Smoking Impacts Alzheimer's Disease Progression Through Oral Microbiota Modulation.

Alzheimer's disease (AD) is an important public health challenge with a limited understanding of its pathogenesis. Smoking is a significant modifiable risk factor for AD progression, and its specific mechanism is often interpreted from a toxicological perspective. However, microbial infections also contribute to AD, with oral microbiota playing a crucial role in its progression. Notably, smoking alters the ecological structure and pathogenicity of the oral microbiota. Currently, there is no systematic review or summary of the relationship between these three factors; thus, understanding this association can help in the development of new treatments. This review summarizes the connections between smoking, AD, and oral microbiota from existing research. It also explores how smoking affects the occurrence and development of AD through oral microbiota, and examines treatments for oral microbiota that delay the progression of AD. Furthermore, this review emphasizes the potential of the oral microbiota to act as a biomarker for AD. Finally, it considers the feasibility of probiotics and oral antibacterial therapy to expand treatment methods for AD.

Identification of Immune Infiltration and Iron Metabolism-Related Subgroups in Autism Spectrum Disorder.

Autism spectrum disorder (ASD) is a prevalent neurodevelopmental disorder with a broad spectrum of symptoms and prognoses. Effective therapy requires understanding this variability. ASD children's cognitive and immunological development may depend on iron homoeostasis. This study employs a machine learning model that focuses on iron metabolism hub genes to identify ASD subgroups and describe immune infiltration patterns. A total of 97 control and 148 ASD samples were obtained from the GEO database. Differentially expressed genes (DEGs) and an iron metabolism gene collection achieved the intersection of 25 genes. Unsupervised cluster analysis determined molecular subgroups in individuals with ASD based on 25 genes related to iron metabolism. We assessed gene ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment, gene set variation analysis (GSVA), and immune infiltration analysis to compare iron metabolism subtype effects. We employed machine learning to identify subtype-predicting hub genes and utilized both training and validation sets to assess gene subtype prediction accuracy. ASD can be classified into two iron-metabolizing molecular clusters. Metabolic enrichment pathways differed between clusters. Immune infiltration showed that clusters differed immunologically. Cluster 2 had better immunological scores and more immune cells, indicating a stronger immune response. Machine learning screening identified SELENBP1 and CAND1 as important genes in ASD's iron metabolism signaling pathway. These genes express in the brain and have AUC values over 0.8, implying significant predictive power. The present study introduces iron metabolism signaling pathway indicators to predict ASD subtypes. ASD is linked to immune cell infiltration and iron metabolism disorders.

Efficacy and safety of the compound Chinese herb medicine mouthwash on oral ulcer model in rats.

The present study aimed to evaluate the effect of a mouthwash containing a novel compound Chinese herbal medicine (artemisia capillaris, chrysanthemum, honeysuckle, angelica dahurica and asarum sieboldii) on oral ulcers and analyze sub chronic oral toxicity in rats. For efficacy study, mouthwash was administered on the ulcer area twice daily. Compared with the control group, healing time in the test group was shorter and the ulcer area was smaller. Histological analysis showed less inflammatory cell infiltration in the test group. For sub chronic oral toxicity, mouthwash was administered by oral gavage for 93 consecutive days. There were no significant differences in body weight, food consumption or organ coefficients between the test and control groups. Some parameters of haematology and serum chemistry were statistically different but within normal physiological ranges. No obvious abnormalities were found in the necropsies and histopathological observations. In conclusion, the compound Chinese herbal medicine mouthwash promoted oral ulcer healing in rats with no obvious sub chronic toxicity, providing a potential alternative therapeutic strategy for oral ulcers.

Role of Enterococcus faecalis in refractory apical periodontitis: from pathogenicity to host cell response.

BACKGROUND: Refractory apical periodontitis (RAP) is an oral infectious disease characterised by persistent inflammation, progressive alveolar bone destruction, and delayed bone healing. RAP has received increasing attention, because it cannot be cured after repeated root canal therapies. The aetiology of RAP is related to the complex interplay between the pathogen and its host. However, the exact pathogenesis of RAP remains unclarified and includes several factors, such as microorganism immunogenicity, host immunity and inflammation, and tissue destruction and repair. Enterococcus faecalis is the dominant pathogen involved in RAP, and has evolved multiple strategies to ensure survival, which cause persistent intraradicular and extraradicular infections. OBJECTIVE: To review the crucial role of E. faecalis in the pathogenesis of RAP, and open new avenues for prevention and treatment of RAP. METHODS: The PubMed and Web of Science databases were searched for pertinent publications, employing the search terms "Enterococcus faecalis", "refractory apical periodontitis", "persistent periapical periodontitis", "pathogenicity", "virulence", "biofilm formation", "dentine tubule", "immune cell", "macrophage", and "osteoblast". RESULTS AND CONCLUSION: Besides its high pathogenicity due to various virulence mechanisms, E. faecalis modulates the macrophage and osteoblast responses, including regulated cell death, cell polarisation, cell differentiation, and inflammatory response. An in-depth understanding of the multifaceted host cell responses modulated by E. faecalis will help to design potential future therapeutic strategies and overcome the challenges of sustained infection and delayed tissue healing in RAP.

Comparison of the effect of iRoot BP Plus and MTA on direct pulp capping of carious pulp exposed mature permanent teeth / 口腔疾病防治

Objective@#To study the clinical efficacy of biological ceramics (iRoot BP Plus ) and mineral trioxide aggregate (MTA) in direct pulp capping of mature permanent teeth, to provide referrence for clinical application.@* Methods @# Seventy-four patients with pulp exposure due to deep caries or reversible pulpitis in 75 mature permanent teeth were selected and were randomly divided into two groups. iRoot BP Plus were used as pulp capping agents in the treatment group and MTA were used as pulp capping agents in the control group respectively. The clinical efficacy and imaging analysis were performed at 1, 3, 6 and 12 months after operation. Treatment success rate of the two groups were calculated, and the influence of various factors including gender, age, tooth position, cavity, number and size of pulp exposure on the efficacy of direct pulp capping were analyzed. @* Results @#Sixty patients with 61 mature permanent teeth were selected. Twelve mouths after treatment, 61 teeth of 60 patents were completely investigated (iRoot group: 31 teeth 30 patients; MTA group: 30 teeth 30 patients). The success rates of the 2 groups were 90.3% (iRoot BP Plus) and 90.0% (MTA), respectively. There was no statistical difference between 2 groups (P>0.05). Statistical analysis also showed that gender, age, tooth position, cavity, number and size of pulp exposure had no significant difference between the two groups (P>0.05). @*Conclusion@# Both iRoot BP Plus and MTA are effective in direct pulp capping of mature permanent teeth with carious pulp exposure, while the operation of iRoot is simple and convenient.

Enterococcus faecalis-Induced Macrophage Necroptosis Promotes Refractory Apical Periodontitis.

The persistence of residual bacteria, particularly Enterococcus faecalis, contributes to refractory periapical periodontitis, which still lacks effective therapy. The role of receptor-interacting protein kinase 3 (RIPK3)- and mixed lineage kinase domain-like protein (MLKL)-mediated necroptosis, a highly proinflammatory form of regulated cell death, has recently drawn much attention. However, the role of necroptosis in the pathogenesis of refractory periapical periodontitis remains unclear. We investigated whether the RIPK3/MLKL signaling pathway was activated in periapical lesion specimens obtained from patients diagnosed with refractory periapical periodontitis. RIPK3-deficient mice were then used to determine the role of necroptosis under this condition in vivo. We found that the phosphorylation levels of RIPK3 and MLKL were elevated in periapical lesion specimens of patients with refractory periapical periodontitis. In addition, necroptosis was induced in an E. faecalis-infected refractory periapical periodontitis mouse model, in which inhibition of necroptosis by RIPK3 deficiency could markedly alleviate inflammation and bone destruction. Moreover, double-labeling immunofluorescence suggested that macrophage necroptosis may be involved in the development of refractory periapical periodontitis. Then, we established an in vitro macrophage infection model with E. faecalis. E. faecalis infection was found to induce necroptotic cell death in macrophages through the RIPK3/MLKL signaling pathway, which was markedly alleviated by the RIPK3- or MLKL-specific inhibitor. Our study revealed that RIPK3/MLKL-mediated macrophage necroptosis contributes to the development of refractory periapical periodontitis and suggests that inhibitors or treatments targeting necroptosis represent a plausible strategy for the management of refractory periapical periodontitis. IMPORTANCE Oral infectious diseases represent a major neglected global population health challenge, imposing an increasing burden on public health and economy. Refractory apical periodontitis (RAP), mainly caused by Enterococcus faecalis, is a representative oral infectious disease with considerable therapeutic challenges. The interplay between E. faecalis and the host often leads to the activation of programmed cell death. This study identifies an important role of macrophage necroptosis induced by E. faecalis in the pathogenesis of RAP. Manipulating RIPK3/MLKL-mediated necroptosis may represent novel therapeutic targets, not only for RAP but also for other E. faecalis-associated infectious diseases.

Effects of N-cadherin silencing on the proliferation and migration of human dental pulp stem cells / 口腔疾病防治

Objective @#To investigate the effects of N-cadherin silencing on the proliferation and migration of human dental pulp stem cells (DPSCs) and to provide experimental evidence for DPSCs-based dental pulp regeneration.@* Methods@# DPSCs were transfected with N-cadherin shRNA lentivirus, and the knockdown efficiency of N-cadherin at both the mRNA and protein levels was confirmed by qRT-PCR and Western blot. The experiment included a negative control group (shRNA -NC) and an N-cadherin shRNA silencing group. Cell proliferation was detected by the CCK-8 method. Cell cycle and apoptosis were assessed by flow cytometry, and cell migration was detected using the Transwell method.@*Results@#N-cadherin shRNA significantly reduced the expression levels of N-cadherin mRNA and protein in DPSCs (P<0.001). The proliferation activity of the N-cadherin shRNA group was significantly greater than that of the shRNA-NC group on the 3rd and 4th days after cell inoculation and lower than that of the shRNA-NC group from the 6th to 8th days (P<0.05). On the 3rd day after cell inoculation, the proportion of cells in S phase and G2 phase in the N-cadherin shRNA group was greater than that in the shRNA-NC group (P<0.05). On the 6th day after cell inoculation, the proportion of cells in S phase and G2 phase in the N-cadherin shRNA group was lower than that in the shRNA-NC group (P<0.05), and the proportion of apoptotic cells in the N-cadherin shRNA group was greater than that in the shRNA-NC group (P<0.01). Low densities cells and high densities cells were inoculated into Transwell upper chamber for 20 h, the number of cells passing through the membrane pores of upper chamber in the N-cadherin shRNA group was greater than that in the shRNA-NC group (P<0.001).@*Conclusion@#Silencing N-cadherin expression can promote the early proliferation and migration of DPSCs.

PANoptosis: A New Insight Into Oral Infectious Diseases.

The oral microbiome, one of the most complex and intensive microbial ecosystems in the human body, comprises bacteria, archaea, fungi, protozoa, and viruses. Dysbiosis of the oral microbiome is the initiating factor that leads to oral infectious diseases. Infection is a sophisticated biological process involving interplay between the pathogen and the host, which often leads to activation of programmed cell death. Studies suggest that pyroptosis, apoptosis, and necroptosis are involved in multiple oral infectious diseases. Further understanding of crosstalk between cell death pathways has led to pyroptosis, apoptosis, and necroptosis being integrated into a single term: PANoptosis. PANoptosis is a multifaceted agent of the immune response that has important pathophysiological relevance to infectious diseases, autoimmunity, and cancer. As such, it plays an important role in innate immune cells that detect and eliminate intracellular pathogens. In addition to the classical model of influenza virus-infected and Yersinia-infected macrophages, other studies have expanded the scope of PANoptosis to include other microorganisms, as well as potential roles in cell types other than macrophages. In this review, we will summarize the pathophysiological mechanisms underlying inflammation and tissue destruction caused by oral pathogens. We present an overview of different pathogens that may induce activation of PANoptosis, along with the functional consequences of PANoptosis in the context of oral infectious diseases. To advance our understanding of immunology, we also explore the strategies used by microbes that enable immune evasion and replication within host cells. Improved understanding of the interplay between the host and pathogen through PANoptosis will direct development of therapeutic strategies that target oral infectious diseases.

Efficacy of i-PRF in regenerative endodontics therapy for mature permanent teeth with pulp necrosis: study protocol for a multicentre randomised controlled trial.

BACKGROUND: Dental pulp necrosis, a common health problem, is traditionally treated with root canal therapy; however, it fails in restoring the vitality of damaged pulp. Most studies regarding regenerative endodontic therapy (RET) are limited to the treatment of immature necrotic teeth. Given that injectable platelet-rich fibrin (i-PRF) has shown great potential in regenerative medicine as a novel platelet concentration, this study is designed to explore whether i-PRF can serve as a biological scaffold, extending the indications for RET and improving the clinical feasibility of RET in mature permanent teeth with pulp necrosis. METHODS: This is a randomised, double-blind, controlled, multicentre clinical trial designed to evaluate the clinical feasibility of RET for mature permanent teeth with pulp necrosis and to compare the efficacy of i-PRF and blood clots as scaffolds in RET. A total of 346 patients will be recruited from three centres and randomised at an allocation ratio of 1:1 to receive RET with either a blood clot or i-PRF. The changes in subjective symptoms, clinical examinations, and imaging examinations will be tracked longitudinally for a period of 24 months. The primary outcome is the success rate of RET after 24 months. The secondary outcome is the change in pulp vitality measured via thermal and electric pulp tests. In addition, the incidence of adverse events such as discolouration, reinfection, and root resorption will be recorded for a safety evaluation. DISCUSSION: This study will evaluate the clinical feasibility of RET in mature permanent teeth with pulp necrosis, providing information regarding the efficacy, benefits, and safety of RET with i-PRF. These results may contribute to changes in the treatment of pulp necrosis in mature permanent teeth and reveal the potential of i-PRF as a novel biological scaffold for RET. TRIAL REGISTRATION: ClinicalTrials.gov NCT04313010 . Registered on 19 March 2020.

N-Cadherin Regulates the Odontogenic Differentiation of Dental Pulp Stem Cells via ß-Catenin Activity.

Dental pulp stem cell (DPSC) transplantation has shown new prospects in dental pulp regeneration, and is of great significance in the treatment of pulpitis and pulp necrosis. The fate and regenerative potential of stem cells are dependent, to a great extent, on their microenvironment, which is composed of various tissue components, cell populations, and soluble factors. N-cadherin-mediated cell-cell interaction has been implicated as an important factor in controlling the cell-fate commitment of mesenchymal stem cells. In this study, the effect of N-cadherin on odontogenic differentiation of DPSCs and the potential underlying mechanisms, both in vitro and in vivo, was investigated using a cell culture model and a subcutaneous transplantation mouse model. It was found that the expression of N-cadherin was reversely related to the expression of odontogenic markers (dentin sialophosphoprotein, DSPP, and runt-related transcription factor 2, Runx2) during the differentiation process of DPSCs. Specific shRNA-mediated knockdown of N-cadherin expression in DPSCs significantly increased the expression of DSPP and Runx2, alkaline phosphatase (ALP) activity, and the formation of mineralized nodules. Notably, N-cadherin silencing promoted nucleus translocation and accumulation of ß-catenin. Inhibition of ß-catenin by a specific inhibitor XAV939, reversed the facilitating effects of N-cadherin downregulation on odontogenic differentiation of DPSCs. In addition, knockdown of N-cadherin promoted the formation of odontoblast-like cells and collagenous matrix in ß-tricalcium phosphate/DPSCs composites transplanted into mice. In conclusion, N-cadherin acted as a negative regulator via regulating ß-catenin activity during odontogenic differentiation of DPSCs. These data may help to guide DPSC behavior by tuning the N-cadherin-mediated cell-cell interactions, with implications for pulp regeneration.

Dentinal tubule occlusion using Er:YAG Laser: an in vitro study.

OBJECTIVES: We analyzed the effects of the Er:YAG laser used with different parameters on dentinal tubule (DT) occlusion, intrapulpal temperature and pulp tissue morphology in order to determine the optimal parameters for treating dentin hypersensitivity. METHODOLOGY: Dentin specimens prepared from 36 extracted human third molars were randomized into six groups according to the treatment method (n=6 each): control (A); Gluma desensitizer (B); and Er:YAG laser treatment at 0.5 W , 167 J/cm2 (50 mJ, 10 Hz) (C), 1 W , 334 J/cm2 (50 mJ, 20 Hz) (D), 2 W , 668 J/cm2 (100 mJ, 20 Hz) (E), and 4 W and 1336 J/cm2 (200 mJ, 20 Hz) (F). Treatment-induced morphological changes of the dentin surfaces were assessed using scanning electron microscopy (SEM) to find parameters showing optimal dentin tubule occluding efficacy. To further verify the safety of these parameters (0.5 W, 167 J/cm2), intrapulpal temperature changes were recorded during laser irradiation, and morphological alterations of the dental pulp tissue were observed with an upright microscope. RESULTS: Er:YAG laser irradiation at 0.5 W (167 J/cm2) were found to be superior in DT occlusion, with an exposure rate significantly lower than those in the other groups (P<0.05). Intrapulpal temperature changes induced by Er:YAG laser irradiation at 0.5 W (167 J/cm2) with (G) and without (H) water and air cooling were demonstrated to be below the threshold. Also, no significant morphological alterations of the pulp and odontoblasts were observed after irradiation. CONCLUSION: Therefore, 0.5 W (167 J/cm2) is a suitable parameter for Er:YAG laser to occlude DTs, and it is safe to the pulp tissue.

Effects of a derivative of reutericin 6 and gassericin A on the biofilm of Streptococcus mutans in vitro and caries prevention in vivo.

It is known that Streptococcus mutans (S. mutans) is the leading cariogenic pathogen. Recently, an increasing number of antimicrobial peptides (AMPs) have been brought into consideration as anti-caries agents. Here, we designed and synthesized an AMP derived from reutericin 6 and/or gassericin A, named LN-7, and explored its effect on biofilm of S. mutans UA159 in vitro and development of dental caries in vivo. Antibacterial assays showed that LN-7 was more active against S. mutans (3.2 µM) than many peptide-based agents, capable of killing other types of Streptococci in oral cavity. In addition, LN-7 presented fast killing kinetics, with more than 97% S. mutans killed within 5 min. The mechanism of the antimicrobial activity mainly lies on the disruption of bacterial membrane. Effects of LN-7 on the biofilm formation and the viability of preformed biofilm were quantified by crystal violet staining, which showed that LN-7 could effectively inhibit the biofilm accumulation of S. mutans. Moreover, the biofilm of S. mutans treated with LN-7 displayed notable changes in bacterial viability and morphology, observed by confocal laser scanning microscopy and scanning electron microscopy. In addition, topical oral treatment with LN-7 could suppress the development of dental caries in vivo, reducing the occurrence of severe dental lesion in a rodent model. These results reveal a new peptide-based agent as a topical treatment for dental caries, opening the door to clinical studies to explore its potential for caries prevention.

Dentinal tubule occlusion using Er:YAG Laser: an in vitro study

Abstract Objectives We analyzed the effects of the Er:YAG laser used with different parameters on dentinal tubule (DT) occlusion, intrapulpal temperature and pulp tissue morphology in order to determine the optimal parameters for treating dentin hypersensitivity. Methodology Dentin specimens prepared from 36 extracted human third molars were randomized into six groups according to the treatment method (n=6 each): control (A); Gluma desensitizer (B); and Er:YAG laser treatment at 0.5 W , 167 J/cm2 (50 mJ, 10 Hz) (C), 1 W , 334 J/cm2 (50 mJ, 20 Hz) (D), 2 W , 668 J/cm2 (100 mJ, 20 Hz) (E), and 4 W and 1336 J/cm2 (200 mJ, 20 Hz) (F). Treatment-induced morphological changes of the dentin surfaces were assessed using scanning electron microscopy (SEM) to find parameters showing optimal dentin tubule occluding efficacy. To further verify the safety of these parameters (0.5 W, 167 J/cm2), intrapulpal temperature changes were recorded during laser irradiation, and morphological alterations of the dental pulp tissue were observed with an upright microscope. Results Er:YAG laser irradiation at 0.5 W (167 J/cm2) were found to be superior in DT occlusion, with an exposure rate significantly lower than those in the other groups (P<0.05). Intrapulpal temperature changes induced by Er:YAG laser irradiation at 0.5 W (167 J/cm2) with (G) and without (H) water and air cooling were demonstrated to be below the threshold. Also, no significant morphological alterations of the pulp and odontoblasts were observed after irradiation. Conclusion Therefore, 0.5 W (167 J/cm2) is a suitable parameter for Er:YAG laser to occlude DTs, and it is safe to the pulp tissue.

Effect of ciliary neurotrophic factor on neural differentiation of stem cells of human exfoliated deciduous teeth.

The stem cells of human exfoliated deciduous teeth (SHEDs) are considered to be one of the main sources of seed cells in stem cell therapy. The aim of this study was to examine the effect of ciliary neurotrophic factor (CNTF) on neurogenic differentiation of SHEDs. With the consent of parents, SHEDs from 6 to 8 year old children were isolated and cultured. The mesenchymal stemness and the potential of multidirectional (adipogenic and osteogenic) differentiation for the isolated SHEDs were firstly determined. The effect of CNTF on specific neurogenic differentiation of SHEDs was then examined by detecting the expression of marker genes and proteins via RT-PCR, immunoblotting, and immunofluorescence microscopy. The isolated SHEDs expressed specific surface markers of mesenchymal stem cells, and their potential of osteogenic and adipogenic differentiation were confirmed. CNTF promoted the differentiation of SHEDs into neuron-like cells with a high expression of acetylcholine transferase (CHAT), a marker of cholinergic neurons. The expression of other neuron markers including nestin, microtubule-associated protein 2 (MAP 2), and ß-tublin III was also detected. Interestingly, the expression of neurogenic markers was maintained at a high level after neurogenic induction. SHEDs can be induced by CNTF to differentiate into cholinergic neuron-like cells under appropriate culture conditions. Our findings have laid a foundation for future use of SHEDs to treat neurological diseases.

Helicobacter pylori inhibited cell proliferation in human periodontal ligament fibroblasts through the Cdc25C/CDK1/cyclinB1 signaling cascade.

PURPOSE: Several studies have shown that the oral cavity is a secondary location for Helicobacter pylori colonization and that H. pylori is associated with the severity of periodontitis. This study investigated whether H. pylori had an effect on the periodontium. We established an invasion model of a standard strain of H. pylori in human periodontal ligament fibroblasts (hPDLFs), and evaluated the effects of H. pylori on cell proliferation and cell cycle progression. METHODS: Different concentrations of H. pylori were used to infect hPDLFs, with 6 hours of co-culture. The multiplicity of infection in the low- and high-concentration groups was 10:1 and 100:1, respectively. The Cell Counting Kit-8 method and Ki-67 immunofluorescence were used to detect cell proliferation. Flow cytometry, quantitative real-time polymerase chain reaction, and western blots were used to detect cell cycle progression. In the high-concentration group, the invasion of H. pylori was observed by transmission electron microscopy. RESULTS: It was found that H. pylori invaded the fibroblasts, with cytoplasmic localization. Analyses of cell proliferation and flow cytometry showed that H. pylori inhibited the proliferation of periodontal fibroblasts by causing G2 phase arrest. The inhibition of proliferation and G2 phase arrest were more obvious in the high-concentration group. In the low-concentration group, the G2 phase regulatory factors cyclin dependent kinase 1 (CDK1) and cell division cycle 25C (Cdc25C) were upregulated, while cyclin B1 was inhibited. However, in the high-concentration group, cyclin B1 was upregulated and CDK1 was inhibited. Furthermore, the deactivated states of tyrosine phosphorylation of CDK1 (CDK1-Y15) and serine phosphorylation of Cdc25C (Cdc25C-S216) were upregulated after H. pylori infection. CONCLUSIONS: In our model, H. pylori inhibited the proliferation of hPDLFs and exerted an invasive effect, causing G2 phase arrest via the Cdc25C/CDK1/cyclin B1 signaling cascade. Its inhibitory effect on proliferation was stronger in the high-concentration group.

[Design, screening and antimicrobial activity of novel peptides against Streptococcus mutans].

OBJECTIVE: To construct antimicrobial peptides with potent antimicrobial activity, low cytotoxicity and efficient killing rate of Streptococcus mutans for prevention and treatment of dental caries. METHODS: We exploited the existing design strategies to modify reutericin 6 or gassericin A produced by Lactobacillus species in the oral cavity based on their cationicity, amphipathicity and α-helical structure. We examined their antimicrobial activities using bacterial susceptibility assay, their cytotoxicity through cytotoxicity assay and their killing rate of Streptococcus mutans with time-kill assay. We further evaluated the candidate derivatives for their killing rate against Streptococcus mutans, their antimicrobial activity against different oral pathogens and the development of drug resistance. RESULTS: We constructed 6 AT-1 derivatives, among which AT-7 showed an MIC of 3.3 µmol/L against Streptococcus mutans, Porphyromonas gingivalis and Actinomyces viscosus with a killing rate of 88.7% against Streptococcus mutans within 5 min. We did not obtain de novo strains of Streptococcus mutans resistant to AT- 7 after induction for 10 passages. CONCLUSIONS: Hydrophobicity and imperfect amphipathic structure are two key parameters that define the antimicrobial potency of the antimicrobial peptides. The imperfectly amphipathic peptide AT-7 shows the potential for clinical application in dental caries treatment.

Rational design of peptides with enhanced antimicrobial and anti-biofilm activities against cariogenic bacterium Streptococcus mutans.

Streptococcus mutans (S. mutans) is known to be a leading cariogenic pathogen in the oral cavity. Antimicrobial peptides possess excellent properties to combat such pathogens. In this study, we compared the antimicrobial activity of novel linear reutericin 6- and/or gassericin A-inspired peptides and identified LR-10 as the leading peptide. Antibacterial assays demonstrate that LR-10 is more active against S. mutans (3.3 µM) than many peptide-based agents without resistance selection, capable of killing many oral pathogens, and tolerant of physiological conditions. LR-10 also presented a faster killing rate than chlorhexidine and erythromycin, and appeared to display selective activity against S. mutans within 10 s. S. mutans is usually encased in plaque biofilms. Biofilm inhibitory assays indicated that LR-10 had excellent inhibitory effect on the biofilm formation of S. mutans and biofilm-encased cells in vitro at low concentrations (6.5 µM). Consistent with most peptides, LR-10 kills S. mutans mainly by disrupting the cell membranes. Notably, both hemolytic activity assays and cytotoxicity tests indicated that LR-10 could keep biocompatible at the effective concentrations. Hence, LR-10 could be a good candidate for clinical treatment of dental caries.

IGF2BP1 promotes LPS-induced NFκB activation and pro-inflammatory cytokines production in human macrophages and monocytes.

BACKGROUND: Lipopolysaccharide (LPS)-induced macrophage/monocyte activation and pro-inflammatory cytokines production are important mediators for periodontitis progression. The current study tested the potential role of insulin-like growth factor 2 mRNA-binding protein 1 (IGF2BP1) in the process. METHODS: THP-1 human macrophages and primary human peripheral blood mononuclear cells (PBMCs) were treated with LPS. mRNA and protein expression of IGF2BP1 were tested by qPCR and Western blotting assay. IGF2BP1 expression was altered by shRNAs or CRISPR/Cas-9 gene editing methods. LPS-induced cytokine production was tested by ELISA assay. Cytokine mRNA expression was tested by the quantitative real-time reverse transcriptase polymerase chain reaction (qPCR) assay. RESULTS: In THP-1 human macrophages and PBMCs, treatment with LPS induced mRNA and protein expression of IGF2BP1. IGF2BP1 silencing (by targeted shRNAs) or CRISPR/Cas-9 knockout largely inhibited LPS-induced production of multiple pro-inflammatory cytokines, including tumor necrosis factor-α (TNF-α), interleukin (IL)-1ß and IL-6. Conversely, forced over-expression of IGF2BP1 facilitated LPS-induced pro-inflammatory cytokines production in THP-1 cells. For the mechanism study, we show that IGF2BP1 co-immunoprecipitated with p65-p52 nuclear factor kappa B (NFκB) complex in nuclei of LPS-treated THP-1 cells. Significantly, LPS-induced p65-p52 nuclear translocation and NFκB activation were inhibited by IGF2BP1 silencing or CRISPR/Cas-9 knockout. CONCLUSION: IGF2BP1 promotes LPS-induced NFκB signalling and transcriptional activation in human macrophages and monocytes.

A Membrane-Targeted Peptide Inhibiting PtxA of Phosphotransferase System Blocks Streptococcus mutans.

Streptococcus mutans, the primary cause of dental caries, takes up carbohydrates through the phosphoenolpyruvate sugar phosphotransferase system (PTS). This study aimed to identify a novel membrane-targeted antimicrobial peptide (AMP) that could also target the L-ascorbate-specific PtxA component of the S. mutans PTS system. C10-KKWW was identified and selected using virtual screening of a lipopeptide library, a minimum inhibiting concentration (MIC) assay, cytotoxicity assays and a hemolysis assay. Surface plasmon resonance confirmed that C10-KKWW had a high binding affinity for PtxA. Combining with scanning electron microscopy and cell permeability assay, it was shown that the effects of C10-KKWW could be attributed to both membrane and PtxA. Wild type (WT) S. mutans, a ptxA deletion mutant (ΔptxA), and a mutant-complemented strain (CptxA), were cultured consistently in brain heart infusion (BHI) medium, tryptone-vitamin medium supplemented with 15 mM L-ascorbate (TVL), or for 5 h in BHI supplemented with 7.4 mM sodium L-ascorbate. Compared to ∆ptxA, in WT S. mutans and CptxA, C10-KKWW had a stronger MIC (3.9 µg/mL), and distinctively decreased biofilm viability. The extracellular concentrations of L-ascorbate/sodium L-ascorbate were not changed before and after WT treated with C10-KKWW. L-ascorbate-induced operon genes, or other PTS genes, were significantly suppressed by C10-KKWW. In conclusion, C10-KKWW has been developed; it acts through interaction with the bacterial membrane and interferes with L-ascorbate translocation to inhibit S. mutans growth and eradicate its biofilm. C10-KKWW may be especially effective at optimal oral ascorbate levels. A combination of C10-KKWW with sodium L-ascorbate might also be a novel strategy for dental caries treatment.