevant intestinal metabolites and exhibited correlations with disease indicators. In conclusion, our research suggested that EP holds promise as a prospective therapeutic agent for alleviating P. gingivalis-aggravated intestinal inflammation.

The role of periodontitis in cancer development, with a focus on oral cancers.

Periodontitis is a severe gum infection that begins as gingivitis and can lead to gum recession, bone loss, and tooth loss if left untreated. It is primarily caused by bacterial infection, which triggers inflammation and the formation of periodontal pockets. Notably, periodontitis is associated with systemic health issues and has been linked to heart disease, diabetes, respiratory diseases, adverse pregnancy outcomes, and cancers. Accordingly, the presence of chronic inflammation and immune system dysregulation in individuals with periodontitis significantly contributes to the initiation and progression of various cancers, particularly oral cancers. These processes promote genetic mutations, impair DNA repair mechanisms, and create a tumor-supportive environment. Moreover, the bacteria associated with periodontitis produce harmful byproducts and toxins that directly damage the DNA within oral cells, exacerbating cancer development. In addition, chronic inflammation not only stimulates cell proliferation but also inhibits apoptosis, causes DNA damage, and triggers the release of pro-inflammatory cytokines. Collectively, these factors play a crucial role in the progression of cancer in individuals affected by periodontitis. Further, specific viral and bacterial agents, such as hepatitis B and C viruses, human papillomavirus (HPV), Helicobacter pylori (H. pylori), and Porphyromonas gingivalis, contribute to cancer development through distinct mechanisms. Bacterial infections have systemic implications for cancer development, while viral infections provoke immune and inflammatory responses that can lead to genetic mutations. This review will elucidate the link between periodontitis and cancers, particularly oral cancers, exploring their underlying mechanisms to provide insights for future research and treatment advancements.

Porphyromonas gingivalis virulence factors induce toxic effects in SH-SY5Y neuroblastoma cells: GRK5 modulation as a protective strategy.

Periodontitis (PDS) is a chronic inflammatory disease initiated by a dysbiosis of oral pathogenic bacterial species, such as Porphyromonas gingivalis (Pg). These bacteria can penetrate the bloodstream, releasing various endo and exotoxins that fuel the infection, and stimulate toxic inflammation in different compartments, including the brain. However, the specific mechanisms by which PDS/Pg contribute to brain disorders, such as Alzheimer's disease (AD), remain unclear. This study assessed the effects of Pg's virulence factors - lipopolysaccharide (LPS-Pg) and gingipains (gps) K (Kgp) and Rgp - on SH-SY5Y cells. Our results demonstrated that LPS-Pg activated signaling through the Toll-like receptor (TLR)-2/4 induced a significant downregulation of G protein-coupled receptor kinase 5 (GRK5). Additionally, LPS-Pg stimulation resulted in a robust increase in Tau phosphorylation (pTau) and p53 levels, while causing a marked reduction in Bcl2 and increased cell death compared to unstimulated cells (Ns). LPS-Pg also elevated inducible nitric oxide synthase (iNOS) expression, leading to oxidative damage. In cells overexpressing GRK5 via Adenovirus, LPS-Pg failed to increase iNOS and pTau levels compared to GFP control cells. High GRK5 levels also prevented the nuclear accumulation of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kB). Furthermore, the overexpression of a GRK5 mutant form lacking the nuclear localization signal (ΔNLS) nearly abolished LPS-Pg induced p53 and iNOS upregulation. Finally, we tested whether Kgp and Rgp mediated similar effects and our data showed that both gps caused a marked downregulation of GRK5 leading to increased p53 and pTau levels. In conclusion, this study provides further insight into the toxic effects elicited by Pg in cells and suggests that preventing GRK5 deficiency may be a valid strategy to mitigate Pg-induced toxic effects (i.e. cell death, oxidative damage, and Tau hyperphosphorylation) in SH-SY5Y cells, which are typical molecular hallmarks of neurodegenerative disorders.

Unravelling the intricacies of Porphyromonas gingivalis: virulence factors, lifecycle dynamics and phytochemical interventions for periodontal disease management.

Porphyromonas gingivalis is a gram-negative anaerobic bacterium recognized for its pivotal role in the pathogenesis of periodontal diseases. This review covers an overview of the virulence factors and lifecycle stages of P. gingivalis, with a specific focus on attachment and colonization, biofilm formation, growth and multiplication, dormancy survival and dissemination. Additionally, we explore the significance of inter-bacterial cross-feeding within biofilms. Furthermore, we discuss potential phytochemical-based strategies to target P. gingivalis, including the use of curcumin, apigenin, quercetin and resveratrol. Understanding the virulence factors and lifecycle stages of P. gingivalis, along with the promising phytochemical-based interventions, holds promise for advancing strategies in periodontal disease management and oral health promotion.

Dipotassium glycyrrhizate prevents oral dysbiosis caused by Porphyromonas gingivalis in an in vitro saliva-derived polymicrobial biofilm model.

OBJECTIVES: Oral microbiome dysbiosis prevention is important to avoid the onset and progression of periodontal disease. Dipotassium glycyrrhizate (GK2) is a licorice root extract with anti-inflammatory effects, and its associated mechanisms have been well-reported. However, their effects on the oral microbiome have not been investigated. This study aimed to elucidate the effects of GK2 on the oral microbiome using an in vitro polymicrobial biofilm model. METHODS: An in vitro saliva-derived polymicrobial biofilm model was used to evaluate the effects of GK2 on the oral microbiome. One-week anaerobic culture was performed, in which GK2 was added to the medium. Subsequently, microbiome analysis was performed based on the V1-V2 region of the 16 S rRNA gene, and pathogenicity indices were assessed. We investigated the effects of GK2 on various bacterial monocultures by evaluating its inhibitory effects on cell growth, based on culture turbidity. RESULTS: GK2 treatment altered the microbiome structure and decreased the relative abundance of periodontal pathogenic bacteria, including Porphyromonas. Moreover, GK2 treatment reduced the DPP4 activity -a pathogenicity index of periodontal disease. Specifically, GK2 exhibited selective antibacterial activity against periodontal pathogenic bacteria. CONCLUSIONS: These findings suggest that GK2 has a selective antibacterial effect against periodontal pathogenic bacteria; thus, preventing oral microbiome dysbiosis. Therefore, GK2 is expected to contribute to periodontal disease prevention by modulating the oral microbiome toward a state with low inflammatory potential, thereby utilizing its anti-inflammatory properties on the host.

P. gingivalis in oral-prostate axis exacerbates benign prostatic hyperplasia via IL-6/IL-6R pathway.

BACKGROUND: Benign prostatic hyperplasia (BPH) is the most common disease in elderly men. There is increasing evidence that periodontitis increases the risk of BPH, but the specific mechanism remains unclear. This study aimed to explore the role and mechanism of the key periodontal pathogen Porphyromonas gingivalis (P. gingivalis) in the development of BPH. METHODS: The subgingival plaque (Sp) and prostatic fluid (Pf) of patients with BPH concurrent periodontitis were extracted and cultured for 16S rDNA sequencing. Ligature-induced periodontitis, testosterone-induced BPH and the composite models in rats were established. The P. gingivalis and its toxic factor P. gingivalis lipopolysaccharide (P.g-LPS) were injected into the ventral lobe of prostate in rats to simulate its colonization of prostate. P.g-LPS was used to construct the prostate cell infection model for mechanism exploration. RESULTS: P. gingivalis, Streptococcus oralis, Capnocytophaga ochracea and other oral pathogens were simultaneously detected in the Pf and Sp of patients with BPH concurrent periodontitis, and the average relative abundance of P. gingivalis was found to be the highest. P. gingivalis was detected in both Pf and Sp in 62.5% of patients. Simultaneous periodontitis and BPH synergistically aggravated prostate histological changes. P. gingivalis and P.g-LPS infection could induce obvious hyperplasia of the prostate epithelium and stroma (epithelial thickness was 2.97- and 3.08-fold that of control group, respectively), and increase of collagen fibrosis (3.81- and 5.02-fold that of control group, respectively). P. gingivalis infection promoted prostate cell proliferation, inhibited apoptosis, and upregulated the expression of inflammatory cytokines interleukin-6 (IL-6; 4.47-fold), interleukin-6 receptor-α (IL-6Rα; 5.74-fold) and glycoprotein 130 (gp130; 4.47-fold) in prostatic tissue. P.g-LPS could significantly inhibit cell apoptosis, promote mitosis and proliferation of cells. P.g-LPS activates the Akt pathway through IL-6/IL-6Rα/gp130 complex, which destroys the imbalance between proliferation and apoptosis of prostate cells, induces BPH. CONCLUSION: P. gingivalis was abundant in the Pf of patients with BPH concurrent periodontitis. P. gingivalis infection can promote BPH, which may affect the progression of BPH via inflammation and the Akt signaling pathway.

Comprehensive comparative analysis of the periodontal pathogen Porphyromonas gingivalis: exploring the pan-genome, the reconstruction of the gene regulatory network and genome-scale metabolic network.

Porphyromonas gingivalis is a nonmotile, obligate anaerobic, Gram-negative bacterium known for its association with periodontal disease and its involvement in systemic diseases such as atherosclerosis, cardiovascular disease, colon cancer, and Alzheimer's disease. This bacterium produces several virulence factors, including capsules, fimbriae, lipopolysaccharides, proteolytic enzymes, and hemagglutinins. A comparative genomic analysis revealed the open pangenome of P. gingivalis and identified complete type IV secretion systems in strain KCOM2805 and almost complete type VI secretion systems in strains KCOM2798 and ATCC49417, which is a new discovery as previous studies did not find the proteins involved in secretion systems IV and VI. Conservation of some virulence factors between different strains was observed, regardless of their genetic diversity and origin. In addition, we performed for the first time a reconstruction analysis of the gene regulatory network, identifying transcription factors and proteins involved in the regulatory mechanisms of bacterial pathogenesis. In particular, QseB regulates the expression of hemagglutinin and arginine deaminase, while Rex may suppress the release of gingipain through interactions with PorV and the formatum/nitrate transporter. Our study highlights the central role of conserved virulence factors and regulatory pathways, particularly QseB and Rex, in P. gingivalis and provides insights into potential therapeutic targets.

Cinnamaldehyde Protects against P. gingivalis Induced Intestinal Epithelial Barrier Dysfunction in IEC-6 Cells via the PI3K/Akt-Mediated NO/Nrf2 Signaling Pathway.

Porphyromonas gingivalis (Pg), a Gram-negative oral pathogen, promotes and accelerates periodontitis-associated gut disorders. Intestinal epithelial barrier dysfunction is crucial in the pathogenesis of intestinal and systemic diseases. In this study, we sought to elucidate the protective role of cinnamaldehyde (CNM, an activator of Nrf2) against P. gingivalis (W83) and Pg-derived lipopolysaccharide (Pg-LPS) induced intestinal epithelial barrier dysfunction via antioxidative mechanisms in IEC-6 cells. IEC-6 (ATCC, CRL-1592) cells were pretreated with or without CNM (100 µM), in the presence or absence of P. gingivalis (strain W83, 109 MOI) or Pg-LPS (1, 10, and 100 µg/mL), respectively, between 0-72 h time points by adopting a co-culture method. Intestinal barrier function, cytokine secretion, and intestinal oxidative stress protein markers were analyzed. P. gingivalis or Pg-LPS significantly (p < 0.05) increased reactive oxygen species (ROS) and malondialdehyde (MDA) levels expressing oxidative stress damage. Pg-LPS, as well as Pg alone, induces inflammatory cytokines via TLR-4 signaling. Furthermore, infection reduced Nrf2 and NAD(P)H quinone dehydrogenase 1 (NQO1). Interestingly, inducible nitric oxide synthase (iNOS) protein expression significantly (p < 0.05) increased with Pg-LPS or Pg infection, with elevated levels of nitric oxide (NO). CNM treatment suppressed both Pg- and Pg-LPS-induced intestinal oxidative stress damage by reducing ROS, MDA, and NO production. Furthermore, CNM treatment significantly upregulated the expression of tight junction proteins via increasing the phosphorylation levels of PI3K/Akt/Nrf2 suppressing inflammatory cytokines. CNM protected against Pg infection-induced intestinal epithelial barrier dysfunction by activating the PI3K/Akt-mediated Nrf2 signaling pathway in IEC-6 cells.

About a Possible Impact of Endodontic Infections by Fusobacterium nucleatum or Porphyromonas gingivalis on Oral Carcinogenesis: A Literature Overview.

Periodontitis is linked to the onset and progression of oral squamous cell carcinoma (OSCC), an epidemiologically frequent and clinically aggressive malignancy. In this context, Fusobacterium (F.) nucleatum and Porphyromonas (P.) gingivalis, two bacteria that cause periodontitis, are found in OSCC tissues as well as in oral premalignant lesions, where they exert pro-tumorigenic activities. Since the two bacteria are present also in endodontic diseases, playing a role in their pathogenesis, here we analyze the literature searching for information on the impact that endodontic infection by P. gingivalis or F. nucleatum could have on cellular and molecular events involved in oral carcinogenesis. Results from the reviewed papers indicate that infection by P. gingivalis and/or F. nucleatum triggers the production of inflammatory cytokines and growth factors in dental pulp cells or periodontal cells, affecting the survival, proliferation, invasion, and differentiation of OSCC cells. In addition, the two bacteria and the cytokines they induce halt the differentiation and stimulate the proliferation and invasion of stem cells populating the dental pulp or the periodontium. Although most of the literature confutes the possibility that bacteria-induced endodontic inflammatory diseases could impact on oral carcinogenesis, the papers we have analyzed and discussed herein recommend further investigations on this topic.

Mechanism and functional verification of genes by virulence factors of P. gingivalis in ferroptosis.

OBJECTIVE: Porphyromonas gingivalis (P. gingivalis) is a key etiological agent in periodontitis and functions as a facultative intracellular microorganism and involves many virulence factors. These virulence factors participate in multiple intracellular processes, like ferroptosis, the mechanistic underpinnings remain to be elucidated. Aim of this study was to investigate the effects of virulence factors on the host cells. DESIGN: Human umbilical vein endothelial cells (HUVECs) were treated with 4% paraformaldehyde-fixed P. gingivalis, and subsequent alterations in gene expression were profiled via RNA-seq. Further, the molecules associated with ferroptosis were quantitatively analyzed using qRT-PCR and Western blot. RESULTS: A total of 1125 differentially expressed genes (DEGs) were identified, encompassing 225 upregulated and 900 downregulated. Ferroptosis was conspicuously represented in the kyoto encyclopedia of genes and genomes (KEGG) enrichment analysis, with notable upregulation of Heme oxygenase 1 (HMOX1), Ferritin light chain (FTL), and Solute carrier family 3 member 2 (SLC3A2) and downregulation of Scavenger receptor class A member 5 (SCARA5) and glutaminase (GLS). Random selection of DEGs for validation through qRT-PCR corroborated the RNA-Seq data (R2 = 0.93). Kelch like ECH associated protein 1 (Keap1) protein expression decreased after 4 and 8 h, while NFE2 like bZIP transcription factor 2 (Nrf2) and HMOX1 were elevated, with significant nuclear translocation of Nrf2. CONCLUSIONS: The virulence factors of P. gingivalis may potentially instigating ferroptosis through activation of the Keap1-Nrf2-HMOX1 signaling cascade, in conjunction with modulating the expression of other ferroptosis-associated elements. Further research is necessary to achieve a thorough comprehension of these complex molecular interactions.

Oral Microbially-Induced Small Extracellular Vesicles Cross the Blood-Brain Barrier.

Porphyromonas gingivalis (Pg) and its gingipain proteases contribute to Alzheimer's disease (AD) pathogenesis through yet unclear mechanisms. Cellular secretion of small extracellular vesicles or exosomes (EXO) increases with aging as part of the senescence-associated secretory phenotype (SASP). We have shown that EXO isolated from Pg-infected dendritic cells contain gingipains and other Pg antigens and transmit senescence to bystander gingival cells, inducing alveolar bone loss in mice in vivo. Here, EXO were isolated from the gingiva of mice and humans with/without periodontitis (PD) to determine their ability to penetrate the blood-brain barrier (BBB) in vitro and in vivo. PD was induced by Pg oral gavage for 6 weeks in C57B6 mice. EXO isolated from the gingiva or brain of donor Pg-infected (PD EXO) or control animals (Con EXO) were characterized by NTA, Western blot, and TEM. Gingival PD EXO or Con EXO were labeled and injected into the gingiva of uninfected WT mouse model. EXO biodistribution in brains was tracked by an in vivo imaging system (IVIS) and confocal microscopy. The effect of human PD EXO on BBB integrity and permeability was examined using TEER and FITC dextran assays in a human in vitro 3D model of the BBB. Pg antigens (RGP and Mfa-1) were detected in EXO derived from gingival and brain tissues of donor Pg-infected mice. Orally injected PD EXO from donor mice penetrated the brains of recipient uninfected mice and colocalized with hippocampal microglial cells. IL-1ß and IL-6 were expressed in human PD EXO and not in Con EXO. Human PD EXO promoted BBB permeability and penetrated the BBB in vitro. This is the first demonstration that microbial-induced EXO in the oral cavity can disseminate, cross the BBB, and may contribute to AD pathogenesis.