The rising prevalence of acetyl tributyl citrate (ATBC) as an environmental pollutant has raised considerable concern about its potential role in oral diseases. This study focuses on the effects of ATBC exposure on oral squamous cell carcinoma (OSCC), with the specific aim of identifying potential targets and elucidating the associated molecular mechanisms, employing network toxicology, molecular docking, and molecular dynamics (MD) simulation. Relevant targets of OSCC were collected from the TTD, GeneCards, and OMIM databases. The ChEMBL, STITCH, TargetNet, and Swiss Target Prediction databases were utilized to screen ATBC compounds and identify associated compound targets. We selected 107 potential targets for ATBC-induced OSCC and extracted 22 core targets using STRING 12.0 and Cytoscape 3.9.1, including AKT1, HSP90AA1, ESR1, CASP3, BCL2, PPARG, MMP9, and EGFR. Gene ontology (GO) analysis revealed that ATBC-induced OSCC was associated with cell proliferation and apoptosis caused by exogenous chemicals. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that ATBC participates in the cancer signaling pathway through Heat Shock Protein 90 Alpha Family Class A Member 1 (HSP90AA1), Epidermal Growth Factor Receptor (EGFR), and Matrix Metalloproteinase-9 (MMP9). Molecular docking and MD simulations indicate the high stability and reliability of ATBC binding to these core targets. This study elucidates the role of ATBC in the induction of OSCC and its underlying molecular mechanisms, offering significant support for uncovering the toxicological mechanisms of ATBC. Moreover, it provides a theoretical foundation for developing preventive strategies and therapeutic interventions for oral diseases associated with ATBC exposure.

Background: This study aimed to investigate the regulation of CpG oligodeoxynucleotides (ODNs) on proliferation and osteogenic differentiation of MC3T3 cells. Methods: The laser co-focusing and flow cytometry assay were employed to detect cell uptake of CpG ODN 2006. Twelve ODNs were sythesized, and their effects on proliferation and differentiation were detected by MTT and alkaline phosphatase (ALP) activity assay. Flow cytometry assay was used to examine the regulation of CpG ODN on cell cycle. Quantitative real-time PCR (qRT-PCR) and western blot were used to evaluate the regulation of CpG ODN on mRNA and protein expression of osteogenic differentiation genes. Results: The phosphorothioate CpG ODN 2006 could efficiently enter the MC3T3 cells in 1 h and locate in the cytoplasm. The MTT assay demonstrated CpG ODNs could promote MC3T3 cell proliferation and differentiation in the early stage, and gradually attenuated along with the increase of treating time, except for BW001 and FC001. qRT-PCR assay demonstrated that all the 12 CpG ODNs could promote the relative expression level of osteogenic differentiated genes, SP7 and OCN. In addition, western blot analysis suggested the CpG ODNs of BW001 and FC001 could increase the protein expression of P27^Kip1 and Runx2 and decrease the protein expression of cyclin D1. Conclusion: The selected CpGODNs may be a potential gene therapy for bone regeneration of periodontitis.

An oligodeoxynucleotide (ODN), with its design based on human mitochondrial DNA (MT01), has been demonstrated to promote the proliferation and activation of osteoblasts. However, critical issues such as poor oligonucleotide stability in the presence of nucleases currently limit its broad application. This study evaluated whether N-isopropylacrylamide-modified polyethylenimine (PEN) nanoparticles could effectively deliver ODN MT01 locally with low or no toxicity and promote osteoblast differentiation in vitro. Our data demonstrate that at a w/w ratio of 6 : 1, PEN/MT01 nanoparticles were effectively transfected into MG63 cells without cytotoxicity, whereby they promoted cell proliferation in vitro. Quantitative PCR and western blotting analyses showed that the PEN/MT01 complexes enhanced mRNA and protein expression of osteoprotegerin, Sp7, and runt-related transcription factor 2, and decreased levels of receptor activator of nuclear factor kappa-B ligand, which are markers indicative of MG63 differentiation into preosteoblasts or mature osteoblasts. This approach demonstrates the potential of using PEN as a biocompatible gene therapy carrier to address regeneration of bone defects.

Clinical prevention and treatment of periodontitis-induced bone absorption remains a challenge. The anti-infection role of CpG oligodeoxynucleotides (CpG ODNs) is well known; however, their effect on osteoclasts is still unclear. Here, we show that some CpG ODNs can regulate osteoclastogenesis of RAW264.7 cells. The phosphorothioate CpG ODN was efficiently taken up by the cells within 1 h and distributed in the cytoplasm. BW006, YW001, YW002, and FC004 CpG ODNs significantly repressed cell proliferation by targeting several cyclin proteins to arrest the cells in the G2 phase and to further initiate cell apoptosis. Regarding differentiation, we selected six CpG ODNs (FC002, BW006, YW002, YW001, FC004, and MT01) that markedly inhibited the gene expression levels of Nfatc, c-fos, RANK, and MMP9. TRAP staining showed that only YW002, YW001, and FC004 suppressed osteoclast generation and maturation. These three CpG ODNs dramatically declined the protein levels of osteoclastogenic proteins by elevating the ratio of OPG/RANKL and also downregulating the inflammatory factors (TNF-α, IL-1β, IL-6, and IL-17) at different stages. Thus, the selected CpG ODNs may be a potential molecular therapy for the prevention and treatment of periodontitis-mediated bone resorption.

Purpose: In a previous study, we found that oligodeoxynucleotide (ODN) YW002 could induce the activity of alkaline phosphatase of early osteogenesis in human periodontal membrane stem cells, and downregulate the synthesis of nitric oxide in RAW 264.7 cells in the late inflammatory stage, laying the experimental foundation for the subsequent application of ODN YW002 in periodontitis. However, free ODN does not easily adhere to cells and is easily hydrolyzed by nuclease, so the immune effect of ODN is greatly reduced. Therefore, the nano-drug delivery system provides a method for efficient delivery and uptake of ODN. Methods: We synthesized a polyethyleneimine (PEI) modified chondroitin sulfate (CS) derivative (PEI-CS) via Michael addition to deliver ODN YW002. We aimed to evaluate whether PEI-CS could effectively deliver YW002 to RAW 264.7 cells and if it can regulate inflammation in vitro. PEI-CS/YW002 nanocomplexes were locally injected into a mouse periodontitis model, and the therapeutic effects were evaluated by microcomputed tomography (micro-CT) and hematoxylin–eosin (H&E) staining. Results: The results indicated that PEI-CS had good biocompatibility and could form a stable nanocomplex with YW002 at a mass ratio of 4 : 1. Moreover, PEI-CS could deliver YW002 into RAW 246.7 cells and markedly decreased the expression levels of interleukin (IL)-1β, IL-6 and tumor necrosis factor (TNF)-α. Histological evaluation and micro-CT scanning showed that PEI-CS/YW002 nanocomplexes effectively inhibited periodontitis and reduced alveolar bone resorption in mice. Conclusion: Our study has underscored the potential of PEI-CS/YW002 nanocomplexes as promising agents for the prevention and treatment of periodontitis due to their potent anti-inflammatory effects.

Bisphenol A (BPA) is widely used in producing polycarbonate plastics and epoxy resins, but it may pose potential health risks to the endocrine, reproductive, and immune systems and may also promote cancer. Researchers found that BPA leaching from dental materials into saliva could pose a health risk. Evidence linking BPA to oral squamous cell carcinoma (OSCC) is insufficient, and its carcinogenic potential and role in OSCC pathogenesis warrant further exploration. We utilized network toxicology to investigate pathogenic targets and pathways in BPA with OSCC. Molecular docking and molecular dynamics simulations explored BPAs binding efficiency and stability with core targets. Single-cell sequencing analysis was conducted to determine the expression of core targets in different cell types. Network toxicology revealed that BPA promotes OSCC by affecting key proteins such as prostaglandin-endoperoxide synthase 2 (PTGS2) in the neuroactive ligand-receptor interaction pathway. Molecular docking demonstrated that BPA has the strongest binding affinity to PTGS2 and Kelch-like ECH-associated protein 1 (KEAP1) (−7.6 kcal/mol), followed by binding to heat shock protein 90 alpha family class A member 1 (HSP90AA1) (−6.7 kcal/mol). Molecular dynamics simulations further confirmed the stability, and single-cell sequencing demonstrated the distribution of three core targets. This study explored the mechanisms of BPA exposure on OSCC, identifying PTGS2, HSP90AA1, and KEAP1 as key targets. It provides a method for assessing chemical safety and offers new insights into BPAs carcinogenic effects in dentistry, which needs re-evaluation and validation in further research. Not applicable.