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.

Chemical, structural and cytotoxicity characterisation of experimental fluoride-doped calcium phosphates as promising remineralising materials for dental applications.

OBJECTIVES: This study aimed at evaluating the cytotoxicity, chemical and structural properties of experimental fluoride-doped calcium-phosphates as potential remineralising materials for dental applications. METHODS: Experimental calcium phosphates were formulated using ß-tricalcium phosphate, monocalcium phosphate monohydrate, calcium hydroxide, and different concentrations of calcium/sodium fluoride salts [(5 wt%: VSG5F), (10 wt%: VSG10F), (20 wt%: VSG20F)]. A fluoride-free calcium phosphate (VSG) was used as control. Each tested material was immersed in simulated body fluid (SBF), (24 h, 15 and 30 days) to assess their ability to crystallise into apatite-like. Cumulative fluoride release was assayed up to 45 days. Moreover, each powder was placed into a medium containing human dental pulp stem cells (200 mg/mL) and their cytotoxicity was analysed using the 3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyltetrazolium bromide (MTT) assay (24 h, 48 h and 72 h incubation). These latter results were statistically analysed by ANOVA and Tukey's test (α = 0.05). RESULTS: All the experimental VSG-F materials produced fluoride-containing apatite-like crystals after SBF immersion. VSG20F presented prolonged release of fluoride ions into the storage media (45d). VSG, VSG10F and VSG20F showed a significant cytotoxicity at dilution of 1:1, while at 1:5, only VSG and VSG20F demonstrated a reduction in cell viability. At lower dilutions (1:10, 1:50 and 1:100) all specimens showed no significant toxicity to hDPSCs, but an increase in cell proliferation. SIGNIFICANCE: The experimental fluoride-doped calcium-phosphates are biocompatible and possess a clear ability to evoke fluoride-containing apatite-like crystallisation. Hence, they may be promising remineralising materials for dental applications.

Infective Endocarditis: A Focus on Oral Microbiota.

Infective endocarditis (IE) is an inflammatory disease usually caused by bacteria entering the bloodstream and settling in the heart lining valves or blood vessels. Despite modern antimicrobial and surgical treatments, IE continues to cause substantial morbidity and mortality. Thus, primary prevention and enhanced diagnosis remain the most important strategies to fight this disease. In this regard, it is worth noting that for over 50 years, oral microbiota has been considered one of the significant risk factors for IE. Indeed, among the disparate recommendations from the American heart association and the European Society of Cardiology, there are good oral hygiene and prophylaxis for high-risk patients undergoing dental procedures. Thus, significant interest has grown in the role of oral microbiota and it continues to be a subject of research interest, especially if we consider that antimicrobial treatments can generate drug-resistant mutant bacteria, becoming a severe social problem. This review will describe the current knowledge about the relationship between oral microbiota, dental procedures, and IE. Further, it will discuss current methods used to prevent IE cases that originate from oral pathogens and how these should be focused on improving oral hygiene, which remains the significant persuasible way to prevent bacteremia and systemic disorders.