Decellularized matrix could affect the proliferation and differentiation of periodontal ligament stem cells in vitro.

OBJECTIVE AND BACKGROUND: Recently, decellularized matrix (DCM) is considered as a new biomaterial for tissue regeneration. To explore the possible application of DCM in periodontal regeneration, the effect of DCM from three different cells on the proliferation and differentiation of human periodontal ligament stem cells (PDLSCs) was investigated. METHODS: DCM derived from human periodontal ligament cells (PDLCs), dental pulp cells (DPCs), and gingival fibroblasts (GFs) were fabricated using Triton X-100/NH4 OH combined with DNase I. Allogeneic PDLSCs were cultured on PDLC-DCM, DPC-DCM, and GF-DCM, respectively. The proliferative capacity of PDLSCs was evaluated by PicoGreen assay kit. The expression of alkaline phosphatase (ALP), runt-related transcription factor-2 (RUNX2), osteocalcin (OCN), collagen I (COL1), periostin (POSTN), and cementum protein 1 (CEMP1) were detected by qRT-PCR and western blotting. RESULTS: PDLC-DCM, DPC-DCM, and GF-DCM had similar and integrated networks of extracellular matrix, as well as significantly decreased DNA content. Compared with control group in which PDLSCs were directly seeded in culture plates, PDLC-DCM, DPC-DCM, and GF-DCM promoted the proliferation of re-seeded PDLSCs. Additionally, PDLSCs on DCM exhibited higher mRNA and protein expression levels of ALP, RUNX2, OCN, and COL1. The expression of POSTN in PDLC-DCM group was significantly higher than control group at both mRNA and protein levels. CONCLUSIONS: PDLC-DCM, DPC-DCM, and GF-DCM could enhance the proliferation of PDLSCs. PDLC-DCM facilitated osteogenic differentiation and periodontal ligament differentiation of PDLSCs, while DPC-DCM and GF-DCM promoted osteogenic differentiation.

Clinical evaluation of xenogeneic collagen matrix versus free gingival grafts for keratinized mucosa augmentation around dental implants: A randomized controlled clinical trial.

AIM: To evaluate the outcomes of an apically repositioned flap (ARF) plus xenogeneic collagen matrix (XCM) in augmenting keratinized mucosa width (KMW) around dental implants when compared with ARF plus free gingival grafts (FGG). MATERIALS AND METHODS: Twenty-six participants with at least one site with KMW ≤2 mm were randomized into FGG or XCM group. Clinical examinations were performed at baseline and at 2 and 6 months after surgery, including KMW, keratinized mucosa thickness, gingival index (GI), and probing depth (PD). Post-operative pain and patient satisfaction were also evaluated. RESULTS: At 6 months, FGG attained a greater increase of KMW and thicker mucosa than XCM (4.1 ± 1.6 mm vs. 1.8 ± 1.0 mm, p < .001; 1.7 ± 0.6 mm vs. 1.2 ± 0.3 mm, p < .01). Regarding GI, PD, post-operative pain, aesthetic outcomes, and patient satisfaction, no significant difference could be detected. Moreover, the operation time of XCM group was shorter (60 ± 9 min vs. 39 ± 8 min, p < .001). CONCLUSIONS: FGG could result in greater increase of KMW than XCM, though both could increase KMW, maintain peri-implant health, and attain comparable aesthetic outcomes. The use of XCM was associated with reduced operation time.

Efficacy of xenogeneic collagen matrix in the treatment of gingival recessions: A systematic review and meta-analysis.

OBJECTIVES: This review aims to evaluate the efficacy of xenogeneic collagen matrix (XCM) for the treatment of single or multiple gingival recessions in terms of clinical parameters and patient-related outcomes. MATERIALS AND METHODS: Various electronic databases (The Cochrane Central Register of Controlled Trials, MEDLINE, EMBASE, etc.) from 1966 to April 2018 and hand literatures were searched. Quality of the included studies was assessed through the Cochrane Collaboration's Risk of Bias tool. A meta-analysis was performed to calculate risk ratios and mean differences. RESULTS: Nine randomized controlled trials were included. The results revealed a higher percentage of mean root coverage (MRC) and a greater recession reduction (RecRed) for single recessions for the combination of coronally advanced flap (CAF) with XCM compared to CAF alone (n = 3; MD = 10.00%; 95%CI [3.56%; 16.43%]; p = 0.002) (n = 3; MD = 0.35 mm; 95%CI [0.10 mm; 0.60 mm]; p = 0.005). Comparing XCM with connective tissue graft (CTG), no significant differences were detected in MRC or RecRed for single and multiple recessions. CONCLUSIONS: The addition of XCM under CAF could improve MRC and RecRed at single tooth recessions. Initial data suggest that XCM shows promising results to improve the clinical efficacy of CAF for multiple recessions. In addition, XCM could be a valid alternative to CTG in terms of MRC and RecRed at both single and multiple recessions. Based on limited evidence, XCM may decrease postoperative morbidity and operation time compared to CTG.

Possible socioeconomic and oral hygiene behavioural risk factors for self-reported periodontal diseases in women of childbearing age in a Chinese population.

PURPOSE: To examine the socioeconomic and behavioural risk factors for periodontal disease in women of childbearing age and evaluate the extent of public awareness of the association between oral health and pregnancy in China. MATERIALS AND METHODS: Cross-sectional data from 832 women (including 188 pregnant women) from Yuyao, Zhejiang Province were collected using a structured questionnaire. Demographic data were used to measure the participants' socioeconomic status. The questionnaire assessed knowledge and behaviours related to personal oral hygiene and utilisation of dental care services. Data were divided into pregnant and non-pregnant groups for multivariate logistic regression analysis. RESULTS: In total, 88.3% pregnant women and 74.2% non-pregnant women reported periodontal symptoms. Abnormal body mass index (BMI ≤ 18.5, odds ratio, OR = 0.76, 95% CI 0.27-0.97, P = 0.024; BMI ≥ 23.9, OR = 1.83, 95% CI 1.12-3.35, P = 0.035) was significantly associated with self-reported periodontal disease. Minimal mental stress (OR = 0.56, 95% CI 0.43-0.94, P = 0.028), high annual household income (OR = 0.69, 95% CI 0.17-0.82, P = 0.008), advanced oral hygiene aids (OR = 0.33, 95% CI 0.18-0.49, P < 0.001) and knowledge of the link between pregnancy and periodontal disease (OR = 0.57, 95% CI 0.33-0.96, P = 0.016) were associated with decreased incidence of self reported periodontal disease. CONCLUSIONS: A low socioeconomic background was correlated with the high incidence of self-reported periodontal disease among women of childbearing age in China. Education about primary oral health and equitable distribution of dental services might be expected to improve oral health in this specific population.

Regeneration potential of decellularized periodontal ligament cell sheets combined with 15-deoxy-Δ12,14-prostaglandin J2 nanoparticles in a rat periodontal defect.

Periodontitis is a chronic inflammatory disease characterized by loss of attachment and destruction of the periodontium. Decellularized sheet, as an advanced tissue regeneration engineering biomaterial, has been researched and applied in many fields, but its effects on periodontal regeneration remain unclear. In this study, the biological properties of decellularized human periodontal ligament cell (dHPDLC) sheets were evaluated in vitro. Polycaprolactone/gelatin (PCL/GE) nanofibers were fabricated as a carrier to enhance the mechanical strength of the dHPDLC sheet. 15-deoxy-[Formula: see text]-prostaglandin J2 (15d-PGJ2) nanoparticles were added for anti-inflammation and regeneration improvement. For in vivo analysis, dHPDLC sheets combined with 15d-PGJ2 nanoparticles, with or without PCL/GE, were implanted into rat periodontal defects. The periodontal regeneration effects were identified by microcomputed tomography (micro-CT) and histological staining, and immunohistochemistry. The results revealed that DNA content was reduced by 96.6%. The hepatocyte growth factor, vascular endothelial growth factor, and basic fibroblast growth factor were preserved but reduced. The expressions or distribution of collagen I and fibronectin were similar in dHPDLC and nondecellularized cell sheets. The dHPDLC sheets maintained the intact structure of the extracellular matrix. It could be recellularized by allogeneic human periodontal stem ligament cells and retain osteoinductive potential. Newly formed bone, cementum, and PDL were observed in dHPDLC sheets combined with 15d-PGJ2 groups, with or without PCL/GE nanofibers, for four weeks post-operation in vivo. Bringing together all these points, this new construct of dHPDLC sheets can be a potential candidate for periodontal regeneration in an inflammatory environment of the oral cavity.

Oral Mesenchymal Stem/Progenitor Cells: The Immunomodulatory Masters.

Oral mesenchymal stem/progenitor cells (MSCs) are renowned in the field of tissue engineering/regeneration for their multilineage differentiation potential and easy acquisition. These cells encompass the periodontal ligament stem/progenitor cells (PDLSCs), the dental pulp stem/progenitor cells (DPSCs), the stem/progenitor cells from human exfoliated deciduous teeth (SHED), the gingival mesenchymal stem/progenitor cells (GMSCs), the stem/progenitor cells from the apical papilla (SCAP), the dental follicle stem/progenitor cells (DFSCs), the bone marrow mesenchymal stem/progenitor cells (BM-MSCs) from the alveolar bone proper, and the human periapical cyst-mesenchymal stem cells (hPCy-MSCs). Apart from their remarkable regenerative potential, oral MSCs possess the capacity to interact with an inflammatory microenvironment. Although inflammation might affect the properties of oral MSCs, they could inversely exert a multitude of immunological actions to the local inflammatory microenvironment. The present review discusses the current understanding about the immunomodulatory role of oral MSCs both in periodontitis and systemic diseases, their "double-edged sword" uniqueness in inflammatory regulation, their affection of the immune system, and the underlying mechanisms, involving oral MSC-derived extracellular vesicles.

Association between infection of different strains of Porphyromonas gingivalis and Actinobacillus actinomycetemcomitans in subgingival plaque and clinical parameters in chronic periodontitis.

OBJECTIVE: The aim of this study was to investigate subgingival infection frequencies of Porphyromonas gingivalis and Actinobacillus actinomycetemcomitans strains with genetic variation in Chinese chronic periodontitis (CP) patients and to evaluate its correlation with clinical parameters. METHODS: Two multiplex polymerase chain reaction (PCR) assays were developed to detect the 16SrDNA, collagenase (prtC) and fimbria (fimA) genes of P. gingivalis and the 16SrDNA, leukotoxin (lktA) and fimbria-associated protein (fap) genes of A. actinomycetemcomitans in 60 sulcus samples from 30 periodontal healthy subjects and in 122 subgingival plaque samples from 61 patients with CP. The PCR products were further T-A cloned and sent for nucleotide sequence analysis. RESULTS: The 16SrDNA, prtC and fimA genes of P. gingivalis were detected in 92.6%, 85.2% and 80.3% of the subgingival plaque samples respectively, while the 16SrDNA, lktA and fap genes of A. actinomycetemcomitans were in 84.4%, 75.4% and 50.0% respectively. Nucleotide sequence analysis showed 98.62%~100% homology of the PCR products in these genes with the reported sequences. P. gingivalis strains with prtC+/fimA+ and A. actinomycetemcomitans with lktA+ were predominant in deep pockets (>6 mm) or in sites with attachment loss > or =5 mm than in shallow pockets (3~4 mm) or in sites with attachment loss < or =2 mm (P<0.05). P. gingivalis strains with prtC+/fimA+ also showed higher frequency in gingival index (GI)=3 than in GI=1 group (P<0.05). CONCLUSION: Infection of P. gingivalis with prtC+/fimA+ and A. actinomycetemcomitans with lktA+ correlates with periodontal destruction of CP in Chinese. Nonetheless P. gingivalis fimA, prtC genes and A. actinomycetemcomitans lktA gene are closely associated with periodontal destruction, while A. actinomycetemcomitans fap gene is not.

AKT2 is involved in the IL­17A­mediated promotion of differentiation and calcification of murine preosteoblastic MC3T3­E1 cells.

Interleukin (IL)­17A exhibits pleiotropic biological activities and serves a role in the progression of periodontitis. However, data describing the association between IL­17 and osteogenesis are not conclusive. It was previously demonstrated that RAC­ß serine/threonine protein kinase (AKT2)­specific knockdown in MC3T3­E1 cells weakened osteogenic effects. The role of AKT2 in the regulation of IL­17A for osteoblast differentiation and calcification remains unclear. The MTT method was adopted in the present study to assess cell proliferation; cell cycle distribution was analyzed by flow cytometry. Following osteogenic induction treatment, the involvement of phosphatidylinositol 3­kinase (PI3K) and phosphorylated­PI3K was evaluated by western blotting. The effects of IL­17A on osteogenesis­associated markers, including Runt­related transcription factor 2 (Runx­2), alkaline phosphatase (ALP) and osteocalcin (OCN) were evaluated by reverse transcription­quantitative polymerase chain reaction (RT­qPCR) analysis. An ALP activity assay and Alizarin Red S staining were used to assess the differentiation and calcification functions. AKT2 knockdown inhibited MC3T3­E1 cell proliferation, inducing significantly increased G0/G1 cell counts, and reduced S and G2/M cell numbers. IL­17A exerted no significant effects. The protein levels of p­PI3K, gene expression levels of IL­17A, Runx­2, ALP and OCN, and relative ALP activity and calcification areas were increased in the induction group, and these effects were markedly promoted by treatment with IL­17A. AKT2 knockdown in MC3T3­E1 cells resulted in reduced IL­17A­induced differentiation and calcification, although it was not completely inhibited. The results of the present study suggested that AKT2 signaling was required for MC3T3­E1 cell proliferation. IL­17A promoted osteoblast differentiation and calcification in a partly AKT2­dependent manner in MC3T3­E1 cells in vitro, possibly reflecting compensation by other signaling pathways. The results of the present study may offer novel perspectives to guide the clinical strategy for the prevention and treatment of periodontitis.

Infection frequency of Epstein-Barr virus in subgingival samples from patients with different periodontal status and its correlation with clinical parameters.

OBJECTIVE: To detect the infection frequencies of different genotypes of Epstein-Barr virus (EBV) in subgingival samples from chronic periodontitis (CP) patients, and to discuss the correlation between infection with EBV and clinical parameters. METHODS: Nested-PCR assay was used to detect EBV-1 and EBV-2 in subgingival samples from 65 CP patients, 65 gingivitis patients and 24 periodontally healthy individuals. The amplicons were further identified by restriction fragment length polymorphism analysis (RFLP) with endonucleases Afa I and Stu I. Clinical parameters mainly included bleeding on probing (BOP), probing depth (PD), attachment loss (AL) in six sites of the dentition. RESULTS: In CP patients, gingivitis and periodontally healthy individuals, the infection frequencies were 47.7%, 24.6% and 16.7% for EBV-1, and 15.4%, 7.7% and 0% for EBV-2, respectively. In 2 out of the 65 CP patients co-infection of EBV-1 and EBV-2 was found. The positive rate of EBV-1 in chronic periodontitis patients was higher than that in gingivitis patients (P=0.01) and periodontally healthy individuals (P=0.01). But no significant difference was shown in EBV-1 frequency between gingivitis patients and healthy individuals (P>0.05) or in EBV-2 frequency among the three groups (P>0.05). In CP patients, higher mean BOP value was found in EBV-1 or EBV-2 positive patients than that in EBV negative ones (P<0.01), but with no statistical difference in the mean PD or AL value between EBV positive and negative patients (P>0.05). After initial periodontal treatment, 12 out of the 21 EBV-1 positive CP patients did not show detectable EBV-1 in subgingival samples. CONCLUSION: nPCR plus RFLP analysis is a sensitive, specific and stable method to detect EBV-1 and EBV-2 in subgingival samples. Subgingival infection with EBV-1 is closely associated with chronic periodontitis. Infection of EBV in subgingival samples was correlated with BOP.

Association between coinfection of Porphyromonas gingivalis, Actinobacillus actinomycetemcomitans and Treponema denticola and periodontal tissue destruction in chronic periodontitis.

BACKGROUND: The association between the infection of Porphyromonas gingivalis, Actinobacillus actinomycetemcomitans and Treponema denticola in chronic periodontitis (CP) and the severity of periodontal disease remains to be elucidated. The aim of this study was to investigate the subgingival infection frequencies of three periodontopathic bacteria in Chinese CP patients and to evaluate the correlations between infection by these bacteria and periodontal destruction. METHODS: A multiple PCR assay using primers derived from 16SrDNA genes of P. gingivalis, A. actinomycetemcomitans and T. denticola was established to measure simultaneously the presence of the three microbes in 162 subgingival samples from 81 Chinese CP patients. RESULTS: The positive rates of P. gingivalis, A. actinomycetemcomitans and T. denticola in the subgingival samples were 84.6%, 83.3% and 88.3%, respectively. Of the subgingival samples, 68% revealed the coinfection of all the three microbes. The infection rates with P. gingivalis, A. actinomycetemcomitans or T. denticola alone was 5.9% (1/17), 17.6% (3/17) and 76.5% (13/17), respectively. A close association was present between the A. actinomycetemcomitans infection and gingival index (GI) (P < 0.01), but not between P. gingivalis or T. denticola infection and GI (P > 0.05). P. gingivalis and A. actinomycetemcomitans were more frequently detectable in middle and deep pockets than in shallow ones (P < 0.01), while T. denticola was found remarkably often in deep pockets (P < 0.05). The coinfection rate of the three microbes was significantly higher in sites with severe periodontitis than in those with mild periodontitis (P < 0.01). CONCLUSIONS: The multiple PCR established in this study can be used as a sensitive and specific method to simultaneously detect all three microbes in subgingival samples. A. actinomycetemcomitans infection may be associated with CP and play an important role in the periodontal tissue destruction. The coinfection of P. gingivalis, A. actinomycetemcomitans and T. denticola can cause more serious periodontal destruction than infection of any one or two of the three microbes.

[Infection frequency of Epstein-Barr virus in subgingival samples from patients with different periodontal status and its correlation with severity of periodontal lesion].

OBJECTIVE: To detect the infection frequencies of different Epstein-Barr virus (EBV) genotypes in subgingival samples of chronic periodontitis, and the correlation among infection with different genotypes and the severity of periodontal lesion. METHODS: Nested PCR (nPCR) with EBV-1 or EBV-2 specific primers was used to detect EBV-1 and EBV-2 in the subgingival samples from 65 chronic periodontitis patients, 65 gingivitis patients and 24 periodontal healthy individuals. The amplicons were further identified by RFLP with endonucleases Afa I and Stu I. By using periodontal attachment loss (AL) and gingival index (GI) as the observing in, the correlation of infection with different EBV genotypes and the severity of periodontal lesion were analyzed. RESULTS: 0.01 ng of EBV-1 DNA could be detectable by the established nPCR. All the samples showed the same detection results by two separated nPCR. All the EBV-1 amplification products (497 bp) by using endonuclease Afa I digestion could be divided into two fragments with 355 bp and 142 bp respectively. After endonuclease Stu I digestion, all the EBV-2 amplification products (165 bp) displayed two fragments with 118 bp and 47 bp respectively. In the samples of chronic periodontitis patients, gingivitis patients, and healthy periodontal tissues, the positive rates were 28.5% (74/260), 16.9% (44/260), and 14.6% (14/96) for EBV-1; and were 8.1% (21/260), 3.1% (8/260), and 0% for EBV-2 respectively, and the total EBV positive rates were 36.5% (95/260), 20.0% (52/260) and 14.6% (14/96) respectively. None of the positive samples was detectable for both the EBV-1 and EBV-2. The positive rates of EBV-1, EBV-2 and the total EBV positive rates in the chronic periodontitis samples were all higher than those in the gingivitis samples (all P < 0.05) and healthy periodontal tissue samples (all P < 0.01), without a significant difference between the gingivitis samples and healthy periodontal tissue samples (P > 0.05). Infection of EBV or EBV-1 or EBV-2 in CP patients could not be associated with AL or GI. CONCLUSION: Subgingival infection with either EBV-1 or EBV-2 is closely associated with chronic periodontitis. Infection of EBV may not correlate directly with severity of periodontitis.

Periodontal status and associated risk factors among childbearing age women in Cixi City of China.

OBJECTIVE: To investigate the periodontal status and associated risk factors among women of childbearing age to increase the awareness of oral health. METHODS: The study was conducted on childbearing age women in Cixi, a city in Zhejiang Province in the southeast of China. A total of 754 women participated in periodontal examination while receiving prenatal care. Data of the women were collected from the Cixi Family Planning Commission and during an interview. Clinical periodontal indices, such as bleeding on probing (BOP), probing depth (PD), and clinical attachment level (CAL) were measured during the examination. Statistical analysis on subject-based data was performed. RESULTS: The prevalence of periodontal disease among childbearing age women in Cixi was high (84.7%). A significant association was found between the disease and educational level, pregnancy, taking oral contraceptives, stress, alcohol consumption, overweight, dental visit, and teeth brushing (P<0.05). Women who suffered periodontal disease showed deep PD, obvious BOP, and clinical attachment loss. Among this population, pregnancy was closely associated with higher BOP percentage; teeth brushing no more than once per day or brushing for less than 1 min (P<0.001) after adjusting for age and stress. CONCLUSIONS: The periodontal status of childbearing age women in Cixi needs to be improved urgently. Attention towards the periodontal health should be warranted, especially for those in special statuses and with poor awareness.

Osteogenic effect of Drynariae rhizoma extracts and Naringin on MC3T3-E1 cells and an induced rat alveolar bone resorption model.

OBJECTIVE: To investigate if Drynariae rhizoma (DR) and its main ingredient Naringin could reduce alveolar bone loss by stimulating the proliferation and differentiation of osteoblasts. MATERIALS AND METHODS: The effect of DR water (DRWE), ethanolic extract (DREE), and Naringin on MC3T3-E1 cells was evaluated respectively by MTT method and by measuring the activity of alkaline phosphatase (ALP activity) as well as the level of osteocalcin in medium. Bone mineral density (BMD) detection, osteoclast counting by tartrate resistant acid phosphatase staining, and histopathological analysis were performed in an induced rat model of alveolar bone resorption after gastric perfusion with DR extracts or Naringin. RESULTS: DRWE and Naringin effectively increased the proliferation of MC3T3-E1 cells, whilst DREE and Naringin enhanced the differentiation of osteoblastic cells. The in vivo study indicated an elevated BMD value in the tooth-periodontal tissues from DRWE, DREE and Naringin treated groups after 10, 20 and 30 days of perfusion (P<0.05). In DRWE treated group, the number of osteoclasts at days 10, 20 and 30 decreased remarkably as compared to the corresponding negative controls (P<0.05), and no osteoclast could be found at day 30. New non-calcified bone-like matrix attached by osteoblasts at the root furcation was also shown. CONCLUSIONS: DR could be a supplementary medicine for periodontal therapy as it could reduce bone resorption in rat model of alveolar bone resorption and exert osteogenic effect on osteoblasts.

Inflammatory cytokines, adiponectin, insulin resistance and metabolic control after periodontal intervention in patients with type 2 diabetes and chronic periodontitis.

OBJECTIVE: To evaluate the effects of periodontal intervention on inflammatory cytokines, adiponectin, insulin resistance (IR), and metabolic control and to investigate the relationship between type 2 diabetes mellitus (T2DM) and moderately poor glycemic control and chronic periodontitis. METHODS AND PATIENTS: A total of 190 moderately poorly controlled (HbA1c between 7.5% and 9.5%) T2DM patients with periodontitis were randomly divided into two groups according to whether they underwent periodontal intervention: T2DM-NT and T2DM-T group. The levels of serum adiponectin, C-reactive protein (CRP), tumor necrosis factor α (TNF-α), interleukin-6 (IL-6), lipid profile, glucose, insulin, homeostasis model of assessment-insulin resistance (HOMA-IR) and homeostasis model assessment of ß-cell function (HOMA-ß) were measured at baseline and after 3 months. RESULTS: The levels of clinical periodontal variables, the probing depth, attachment loss, bleeding index, and plaque index were improved significantly in T2DM-T group after 3 months compared to T2DM-NT group (all p<0.01). After 3 months, the serum levels of hsCRP, TNF-α, IL-6, fasting plasma glucose (FPG), glycosylated hemoglobin (HbA1c), fasting insulin (FINS) and HOMA-IR index decreased, and adiponectin was significantly increased in T2DM-T group compared to those in the T2DM-NT group (p<0.05 or p<0.01). CONCLUSION: Periodontal intervention can improve glycemic control, lipid profile and IR, reduce serum inflammatory cytokine levels and increase serum adiponectin levels in moderately poorly controlled T2DM patients.

[Effect of lipopolysaccharide of Porphyromonas gingivalis on prostaglandin E2 biosynthetic pathway in human monocytic cell strain THP-1].

OBJECTIVE: To investigate the effect of lipopolysaccharide of Porphyromonas gingivalis (Pg-LPS) on the bio-thythetic pathway of prostaglandin E2 (PGE2) and its difference from lipopolysaccharide of Escherichia coli (Ec-LPS). METHODS: Purified Pg-LPS and Ec-LPS were used to stimulate a human monocytic cell strain THP-1. PGE2 concentration was determined by an enzyme immunoassay kit. The release of tritium labeled arachidonic acid (AA) was detected by a liquid scintillation counter. Reverse transcription polymerase chain reaction and western blot were used to analyse the expression of cytosolic phospholipase A2 (cPLA2) enzyme, cyclooxygenase-2 (COX-2), and microsomal prostaglandin E synthase-1 (mPGES-1). RESULTS: The effect of Pg-LPS on induction of PGE2 and release of AA was significantly weaker than that of Ec-LPS (P < 0.05).Increased secretion of PGE2 was observed after stimulation with Pg-LPS for 6 h, which peak at 24 h at (221.40 +/- 29.46) ng/L; or with Ec-LPS for 1-48 h, at (161.80 +/- 17.31) approximately (379.80 +/- 37.35) ng/L. The highest levels of COX-2 and mPGES-1 were shown after 16 h treatment by Pg-LPS, or after 8 h and 16 h by Ec-LPS respectively.cPLA2 inhibitor AACOCF3 could lower the level of LPS-induced release of AA, while it did not influence the production of PGE2. COX-2 inhibitor NS-398 could remarkably reduce the concentration of PGE2. CONCLUSIONS: Pg-LPS showed delayed and weaker effect on PGE2 biosynthetic pathway than Ec-LPS. Pg-LPS-induced PGE2 synthesis was mainly due to enhanced expression of COX-2 and mPGES-1, whereas cPLA2 played an insignificant role.

Effect of Porphyromonas gingivalis PrtC on cytokine expression in ECV304 endothelial cells and its level in subgingival plaques from patients with chronic periodontitis.

AIM: To investigate the effect of the collagenase gene (prtC) product of Porphyromonas gingivalis on inducing host cells to secrete inflammatory cytokines, and to discuss the correlation between the PrtC level in subgingival plaque samples and clinical parameters. METHODS: A prokaryotic expression system pET32a-prtC-Escheria coli BL21DE3 was constructed. Antigenicity and immunoreactivity of the recombinant PrtC protein (rPrtC) was identified by Western blotting. ELISA was applied to detect interleukin (IL)-1alpha, IL-8, and TNF-alpha levels in supernatants from rPrtC-induced human umbilical vein endothelial cells (HUVEC) originated ECV304 cells. Clinical parameters recorded at baseline and after treatment included bleeding on probing (BOP), probing depth (PD), and attachment loss (AL). ELISA was established to measure the PrtC level in 196 subgingival plaque samples from 49 patients with chronic periodontitis. RESULTS: After coincubation with 1 microg/mL rPrtC for 24 h and with 5 or 10 microg/mL rPrtC for 12 h, the levels of IL-1 alpha, IL-8, and TNF-alpha secreted by the ECV304 cells increased significantly (P<0.05). The PrtC level in the BOP-positive or the > or =5 mm AL or > or = 6 mm PD sites was higher than that in the BOP-negative or the < or =2 mm AL or < or =6 mm PD sites (P<0.05), respectively. Compared with baseline, the PrtC levels in different AL sites or in the < or =6 mm PD pockets decreased remarkably after treatment (P<0.01), but in the BOP-positive or in the > 6 mm PD sites, the PrtC levels changed insignificantly (P>0.05). CONCLUSION: rPrtC is able to directly induce host cells to synthesize and secrete IL-1 alpha, IL-8, and TNF-alpha. The PrtC level in subgingival samples is correlated with BOP, AL, and PD.

Correlation between infections with different genotypes of human cytomegalovirus and Epstein-Barr virus in subgingival samples and periodontal status of patients.

Accumulating evidence indicates that herpesviruses may be putative pathogens in various types of periodontal diseases. The present study was performed to examine infections with different genotypes of human cytomegalovirus (HCMV) and Epstein-Barr virus (EBV) in subgingival samples from a Chinese population and to analyze the correlation with periodontal status. A nested PCR assay was used to identify the presence of HCMV, EBV type 1 (EBV-1), and EBV-2; and the amplicons were further analyzed by restriction fragment length polymorphism analysis. HCMV was detected in 79.0% of 143 chronic periodontitis (CP) patients, 78.5% of 65 gingivitis patients, and 76.3% of 76 periodontally healthy individuals, while EBV was found in 63.6%, 32.3%, and 30.3% of the three groups of subjects, respectively. The HCMV-positive PCR products from all the samples were identified as corresponding to gB genotype I (gB-I) or gB-II. HCMV gB-II (62.9%), EBV-1 (43.4%), and EBV-2 (18.2%) were associated with CP at higher frequencies (P < 0.05), whereas HCMV gB-I was more often observed in gingivitis patients (40.0%) and healthy individuals (40.8%) (P < 0.05). Furthermore, a higher rate of coinfection with HCMV and EBV was shown in CP patients (52.4%), especially dual infections with HCMV gB-II and EBV-1 (30.8%) or HCMV gB-II and EBV-2 (12.6%), compared with the rates of single infections with HCMV or EBV (P < 0.05). Infection with HCMV gB-II, EBV-1, or EBV-2 was correlated with higher rates of bleeding on probing (P < 0.05). In patients infected with HCMV gB-II or both HCMV and EBV, including HCMV gB-II and EBV-1, a deeper probing depth or more serious attachment loss was found (P < 0.05). These findings clearly indicate that HCMV gB-II is the dominant genotype detected in subgingival samples in CP. HCMV gB-II infection and HCMV gB-II coinfection with EBV-1 are closely associated with periodontal tissue inflammation and destruction.