中性粒细胞胞外诱捕网与牙周炎高凝状态的相关性研究

目的 探究中性粒细胞胞外诱捕网(neutrophil extracellular traps,NETs)对重度牙周炎患者凝血指标的影响.方法 纳入20例重度牙周炎患者作为试验组,20例健康人作为对照组,记录牙周指标、凝血指标.ELISA测定外周血中NETs标志物cf-DNA和MPO-DNA的含量,免疫荧光显微镜观察不同...     

The role of phagocytosis,oxidative burst and neutrophil extracellular traps in the interaction between neutrophils and the periodontal pathogen Porphyromonas gingivalis

Neutrophils are regarded as the sentinel cells of innate immunity and are found in abundance within the gingival crevice. Discovery of neutrophil extracellular traps (NETs) within the gingival pockets prompted us to probe the nature of the interactions of neutrophils with the prominent periopathogen Porphyromonas gingivalis. Some of the noted virulence factors of this Gram‐negative anaerobe are gingipains: arginine gingipains (RgpA/B) and lysine gingipain (Kgp). The aim of this study was to evaluate the role of gingipains in phagocytosis, formation of reactive oxygen species, NETs and CXCL8 modulation by using wild‐type strains and isogenic gingipain mutants. Confocal imaging showed that gingipain mutants K1A (Kgp) and E8 (RgpA/B) induced extracellular traps in neutrophils, whereas ATCC33277 and W50 were phagocytosed. The viability of both ATCC33277 and W50 dwindled as the result of phagocytosis and could be salvaged by cytochalasin D, and the bacteria released high levels of lipopolysaccharide in the culture supernatant. Porphyromonas gingivalis induced reactive oxygen species and CXCL8 with the most prominent effect being that of the wild‐type strain ATCC33277, whereas the other wild‐type strain W50 was less effective. Quantitative real‐time polymerase chain reaction revealed a significant CXCL8 expression by E8. All the tested P. gingivalis strains increased cytosolic free calcium. In conclusion, phagocytosis is the primary neutrophil response to P. gingivalis, although NETs could play an accessory role in infection control. Although gingipains do not seem to directly regulate phagocytosis, NETs or oxidative burst in neutrophils, their proteolytic properties could modulate the subsequent outcomes such as nutrition acquisition and survival by the bacteria.     

中性粒细胞细胞外陷阱网与牙周炎的相关性研究进展

中性粒细胞是机体防御系统的成员之一,在免疫反应中发挥着重要作用。中性粒细胞细胞外陷阱网(NET)是新近发现的中性粒细胞发挥作用的一种形式,除了能够诱获并杀灭病原体而发挥作用之外,还可能对机体产生直接或者间接的损伤。NET不仅出现在牙周组织中,还与牙周致病菌息息相关。本文对NET与牙周炎的相关性研究进行综述。     

Modulation of clinical expression of plaque-induced gingivitis: response in aggressive periodontitis subjects

AIM: The aim of this study was to characterize the gingival inflammatory response to de novo plaque accumulation in subjects treated for aggressive periodontitis (AP). The gingival inflammatory response of the AP subjects was retrospectively compared with that of periodontally healthy individuals (PH) matched for exposure to plaque and of periodontally healthy subjects previously identified as "high responders" (HR) and "low responders" (LR). MATERIALS AND METHODS: 13 AP subjects and 26 matched PH subjects participated in a 21-day experimental gingivitis trial. Plaque index (PlI), Gingival index (GI), gingival crevicular fluid volume (GCF) and angulated bleeding score (AngBS) were recorded at days 0, 7, 14 and 21. Cumulative plaque exposure (CPE), i.e. PlI over time, was also calculated. RESULTS: GCF was significantly higher in AP compared with PH group at each observation interval (p< or =0.001). In addition, GCF was significantly higher in AP group compared with either LR or HR groups at each observation interval (p<0.001). CONCLUSIONS: These results suggest that susceptibility to gingival inflammation in response to de novo plaque accumulation may be related to susceptibility to periodontitis.     

The gingival crevicular fluid ciprofloxacin level in subjects with gingivitis and periodontitis, and its effects on clinical parameters

OBJECTIVES: The purpose of this study, conducted on patients with gingivitis and periodontitis, was twofold: to find out the serum and gingival crevicular fluid concentration of ciprofloxacin, which is a common drug used effectively against Actinobacillus actinomycetemcomitans and to determine the effects of ciprofloxacin administration on clinical parameters. METHOD: A total of 32 adult patients, consisting of 16 subjects with gingivitis and 16 subjects with untreated chronic periodontitis, were included in the study. The subjects were divided into four groups: group I included eight subjects with chronic gingivitis who had not previously received any ciprofloxacin; group II included eight subjects with chronic gingivitis to whom three doses of ciprofloxacin were administered (Siprosan 500 mg) to establish adequate gingival crevicular fluid and serum concentrations of the agent; group III consisted of eight subjects with chronic periodontitis who had not received any ciprofloxacin; group IV included eight subjects with chronic periodontitis to whom three doses of ciprofloxacin were administered to establish adequate gingival crevicular fluid and serum concentrations of the agent. All patients were systemically healthy, free of pain and reported no current medication usage. Each patient was treated with scaling and/or root planing using specific hand instruments under local anesthesia. Gingival index, plaque index and clinical attachment levels of the teeth were used to determine the clinical condition of the subjects and findings were recorded at the beginning, seventh day, 21st day and third month of the study. Serum ciprofloxacin level was measured in venous blood. Approximately 5 ml of venous blood was drawn from subjects in groups II and IV using a standard venipuncture technique. Gingival crevicular fluid samples were sampled from six interproximal sites with six paper strips in the posterior region of upper jaw (excluding third molar) and all gingival crevicular fluid and serum samples were evaluated by high-performance liquid chromatography. RESULTS: The serum concentrations of ciprofloxacin at the first and 72nd hour were not significantly different in subjects with periodontitis compared to subjects with gingivitis. But the gingival crevicular fluid concentrations of ciprofloxacin at the same hours were significantly high in subjects with periodontitis compared to subjects with gingivitis. Both subjects with gingivitis and periodontitis had significantly higher ciprofloxacin levels in the gingival crevicular fluid than in serum. The application of ciprofloxacin did not have any positive or statistically significant effect upon the clinical parameters of the subjects with gingivitis. On the other hand, a significant decrease in the clinical attachment level scores of the subjects with periodontitis (group IV) was observed compared to group III in the 21st day and third month. CONCLUSION: According to these results, the use of ciprofloxacin as an alternative drug in subjects with periodontitis but not gingivitis can be recommended. However, long-term studies are also needed to assess the effects of ciprofloxacin on clinical parameters.     

Interleukin-1beta, interleukin-12 and interleukin-18 levels in gingival fluid and serum of patients with gingivitis and periodontitis

INTRODUCTION: Cytokines are of major importance in periodontal disease progression. Interleukin-12 (IL-12) stimulates interferon-gamma production by T helper type 1 (Th1) cells while IL-18 induces Th1 responses when present with IL-12 but Th2 responses in the absence of IL-12. IL-1beta has been correlated with periodontal disease destruction. This study determined the local concentrations of these cytokines in sites of gingivitis and periodontitis. METHODS: Gingival crevicular fluid was collected from two sites in each of 10 gingivitis patients and from two gingivitis sites and two periodontitis sites from each of 10 periodontitis patients. Serum samples were also collected. IL-1beta, biologically active IL-12 p70, the IL-12 p40 subunit and IL-18 concentrations were determined by enzyme-linked immunoabsorbent assay. RESULTS: IL-1beta and IL-18 concentrations were higher in the gingival crevicular fluid from periodontitis patients than in that from gingivitis patients; IL-18 concentrations were higher than those of IL-1beta. Very little IL-12, either p40 or p70, was detected in the gingival crevicular fluid samples. In the serum, very low levels of cytokines were found. The level of serum IL-12 p40, however, was higher than in the fluid from periodontitis sites of periodontitis patients. CONCLUSION: The local production of IL-1beta and IL-18 in the gingival crevicular fluid increased with increasing inflammation and IL-18 was the predominant cytokine at both gingivitis and periodontitis sites. Very little IL-12 was detected with levels decreasing with increasing inflammation. These results suggest that there is an association between severity of periodontal disease and levels of IL-1, IL-12 and IL-18.     

Mycoplasma in relation to gingivitis and periodontitis

In this study, mycoplasma were enumerated in the gingival crevicular fluid (GCF) of gingivitis and periodontitis patients. GCF samples were obtained from the mesial (approximal) gingival crevice or periodontal pocket of the maxillary right first permanent molar. Samples were cultured anaerobically on a medium selective for mycoplasmas as well as on a non-selective medium to obtain a total anaerobic count. Mycoplasmas were present in the GCF of both gingivitis and periodontitis patients, but a greater % was obtained from the GCF of patients with periodontitis. The proportions of mycoplasma appeared to be independent of the disease activity as expressed by bleeding on probing (PBI), but to be related to probable pocket depth, as shown by the higher %s in periodontitis than gingivitis for a given PBI. On the basis of an increased proportion in relation to disease severity, mycoplasmas would therefore appear to be amongst those micro-organisms associated with CIPD.     

Clinical and microbiological findings in elderly subjects with gingivitis or periodontitis

Abstract. The objective of the present study was to study the supra- and subgingival microflora by culture and cDNA probe methods in 20 elderly subjects who were between 62 and 93 years of age, 10 of them had gingivitis only, and 10 had periodontitis. B. forsythus (BF). P. gingivalis (PG), P intermedia (PI), P. nigrescens (PN), A. actinomycetemcomitans (AA), T. denticola (TD), and pathogen-related oral spirochetes (PROS) were studied. Oral hygiene was similar and poor in both groups. The mean probing depth at sample sites was 6.7 mm (S.D±1.3) in the periodontitis group and 2.2 mm (S.D.±1.5) in the gingivitis group (F=17.75, p < 0.001). Mean clinical attachment levels (CAL) were 4.3 mm (S.D.±2.0) and 1.7 mm (S.D.±0.9) respectively (p < 0.001). Total viable counts >1.0×10⁵ in supra-gingival plaque samples were found in all periodontitis and in eight gingivitis subjects, 70× more black-pigmented organisms were found in supra-gingival and 185 times more in sub-gingival plaque from the periodontitis group (p < 0.01). Culture data showed P nigresecens in 10% periodontitis and 50% gingivitis subjects (p < 0.03). In supra-gingival samples by the Affirm? DP test, BF was present in 50% periodontitis and 60% gingivitis while culture data were negative for all subjects, PG was found in 30%. periodontitis and 50% gingivitis subjects with TD in 70% periodontitis and in 30% gingivitis subjects. In the ub-gingival plaque samples 80% periodontitis and 70% gingivitis subjects had >1×10⁵ anaerobes. The total count of black-pigmented organisms was significantly greater in the periodontitis elders (p < 0.001). cDNA probes by the Affirm? DP test identified subgingival presence of BF (80%) PG (80%), PI(80%), AA (0%), TD (50%) in periodontitis subjects with BF (70%), PG (40%), PI (30%) and TD (20%) in gingivitis subjects. PROS were found in (80%) samples from periodontitis and in (60%) of gingivitis elderly. Only the quantities of PI (r=0.48, p < 0.01) and TD (r=0.37, p < 0.01) were associated with the disease definition. The smoking habit in the periodontitis group was significantly higher (p < 0.01). A history of smoking may contribe significantly to periodontitis in the presence of pathogens.     

The association of gingivitis and periodontitis with ischemic stroke

OBJECTIVES: The aim of this study was to assess the associations of different periodontal parameters with cerebral ischemia. METHODS: In a case-control study, 303 consecutive patients with ischemic stroke or transient ischemic attack, and 300 representative population controls received a complete clinical and radiographic dental examination. Patients were examined on average 3 days after ischemia. The individual mean clinical attachment loss measured at four sites per tooth was used as indicator variable for periodontitis. RESULTS: Patients had higher clinical attachment loss than population (p<0.001). After adjustment for age, gender, number of teeth, vascular risk factors and diseases, childhood and adult socioeconomic conditions and lifestyle factors, a mean clinical attachment loss >6 mm had a 7.4 times (95% confidence interval 1.55-15.3) a gingival index >1.2 a 18.3 times (5.84-57.26) and a radiographic bone loss a 3.6 times (1.58-8.28) higher risk of cerebral ischemia than subjects without periodontitis or gingivitis, respectively. CONCLUSION: Periodontitis is an independent risk factor for cerebral ischemia and acute exacerbation of inflammatory processes in the periodontium might be a trigger for the event of cerebral ischemia.     

Quantitative assessment of apoptotic and proliferative gingival keratinocytes in oral and sulcular epithelium in patients with gingivitis and periodontitis

BACKGROUND: Periodontal disease is caused by a chronic infection inducing an inflammatory reaction that leads to a breakdown of tooth-supporting tissue. The maintenance of an equilibrium between the host defence and microorganisms in the sulcus is essential to preserve health. All multicellular organisms have mechanisms for killing their own cells, and use physiological cell death for defence, development, homeostasis and ageing. Apoptosis and proliferation are very important phenomena in regulating this and a disturbance is often associated with disease e.g. cancer, AIDS, Alzheimer's disease, rheumatoid arthritis. OBJECTIVE: The aim of this study was to determine whether the number of apoptotic and proliferative gingival keratinocytes differed between patients with gingivitis and those with periodontitis. MATERIAL AND METHODS: The distribution of neutrophil elastase, PCNA/cyclin, DNA fragmentation (apoptosis) and p53 was determined with immunocytochemical techniques. We used paraffin-embedded sections from gingival biopsies and did quantitative analyses. RESULTS AND CONCLUSION: These showed that 5-12% of the keratinocytes in the basal layers of the epithelium proliferated in the two groups. Fewer apoptotic cells were seen in the oral epithelium than in the sulcus in all subjects in both groups. Only in the most apical part of the sulcus, close to the junctional epithelium, did the number of apoptotic keratinocytes exceed the proliferative ones in patients with periodontitis.     

牙周炎症对牙龈组织巨噬细胞胞外诱捕网形成的影响

目的:探讨牙周炎症对牙龈组织中巨噬细胞胞外诱捕网(METs)形成的影响。方法:纳入基础治疗后,需进行牙周翻瓣手术的牙周炎患者29例,以牙周健康的牙冠延长术患者20例作为对照。术前记录菌斑指数(PLI)、牙龈指数(GI)、探诊深度(PD)、临床附着丧失(CAL)等牙周临床指标,收集龈沟液(GCF)样本,术中收集牙周炎患者牙周破坏最严重部位的牙龈组织及牙冠延长术患者的健康牙龈组织。采用激光共聚焦显微镜观察牙龈组织中METs的形成情况,采用ELISA技术检测GCF中TNF-α和IL-9表达水平,分析两组样本METs形成水平与各牙周临床指标、GCF中TNF-α、IL-9水平的相关性。结果:牙周炎组METs形成分数,PLI、GI、PD、CAL等牙周临床指标,以及TNF-α、IL-9水平均显著高于正常对照组(P<0.05)。METs形成分数与PD、CAL正相关(P<0.05)。结论:牙周炎症破坏可以促进牙龈组织中METs的形成。     

Immunohistochemical expression of RANKL,RANK and OPG in gingival tissue of patients with periodontitis

Abstract

Objectives. To evaluate the expression of the receptor activator of NF-κB (RANK), the receptor activator of NF-κB ligand (RANKL) and osteoprotegerin (OPG), in the gingival tissue of patients with periodontitis. Materials and methods. Gingival tissue was obtained from 14 systemically healthy subjects with chronic periodontitis during conventional periodontal surgery. Immunohistochemistry was used to detect the expression of RANK, RANKL and OPG in the oral and periodontal pocket epithelium as well as in the connective tissue cells. Results. RANKL was negatively expressed in both oral and periodontal pocket epithelium. OPG was also negative or weakly positive in the whole epithelium. RANK showed moderate/strong positive staining mainly in the basal and suprabasal layer of oral and periodontal pocket epithelium. In most of the cases, more than 60% of the inflammatory cell infiltrate stained for RANK and RANKL. In these cases the intensity of the stained cells ranged from moderate-to-strong. In less than half of the cases, OPG was positive in more than 60% of the stained cells of the inflammatory cell infiltrate. Conclusion. The RANK, RANKL and OPG proteins are differentially expressed in periodontal tissues and may play a major role in the bone loss occurring in periodontitis.