Effects of enamel adhesives with different components on Porphyromonas gingivalis and Streptococcus mutans / 中国组织工程研究

BACKGROUND:The light curing and fluoride light curing enamel adhesives have a certain sealing effect on the etched enamel surface.The fluoride light curing enamel adhesives can also achieve the anti-caries function by releasing fluoride ions.However,the existing researches lack the long-term tracing of fluoride release effect,especially the amount of local pathogenic bacteria after 1-3 months of local fluoride application. OBJECTIVE:To analyze the changes in the expression of Porphyromonas gingivalis and Streptococcus mutans in subgingival plaque of the upper anterior teeth adhered by different components of enamel adhesives in adolescent patients with fixed appliance. METHODS:Ninety adolescent patients who received orthodontic treatment in Shanghai Stomatological Hospital from January to December 2016 were enrolled,including 43 males and 47 females,with a mean age of(13.27±1.12)years.These patients were randomly divided into three groups(n=30 per group).In the chemical curing group,Unite? bonding resin was used to bond fixed appliances.In the light curing group,Transbond XT light curing resin was used to bond the fixed appliance.In the fluoride light curing group,GC light curing orthodontic adhesive was used to bond the fixed appliance with cement.The subgingival plaque was collected on the day of bonding,1st,2nd,and 3rd month follow-up reviews.The expressions of Porphyromonas gingivalis and Streptococcus mutans in subgingival plaque were detected by PCR. RESULTS AND CONCLUSION:(1)Intragroup comparison:With the increase of bonding time,Porphyromonas gingivalis expression increased significantly in the 3rd month in the chemical curing group(P<0.05).In the light curing group,Porphyromonas gingivalis showed a significant decrease in the 1st month(P<0.05).Porphyromonas gingivalis expression decreased significantly in the 1st and 2nd months compared with initial data in the fluoride light curing group(P<0.05).The expression of Streptococcus mutans was higher in the chemical curing group in the 1st,2nd,and 3rd months compared with the initial data(P<0.05).In the fluoride light curing group,the expression of Streptococcus mutans was lower in the 1st,2nd,and 3rd months compared with the initial data(P<0.05).There was no significant difference in the proliferation and expression of Streptococcus mutans during follow-up in the light curing group compared to the initial adhesion(P>0.05).(2)Intergroup comparison:In the 1st month,the expression of Porphyromonas gingivalis was lower in the light curing group and fluoride light curing group than that in the chemical curing group(P<0.05).In the 2nd and 3rd months,the expression of Porphyromonas gingivalis was lower in the fluoride light curing group than that in the light curing group and chemical curing group(P<0.05).In the 1st,2nd,and 3rd months,the expression of Streptococcus mutans was lower in the light curing group and fluoride light curing group than that in the chemical curing group(P<0.05).The expression of Streptococcus mutans was lower in the fluoride light curing group than that in the light curing group(P<0.05).(3)The results show that in the fixed orthodontic process,the use of different components of enamel adhesives has different effects on the proliferation and expression of oral Porphyromonas gingivalis and Streptococcus mutans in the short term.Fluoride light curing enamel adhesives at the initial stage can reduce the occurrence of enamel demineralization,caries,and periodontal inflammation.

Effect and mechanism of reactive oxygen species-responsive nanoparticles on the regulation of human gingival fibroblast function and inflammation induced by lipopolysaccharide / 口腔疾病防治

Objective@#To investigate the effects of PssL-NAC reactive oxygen species (ROS)-responsive nanoparticles on intracellular ROS production, inflammatory factor levels, collagen production, cell function and Toll-like receptor 4 (TLR4), NF-κB nuclear factor-κB (p65) pathway protein expression in human gingival fibroblasts (HGFs) induced by Porphyromonas gingivalis-lipopolysaccharide (P.g-LPS).@*Methods@#This study was reviewed and approved by the ethics committee. PssL-NAC microspheres containing oil soluble antioxidant N-acetylcysteine (NAC) were obtained by connecting the hydrophobic end of polycaprolactone (PCL) and the hydrophilic end of polyethylene glycol (PEG) via thioketal (TK) bonds in response to ROS, and self loading in the aqueous and oil phases. After preparation of the PssL-NAC microspheres and aqueous NAC solution, successful synthesis of the nanoparticles was verified by transmission electron microscopy. Then, HGFs were exposed to P.g-LPS (0, 5, or 10 μg/mL), P.g-LPS (0, 5, or 10 μg/mL)+NAC, and P.g-LPS (0, 5, or 10 μg/mL)+PssL-NAC, and the ROS levels in the different groups were observed under confocal microscopy to determine the concentration of P.g-LPS for use in subsequent experiments. The groups were as follows: control group (no treatment), P.g-LPS group (HGFs treated with P.g-LPS), NAC group (HGFs treated with P.g-LPS and NAC), and PssL-NAC group (HGFs treated with P.g-LPS and PssL-NAC). Cell counting kit-8 (CCK-8) assays verified the biosafety of PssL-NAC. The ROS levels in the different groups were detected by DCFH-DA probes and observed via confocal microscopy. Real-time qPCR (RT-qPCR) was used to monitor the gene expression levels of the intracellular inflammatory cytokines interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), collagen 1 (COL1) and collagen 3 (COL3). The effect of PssL-NAC on the migration of HGFs was observed via the scratch test. The protein expression of TLR4-NF-κB, and phosphorylated p65 (p-p65) in the TLR4-NF-κB pathway was evaluated by Western blot.@*Results@#PssL-NAC had no significant effect on HGF proliferation (P>0.05). At elevated P.g-LPS concentrations, PssL-NAC maintained intracellular ROS levels approximately twice those in the control group (P<0.001). PssL-NAC significantly decreased P.g-LPS-induced IL-6 (P<0.001) and TNF-α (P<0.001) gene expression and increased COL1 gene expression (P<0.001). After P.g-LPS stimulation, PssL-NAC restored cell migration to the control level (P>0.05) and decreased the protein expression of TLR4 (P<0.001), p65 (P = 0.006), and p-p65 (P = 0.017) in the TLR4-NF-κB pathway.@*Conclusion@#PssL-NAC maintains the appropriate intracellular ROS concentration, alleviates P.g-LPS-induced inflammation in HGFs through the TLR4-NF-κB pathway, and restores the cell functions of collagen production and migration in an inflammatory environment.

Prevalencia de Porphyromonas gingivalis en fluido gingival y su relación con la periodontitis / Prevalence of Porphyromonas gingivalis in gingival fluid and its relationship with periodontitis

Introducción: la periodontitis es una enfermedad infecciosa multifactorial asociada a un biofilm de microorganismos patógenos. Objetivo: el objetivo del trabajo fue establecer la prevalencia de Porphyromonas gingivalis en pacientes con periodontitis y relacionarla con la severidad de la enfermedad. Material y métodos: participaron 45 pacientes, sistémicamente saludables, con edades entre 35 y 65 años. El grado de periodontitis se definió según los criterios de Papapanou y colaboradores. Como grupo control, se incluyeron 20 sujetos de ambos sexos sin periodontitis y sin enfermedades sistémicas. Se tomaron muestras de fluido gingival en dos sitios más profundos. Porphyromonas gingivalis se detectó por PCR (reacción en cadena de la polimerasa). Resultados: la frecuencia relativa de periodontitis fue de 13.3% grado I, 46.7% grado II y 40% grado III. El sexo masculino presentó periodontitis grado III 72.2% y grado II 52.3%. El grado I se registró con mayor frecuencia en el sexo femenino, 66.7%. La prevalencia de Porphyromonas gingivalis en la población con periodontitis fue de 44.4%. Se obtuvieron diferencias estadísticamente significativas entre los grados de severidad de periodontitis y la presencia de Porphyromonas gingivalis (p = 0.0002, α = 5%). Conclusión: la periodontitis predominó en el sexo masculino. La prevalencia de Porphyromonas gingivalis en la población con periodontitis crónica fue de 44.4% y su presencia está relacionada con la severidad (AU)

Introduction: periodontitis is a multifactorial infectious disease associated with a biofilm of pathogenic microorganisms. Objective: the objective of the work was to establish the prevalence of Porphyromonas gingivalis in patients with periodontitis and relate it to the severity of the disease. Material and methods: 45 systemically healthy patients, aged between 35 and 65 years old, participated. The degree of periodontitis was defined according to the criteria of Papapanou et al. As a control group, 20 patients of both sexes without periodontitis and without systemic diseases were included. Gingival fluid samples were taken from two deeper sites. Porphyromonas gingivalis was detected by PCR (polymerase chain reaction). Results: the relative frequency of periodontitis was 13.3% grade I, 46.7% grade II and 40% grade III. The male sex presented periodontitis grade III 72.2% and grade II 52.3%. Grade I was recorded more frequently in the female sex, 66.7%. The prevalence of Porphyromonas gingivalis in the population with periodontitis was 44.4%. Statistically significant differences were obtained between the degrees of severity of periodontitis and the presence of Porphyromonas gingivalis (p = 0.0002, α = 5%). Conclusion: periodontitis predominated in males. The prevalence of Porphyromonas gingivalis in the population with chronic periodontitis was 44.4% and its presence is related to severity (AU)

Immunological and bacteriological monitoring of periodontal tissue in pregnancy

We determine periodontal pathogens in periodontal pockets from pregnant women with periodontitis and associate it to the C reactive protein (CRP), nitrates, immunoglobulin A and G (Ig A and G), and myeloperoxidase (MPO) levels in saliva to identify some biomarkers as tools to predict the periodontal status from pregnant. The samples were obtained from periodontal pockets and saliva from 100 pregnant women (PW) and 50 non-pregnant women (NPW). Every patient was evaluated by: 1) probing depth (PD) and loss of clinical attachment level (CAL); 2) in saliva; CRP, MPO, Ig A and G) and nitrite concentrations, 3) in periodontal pockets: P.gingivalis, T.forsythia, T.denticola, P.intermedia, A.actinomycetemcomitans. InfoStat/P 2008 software was used with a p-value <0.05. Clinical parameters showed stages I and II of PD in both groups. P.intermedia and A.actinomycetemcomitans were observed only in periodontal pockets from PW. The CAL was higher in pregnant of the 3rd trimester than in the other stages and was associated with low levels of IgA and the presence of P.intermedia and T. forsythia in the same trimester. The levels of IgA in saliva would reflect the immunological situation in pregnant women. This could be used to monitor the immune status of the gingival tissues during pregnancy.

Determinamos patógenos periodontales en bolsas periodontales de gestantes con periodontitis y lo asociamos a los niveles de proteína C reactiva (PCR), nitratos, inmunoglobulina A y G (Ig A y G) y mieloperoxidasa (MPO) en saliva para identificar biomarcadores como herramientas para predecir el estado periodontal de la gestante. Las muestras se obtuvieron de bolsas periodontales y saliva de 100 mujeres embarazadas (ME) y 50 mujeres no embarazadas (NoE). Cada paciente fue evaluado por: 1) profundidad de sondaje(PD) y pérdida del nivel de inserción clínica (NIC); 2) en saliva; PCR, MPO, Ig A y G y concentraciones de nitritos, 3) en bolsas periodontales: P.gingivalis, T.forsythia, T.denticola, P.intermedia, A.actinomycetemcomitans. Se utilizó el software InfoStat/P 2008 con un valor de p<0,05. Los parámetros clínicos mostraron estadios I y II de EP en ambos grupos. P.intermedia y A.actinomycetemcomitans se observaron solo en bolsas periodontales de ME. El NIC fue mayor en gestantes del 3er trimestre que en las demás etapas y se asoció con niveles bajos de IgA y presencia de P.intermedia y T.forsythia en el mismo trimestre. Los niveles de IgA en saliva reflejarían la situación inmunológica en la mujer embarazada. Esto podría usarse para monitorear el estado inmunológico de los tejidos gingivales durante el embarazo.

Research progress on the molecular mechanism of periodontal pathogens promoting the development of cardiovascular diseases / 口腔疾病防治

@#Periodontitis is a multifactorial infectious and inflammatory disease occurring in tooth-supporting tissues. In recent decades, many studies have reported a potential relationship between periodontitis and cardiovascular disease, and periodontal pathogens are an important factor linking periodontitis and cardiovascular disease. In this review, we summarize updated preclinical studies and epidemiological evidence on the association of these two diseases. Moreover, possible mechanisms accounting for such links are introduced, including bacteremia and direct invasion of pathogens, endotoxemia caused by virulence factors of periodontal pathogens leading to systemic inflammation, abnormal lipid metabolism and oxidative stress, which further affect the inflammatory states of the cardiovascular system. The molecular mimicry theory and the intrinsic correlation of apolipoprotein E between periodontitis and cardiovascular disease require further study. Combined with existing studies, it is reasonable to assume that periodontal treatment and oral hygiene can reduce the risk of cardiovascular disease in patients with periodontitis. More studies are needed to focus on the molecular mechanism linking periodontal pathogens and cardiovascular diseases. These studies will provide evidence that periodontal pathogens directly invade the cardiovascular system or indirectly invade host cells as well as isolate and culture bacteria from the tissues of lesions. Studies should also explore how the local inflammatory state, periodontal pathogens and their products directly influence cardiovascular disease-related biomarkers (C-reactive protein, vascular endothelial growth factor, heat shock protein, etc.) and the mechanism. This information may provide a reference for the effective prevention and treatment of periodontitis and cardiovascular disease in the future.

Porphyromonas gingivalis infection causes umbilical vein endothelial barrier dysfunction in vitro by down-regulating ZO-1, occludin and VE-cadherin expression / 南方医科大学学报

OBJECTIVE@#To explore the molecular mechanisms of Porphyromonas gingivalis infection-induced umbilical vein endothelial barrier dysfunction in vitro.@*METHODS@#Human umbilical vein endothelial cells (HUVECs) were cultured in vitro, and after the formation of the endothelial barrier, the cells were infected with P. gingivals at a multiplicity of infection (MOI). The transepithelial electrical resistance (TEER) of the cell barrier was measured, and FITC-dextran trans-endothelial permeability assay and bacterial translocation assay were performed to assess the endothelial barrier function. The expression levels of cell junction proteins including ZO-1, occludin and VE-cadherin in the cells were examined by qRT-PCR and Western blotting.@*RESULTS@#In freshly seeded HUVECs, TEER increased until reaching the maximum on Day 5 (94 Ωcm2), suggesting the formation of the endothelial barrier. P. gingivals infection caused an increase of the permeability of the endothelial barrier as early as 0.5 h after bacterial inoculation, and the barrier function further exacerbated with time, as shown by significantly lowered TEER, increased permeability of FITC-dextran (40 000/70 000), and increased translocation of SYTO9-E. coli cross the barrier. MTT assay suggested that P. gingivals infection did not significantly affect the proliferation of HUVECs (P>0.05), but in P. gingivalsinfected cells, the expressions of ZO-1, occludin and VE-cadherin increased significantly at 24 and 48 h after bacterial inoculation (P < 0.05).@*CONCLUSION@#P. gingivals may disrupt the endothelial barrier function by down-regulating the expressions of the cell junction proteins (ZO-1, occludin, VE-cadherin) and increasing the permeability of the endothelial barrier.

Effect of Porphyromonas gingivalis infection on IFNGR1 palmitoylation in esophageal cancer cells / 南方医科大学学报

OBJECTIVE@#To investigate the effect of Porphyromonas gingivalis (Pg) infection on IFNGR1 palmitoylation and biological behaviors of esophageal squamous cell carcinoma (ESCC) cells and the clinical implications.@*METHODS@#The expression levels of IFNGR1 protein in ESCC cell lines KYSE30 and KYSE70 were detected using Western blotting at 24 and 48 h after Pg infection, and 2-BP was used to detect IFNGR1 palmitoylation in the cells. KYSE70 cells with wild-type IFNGR1 (IFNGR1-WT cells) and with IFNGR1-C122A palmitoylation site mutation induced by site-specific mutagenesis (IFNGR1-C122A cells) were both infected with Pg, and the changes in palmitoylation of IFNGR1-C122A were analyzed using immunofluorescence and Click-iT assays. The changes in proliferation, migration and invasion ability of the infected cells were evaluated using plate cloning assay, scratch assay and Transwell assay, and IFNGR1 co-localization with lysosomal marker LAMP2 was dected using immunofluorescence assay. Immunohistochemistry was used to detect Pg infection and IFNGR1 protein expression in 50 ESCC tissues, and their correlation with the clinicopathological characteristics and survival outcomes of the patients was analyzed.@*RESULTS@#Pg infection down-regulated the protein expression of IFNGR1 in ESCC and promoted IFNGR1 palmitoylation at site 122. In IFNGR1-WT cells, Pg infection significantly enhanced cell proliferation, migration and invasion (P < 0.05). Similarly, Pg also significantly promoted proliferation, migration and invasion of IFNGR1-C122A cells, but to a lesser extent as compared with the wild-type cells (P < 0.05). Immunofluorescence assay showed that Pg and ZDHHC3 promoted IFNGR1 degradation within the lysosome. Immunohistochemical studies of the ESCC tissue samples showed a negative correlation between IFNGR1 and Pg expression, and a reduced IFNGR1 expression was correlated with a poorer survival outcome of the patient.@*CONCLUSION@#Pg infection enhances IFNGR1 palmitoylation to promote progression of ESCC, and elimination of Pg and inhibiting IFNGR1 palmitoylation may effectively control ESCC progression.

Research progress on the structure and function of the type IX secretion system of Porphyromonas gingivalis / 口腔疾病防治

@#The Porphyromonas gingivalis type IX secretion system (T9SS) is a recently discovered protein secretion system that is widely distributed in Bacillus cereus. The T9SS is structurally complex and powerful. More than 20 T9SS components have been verified, and more than 30 virulence factors can be secreted by Porphyromonas gingivalis alone, which contributes significant to the pathogenicity of Porphyromonas gingivalis. T9SS is a large protein complex spanning the inner cell membrane, periplasm, and outer cell membrane. Through the structural and functional connections among its components, it forms a sophisticated functional complex that includes power provision, energy transduction, inner and outer membrane translocation, outer membrane modification, and regulatory systems to recognize, translocate, shear, and modify cargo proteins and translocate bacterial intracellular cargo proteins to the cell surface. In recent years, with advancements in X-ray diffraction and in situ cryoelectron microscopy, the exploration of T9SS has evolved from the functional study of single components to the in situ structural study of multiprotein complexes. Still, the structural resolution of the protein still has shortcomings such as low resolution and an inability to capture dynamic functional structures. Future research directions should focus more on exploring how T9SS interacts and functions with cargo proteins. In this paper, we review the research progress on Porphyromonas gingivalis T9SS on X-ray diffraction and cryoelectron microscopy structure resolution in order to gain a deeper understanding of the transport mechanism of T9SS.

Inhibitory effect of methylene blue mediated photodynamic therapy combined with berberine on Porphyromonas gingivalis in vitro / 国际生物医学工程杂志

Objective:To investigate the in vitro inhibitory effect of methylene blue mediated photodynamic therapy (PDT) combined with berberine on Porphyromonas gingivalis (P.g). Methods:P.g was cultured until the middle to late log phase, and methylene blue was added to P.g suspension at different mass concentrations for 5 min, and a laser (wavelength 660 nm, power 140 mW/cm 2) was irradiated for 2 min to find the optimal concentration of methylene blue combined with the laser for in vitro inhibition of P.g. The effect of methylene blue mediated PDT on the in vitro inhibition of P.g and the effect of berberine on the growth curve of P.g were observed. The inhibitory effect of methylene blue mediated PDT and berberine on P.g was investigated by successive combined applications. The effect of methylene blue mediated PDT on P.g morphology was observed by scanning electron microscopy. The absorption peaks of each component were measured by ultraviolet spectrophotometer. Results:The best inhibition was achieved at a methylene blue mass concentration of 24.414 1 μg/ml under 660 nm laser excitation. The differences were statistically significant in both the methylene blue and PDT groups compared with the control group (all P<0.001). 0.05 mg/ml berberine had an inhibitory effect on the planktonic bacteria of P.g. After P.g was treated with methylene blue mediated PDT, the bacterial cell walls were crumpled into clusters. Compared with the control group, the number of colonies was reduced in the 0.05 mg/ml berberine group, and the difference was statistically significant ( P<0.01). The difference between the 0.05 mg/ml berberine + light group and the control group was not statistically significant ( P>0.05). When PDT was combined with berberine, there was a synergistic inhibitory effect on P.g. PDT followed by berberine shows a better inhibitory effect on bacteria, and the differences were statistically significant (all P<0.01). After the berberine treatment, the bacterial surface became smooth, and the length of the bacterial body increased compared with the control group. Conclusions:Methylene blue mediated PDT has an inhibitory effect on P.g. When combined with berberine, it has a synergistic inhibitory effect on P.g., and the inhibition effect is better when PDT is applied first and then berberine is applied in combination.

Manuka Honey as an Adjunct to Nonsurgical Periodontal Treatment: A Pilot Study

@#This pilot study evaluated the effect of manuka honey as a subgingival adjunct to scaling and root surface debridement in the treatment of periodontitis. This study used a split-mouth design with a 3-month follow-up in seven participants diagnosed with periodontitis Stage III Grade B or C. Root surface debridement was performed on one side of the mouth (control); the other side received debridement plus manuka honey application (test). Clinical parameters were recorded at baseline, 6- and 12-weeks. Gingival crevicular fluid and subgingival plaque were sampled. Microbiological outcomes were analysed using benzoylarginine pnitroanilide assay and polymerase chain reaction assay. Single application of manuka honey to periodontal pockets did not result in additional reduction of pocket depth, improvement of attachment levels or changes in p-nitroaniline enzymes when compared with root surface debridement alone. However, test sites exhibited greater reduction in bleeding than control sites, mean differences 1.3 (95%CI 1.2-1.5) and 1.7 (95%CI 1.5-1.9) at 6-weeks and 12-weeks, respectively. The proportion of mutans streptococci decreased at 6-weeks in test sites but increased at 12-weeks in control sites. Adjunctive application of manuka honey to periodontal pockets improved gingival inflammation but did not demonstrate significant clinical benefits compared with root surface debridement alone.

In vitro evaluation of Andaliman (Zanthoxylum acanthopodium DC) extract on the growth of periodontal pathogenic bacteria

Aims@#Andaliman (Zanthoxylum acanthopodium DC) is a plant spice widely used in the typical cuisine of Sumatera Utara. Several studies have shown that the extract of Andaliman has an antibacterial effect against pathogenic bacteria. This study was aimed to determine the inhibitory power of Andaliman extract on the growth of periodontal pathogenic bacteria.@*Methodology and results @#The sample consisted of Andaliman extract with concentrations of 60%, 30%, 15%, 7.5%and 3.75%. Porphyromonas gingivalis ATCC®33277TM and Fusobacterium nucleatum ATCC®25586TM were selected to observe the antibacterial effect of Andaliman extract. Inhibition was tested using the disc diffusion method. Andaliman extract significantly inhibited (p<0.05) the growth of P. gingivalis and F. nucleatum. This study showed that the inhibition of extract concentrations of 60% and 30% were categorised as strong and the concentration of 15% was categorised as moderate.@*Conclusion, significance and impact of study @#The research findings indicate that Andaliman extract has the potential to inhibit the growth of P. gingivalis and F. nucleatum bacteria. It can be concluded that Andaliman extract has the potential as an alternative antimicrobial therapy against P. gingivalis and F. nucleatum bacteria.

Effects of Porphyromonas gingivalis infection and vitamin D pathway protein expression on survival and prognosis of patients with esophageal squamous cell carcinoma / 安徽医科大学学报

Objective To analyze the effects of Porphyromonas gingivalis(Pg)infection and expression of vitamin D pathway-related proteins on the survival and prognosis of patients with esophageal squamous cell carcinoma(ES-CC).Methods Pg infection and the expression of 24 hydroxylase(CYP24A1),1α hydroxylase(CYP27B1)and vitamin D receptor(VDR)in 173 ESCC tissues were detected by immunohistochemistry.The correlation between each index and the survival time of patients was analyzed.Results The positive rates of Pg,CYP24A1,CYP27B1 and VDR in ESCC were 43.35％,37.57％,20.23％and 21.97％,respectively.The 5-year survival time of ES-CC patients in the Pg+CYP24A1+CYP27B-VDR-high-risk group was shortened(P＜0.05).Conclusion Pg infection and vitamin D pathway-associated proteins can be used as reliable indicators to predict the survival and prognosis of ESCC patients.

Porphyromonas gingivalis Inhibits Ferroptosis in Esophageal Squamous Cell Carcinoma via Upregulation of HIF-1α / 华中科技大学学报(医学版)

Objective To investigate the mechanism underlying the suppressive effect of Porphyromonas gingivalis(P.gingivalis)on ferroptosis in esophageal squamous cell carcinoma(ESCC).Methods ESCC cells infected with P.gingivalis and uninfected control cells were treated with ferroptosis inducer RSL3 followed by measurements of cell viability,malondialde-hyde(MDA),reactive oxygen species(ROS),and the expression of glutathione peroxidase 4(GPX4).Moreover,the expression of hypoxia-inducible factor-1α(HIF-1α)and its target genes were detected by qPT-PCR,Western blotting or immunohistochem-istry in ESCC tissue and cells under the condition of P.gingivalis infection.The effect of P.gingivalis infection combined with the HIF-1α inhibitors LW6 and RSL3 on ferroptosis in ESCC was detected in vitro and in vivo.Results P.gingivalis infection of the ESCC cells resulted in an increase of the cell viability(P＜0.05),decreased levels of intracellular ROS(P＜0.05)and MDA(P＜0.05)and increased the expression of GPX4 compared with RSL3 treatment alone.In ESCC tissues,the increased a-bundance of P.gingivalis was correlated with upregulation of HIF-1α.Furthermore,P.gingivalis infection induced upregula-tion of HIF-1α and its target genes.LW6 promoted ferroptosis via inhibiting the HIF-1α upregulation induced by P.gingivalis infection in vitro and in vivo.Conclusion HIF-1α renders resistance to ferroptosis in P.gingivalis infected ESCC.Combination of HIF-1α inhibitory agents and ferroptosis inducing agents might be a novel therapeutic strategy in ESCC care.

Research progress on neutrophil extracellular traps in oral infectious diseases / 口腔疾病防治

@#Eutrophils are the first innate immune cells to reach the site of inflammation. Neutrophils produce neutrophil extracellular traps (NETs) that can quickly capture and limit the spread of pathogens, facilitating the removal of pathogens and their debris. Neutrophils in the oral cavity are specifically transformed from circulating neutrophils in the blood, and the number of NETs released by oral neutrophils is much higher than that of circulating neutrophils, thus better maintaining the balance of the oral microenvironment. As a bimorphic fungus, only the mycelium phase of Candida albicans can induce NETs, which is related to the neutrophils' ability to sense the size of pathogenic microorganisms through neutrophil elastase. However, spherical Staphylococcus aureus are much smaller than Candida albicans, and they can still induce NETs. Porphyromonas gingivalis, as one of the microorganisms in the periodontitis complex, induces fewer NETs than Streptococcus oralis and Actinomycetes, which are two common oral microorganisms, and there may be a mechanism allowing them to escape neutrophilic immunity in the early stage of periodontitis. Although the two main pathways of NET production have been studied in detail, the mechanisms involved in the induction of NETs by different microorganisms, especially from oral neutrophils, are not well understood. This review describes the mechanism of the immune effects of pathogenic microorganisms on neutrophil NETs in the oral cavity, providing a reference for the search for therapeutic targets and the development of key drugs for treating oral infectious diseases.

Research progress on the pathogenicity of Porphyromonas gingivalis clinical strains / 口腔疾病防治

@#Porphyromonas gingivalis (P. gingivalis) is closely related to the occurrence and development of periodontitis. It is considered to be one of the important pathogens leading to alveolar bone resorption. At present, research on P. gingivalis mostly adopts standard laboratory strains whose genetic characteristics have been confirmed, are guaranteed and are traceable, such as ATCC 33277. The virulence phenotypes (endotoxin, firmbria, etc.) of clinically extracted isolates are quite different from those of standard strains, and the pathogenic effects and ability of the host are also widely different. In addition, P. gingivalis is considered to have a significant correlation with a variety of systemic diseases, and the virulence characteristics and pathogenic ability of different strains will have different effects on systemic diseases. However, at present, there is a lack of research on clinical strains and standard strains, and there is a lack of systematic comparison between the two sources of bacteria. In this paper, the differences in the virulence phenotypes and pathogenic effects between clinical isolates and standard strains of P. gingivalis in the last 5-10 years are reviewed. The aim is to elucidate the important virulence gene loci in the P. gingivalis gene sequence, which will play an important role in improving therapeutic methods and the development of related drugs.

Effects of different debridement methods on titanium surfaces in vitro / 口腔疾病防治

Objective @#To investigate the effect of different decontamination methods, including photodynamic therapy, sandblasting and titanium curette, on titanium surface morphology and bacterial adhesion for the treatment of peri-implant disease. @*Methods@#Porphyromonas gingivalis (Pg) and Fusobacterium nucleatum (Fn) were inoculated on the surface of polished titanium specimens, and titanium specimen surfaces were treated with different decontamination methods after incubation. The titanium specimens were divided into a no-treatment control group, photodynamic group, sandblasting group and titanium curette group according to different decontamination methods. The changes in titanium surface roughness were observed by atomic force microscopy (AFM), and the remaining bacteria on the titanium surface were observed by scanning electron microscopy (SEM) and live/dead bacteria staining tests. After reinoculation of Pg and Fn, bacterial readhesion was observed on the surface of decontaminated titanium specimens. @*Results @#The AFM results showed that the surface roughness of the titanium curette group was significantly higher than that of the no-treatment control group, photodynamic group and sandblasting group (P<0.05), and there was no statistically significant difference between the no-treatment control group, photodynamic group and sandblasting group (P>0.05). The results of contact angle measurement showed that the surface contact angle of each treatment group was smaller than that of the no-treatment control group (P<0.05). The SEM results obtained after the titanium specimen surface was decontaminated showed that the number of bacteria on the no-treatment control group surface was higher and the bacteria were relatively concentrated. The bacteria on the surface of the photodynamic group, sandblasting group and titanium curette group were scattered and distributed in small numbers, and most bacteria on the surface of the photodynamic group were ruptured. The results of the live/dead bacteria staining experiment showed that the percentage of dead bacteria on the surface of the photodynamic group was significantly higher than that of the no-treatment control group, sandblasting group and titanium curette group (P<0.05). The remaining bacteria on the surface of the sandblasting group and titanium curette groups were mainly live bacteria. The remaining bacterial adhesion on the surface was significantly reduced for the sandblasting group compared to the no-treatment control group and the photodynamic and titanium curette groups (P<0.05). SEM and live/dead bacteria staining results of bacterial readhesion on the surface of titanium specimens showed that there was an aggregation of Pg on the surface of the titanium curette group, and its surface bacterial adhesion was significantly higher than that of the no-treatment control group, photodynamic group and sandblasting group. @*Conclusion @#In mechanical decontamination, sandblasting machines are a better option than photodynamic therapy and titanium curettes; however, sandblasting does not remove all bacterial contamination. For sterilization, photodynamic therapy is more effective than sandblasting and titanium curettes. A combination of sandblasting and photodynamic therapy methods for the treatment of peri-implant disease may be considered in clinical practice.

Research progress on the relationship between periodontal pathogens and colorectal cancer / 口腔疾病防治

@#With the deepening of the research on the relationship between oral microbiota and systemic diseases, researchers have found that periodontitis is closely related to diabetes, cardiovascular disease, digestive system disease and other systemic diseases. Fusobacterium nucleatum (Fn) and Porphyromonas gingivalis (Pg) are common periodontal pathogens, which play a key role in the occurrence and development of periodontitis. At present, it is also found that Fn and Pg are closely related to the occurrence and development of colorectal cancer (CRC). They can affect the occurrence and development of CRC and the therapeutic effect and prognosis of CRC patients through a variety of ways. It can promote tumor cell proliferation by regulating cell division cycle and inhibiting cell apoptosis, inhibit immune cell function to mediate immune escape and tumor metastasis, and create a pro-inflammatory microenvironment suitable for tumor survival. The study of the effect of periodontal pathogens on the occurrence and development of colorectal cancer and its mechanism also allows us to think about new methods, such as vaccine development, immune agents and antibiotic use to better prevent and treat colorectal cancer and improve the prognosis of patients with colorectal cancer.

Liver macrophages show an immunotolerance phenotype in nonalcoholic fatty liver combined with Porphyromonas gingivalis-lipopolysaccharide infection / 华西口腔医学杂志

OBJECTIVES@#This study aimed to explore the functions and potential regulatory targets of local macrophages in nonalcoholic fatty liver combined with Porphyromonas gingivalis (P. gingivalis)infection.@*METHODS@#Single-cell RNA sequencing was used to analyze the phenotypes and functional changes in various cells in the liver tissue of nonalcoholic steatohepatitis (NASH) mice fed with P. gingivalis. Real-time polymerase chain reaction (RT-PCR), enzyme-linked immunosorbent assay, and immunofluorescence staining were applied to observe the inflammation and expression levels of macrophage antigen presenting functional markers in the NASH liver. Oil red staining was performed to observe the accumulation of local adipose tissue in the NASH liver. Results were verified through RT-PCRand RNA sequencing using P. gingivalis-lipopolysaccharide treated mouse peritoneal macrophages.@*RESULTS@#In comparison with healthy livers with Kupffer cells, the NASH liver combined with P. gingivalis infection-related macrophages showed significant heterogeneity. C1qb, C1qc, Mafb, Apoe, and Cd14 were highly expressed, but Cd209a, H2-Aa, H2-Ab1, and H2-DMb1, which are related to the antigen presentation function, were weakly expressed. Further in vivo and in vitro investigations indicated that the activation and infiltration of these macrophages may be due to local P. gingivalis-lipopolysaccharide accumulation.@*CONCLUSIONS@#P. gingivalis-lipopolysaccharide induces a local macrophage immunotolerance phenotype in nonalcoholic fatty liver, which may be the key mechanism of periodontitis pathogen infection that promotes NASH inflammation and pathogenesis. This study further clarifies the dysfunction and regulatory mechanisms of macrophages in the pathogenesis of P. gingivalis-infected NASH, thereby providing potential therapeutic targets for its clinical treatment.

Porphyromonas gingivalis enhances the proliferation of colorectal cancer Caco-2 cells via the JAK2-STAT3 pathway / 口腔疾病防治

Objective @# To investigate the effect of pathogenic bacterium-Porphyromonas gingivalis (P.g) on the proliferation and inflammatory factor expression of human colorectal cancer Caco-2 cells, to determine whether the Janus kinase 2-signal transducers and activators of transcription 3 (JAK2-STAT3) pathway is involved in the regulation of Caco-2 cell proliferation by P.g and to provide an experimental basis for further exploring the relationship between P.g and colorectal cancer. @*Methods @# Caco-2 cells were cultured in vitro, and P.g at different multiplicities of infection (MOIs) (0, 1, 10, 25) was selected to stimulate for 12, 24 and 48 h. The effect of P.g on the proliferation of Caco-2 cells was detected by CCK8. The stimulation time was set as 12, 24 and 48 h. MOI=0 was the control group, and MOI=1, 10 and 25 comprised the experimental group. qRT-PCR and Western blot were used to detect the changes in interleukin-6 (IL-6), interleukin-10(IL-10), JAK2 and STAT3 gene and protein (phosphorylated protein) levels in each group. @*Results @# After P.g infection of Caco-2 cells, P.g had a sustained stimulatory effect on the cells for 12, 24 and 48 h at MOI=1 and MOI=10 compared with the control group. Compared with that in the control group, the expression of pro-inflammatory factor IL-6 and related proliferative pathway protein JAK2 and STAT3 in Caco-2 cells with P.g infection increased in a concentration- and time-dependent manner (P<0.05). Additionally, the expression of IL-10, an anti-inflammatory factor, in Caco-2 cells infected with P.g decreased (P<0.05). After the addition of the JAK2 inhibitor AZ960, the proliferation of Caco-2 cells infected with P.g decreased, and the mRNA expression of STAT3 and JAK2 and the protein expression of p-STAT3 and p-JAK2 decreased (P<0.05). @*Conclusion @#P.g can promote the proliferation of the colorectal cancer cell line Caco-2, and the effect of P.g on Caco-2 cells may promote cell proliferation through the JAK2-STAT3 pathway while promoting the expression of the proinflammatory factor IL-6 and inhibiting the expression of the anti-inflammatory factor IL-10, creating an inflammatory environment conducive to cell proliferation, which may be the mechanism by which P.g affects the proliferation of Caco-2 cells.

TIPE2 regulates periodontal inflammation by inhibiting NF-κB p65 phosphorylation

Abstract The roles and molecular mechanisms of tumor necrosis factor-α-induced protein 8-like 2 (TIPE2) in periodontitis remain largely unknown. Objective This study aimed to determine the expression of TIPE2 and NF-κB p65 in rat Porphyromonas gingivalis-induced periodontics in vivo. Methodology Periodontal inflammation and alveolar bone resorption were analyzed using western blotting, micro-computed tomography, TRAP staining, immunohistochemistry, and immunofluorescence. THP-1 monocytes were stimulated using 1 μg/ml Pg. lipopolysaccharide (Pg.LPS) to determine the expression of TIPE2 in vitro. TIPE2 mRNA was suppressed by siRNA transfection, and the transfection efficiency was proven using western blotting and real-time PCR. The NF-κB pathway was activated by treating the cells with 1 μg/ml Pg.LPS to explore related mechanisms. Results The expression of both TIPE2 and NF-κB p65 was increased in the gingival tissues of rat periodontitis compared with normal tissues. Positive expression of TIPE2 was distributed in inflammatory infiltrating cells and osteoclasts in the marginal lacunae of the alveolar bone. However, strong positive expression of TIPE2 in THP-1 was downregulated after Pg.LPS stimulation. TIPE2 levels negatively correlated with TNF-α and IL-1β. Decreased TIPE2 in THP-1 further promoted NF-κB p65 phosphorylation. Mechanistically, TIPE2 knockdown upregulated NF-κB signaling pathway activity. Conclusions Taken together, these findings demonstrate that TIPE2 knockdown aggravates periodontal inflammatory infiltration via NF-κB pathway. Interventions aimed at increasing TIPE2 may help in the therapeutic applications for periodontitis.