二肽基肽酶Ⅳ在Ⅱ型糖尿病大鼠牙龈表达的影响

目的：探讨高脂饮食和Ⅱ型糖尿病对大鼠牙龈组织中二肽基肽酶Ⅳ（DPP—Ⅳ／CD26）表达的影响。方法：将实验动物分为高脂饮食组（HF）、Ⅱ型糖尿病组（DM）和对照组（正常饮食）,根据葡萄糖耐量试验异常确定Ⅱ型糖尿病动物模型成功。采用RT—PCR半定量和Western blot测定DPP—Ⅳ／CD26在大鼠肠（回肠）、肝脏、肾脏和牙龈组织中的表达情况。结果 与对照组比较,RT—PCR半定量测定HF组大鼠小肠、肾脏及牙龈组织内DPP—Ⅳ mRNA表达明显增加（151．58％,239．73％,169．7％,P〈0．05）,肝脏的DPP—Ⅳ无明显变化;DM组大鼠小肠、肾脏、肝脏和牙龈组织DPP—Ⅳ mRNA表达均明显增加（242．36％,324．93％,190．57％,267．4％,P〈0．01）。Western blot证实HF组大鼠DPP—Ⅳ蛋白表达在小肠、肾脏、牙龈组织内中度增加（152．37％,154．17％,157．8％,P〈0．05）,而在肝脏无明显增加;DM组大鼠DPP—Ⅳ蛋白表达在小肠、。肾脏、肝脏和牙龈组织均明显增加（222．5％,210．82％,225．43％,254．7％,P〈0．01）。结论：在高脂饮食和Ⅱ型糖尿病时牙龈组织中DPP—Ⅳ表达上调,而这种上调可能是Ⅱ型糖尿病影响牙周炎发生发展的机制之一。     

口腔鳞癌中Maspin和VEGF基因的表达及意义

目的：研究乳腺丝氨酸蛋白酶抑制剂基因（marray serine proteinase inhibitor,Maspin）、血管内皮生长因子（vascular endothelial growth factor,VEGF）在口腔鳞癌中的表达及其相关性。方法：应用RT—PCR方法,分别对18例口腔鳞癌的癌组织、癌旁组织及相应正常组织标本行Maspin和VEGF mRNA含量的半定量测定,应用随机区组设计的F检验和Person直线相关分析进行统计分析。结果：Maspin mRNA在口腔鳞癌的癌组织、癌旁组织和正常组织中表达逐渐增加（P〈0．01）,且三者之间表达差异均有显著性（P〈0．05）;VEGF mRNA在癌组织、癌旁组织、正常组织中的表达依次降低（P〈0．01）,且癌组织和正常组织、癌组织和癌旁组织之间表达差异有显著性（P〈0．05）;口腔鳞癌中,Maspin和VEGF mRNA表达呈负相关（P〈0．01）;Maspin和VEGF基因mRNA表达与肿瘤淋巴结转移相关（P〈0．05）,与肿瘤的临床分期和病理分级无关。结论：Maspin基因在口腔鳞癌的发生发展过程中可能起着重要抑癌作用：Maspin可能通过下调VEGF来发挥抑癌作用。     

吸烟对慢性牙周炎患者龈沟液及牙龈组织中β防御素2、3的影响

目的 探讨吸烟对慢性牙周炎患者龈沟液（gingival crevicular fluid,GCF）及牙龈组织中β防御素(human beta def-ensin,hBD)2、3表达的影响。方法 将研究对象分为吸烟慢性牙周炎组和非吸烟慢性牙周炎组。采用酶联免疫吸附法（Elisa）检测hBD2、3的浓度;反转录多聚合酶链反应(RT-PCR)检测hBD2、3mRNA的表达,并做半定量分析。相关数据采用SPSS 17．0软件包进行统计学分析。结果 在GCF中,hBD2、3在吸烟组的水平表达均低于非吸烟组;hBD2、3在2组牙龈组织样本中均有mRNA表达,其中吸烟组较非吸烟组mRNA表达水平减弱。结论 吸烟可使GCF及牙龈组织中hBD2、3的表达发生改变,提示吸烟可对牙周宿主免疫防御系统产生消极影响。     

EMMPRIN与牙周炎的相关性研究

目的：研究EMMPRIN在牙周组织中的表达情况,分析其可能发挥的作用。方法：免疫组化、RT-PCR和western blot检测牙龈组织中EMMPRIN mRNA和蛋白表达情况。结果：临床健康和炎性牙龈组织中检测到EMMPRIN蛋白的表达,在炎性牙龈组织中的蛋白表达水平显著高于正常牙龈（P〈0.05）。Pg LPS处理后的牙龈上皮细胞（HGEC）中EMMPRIN和MMPs mRNA的表达存在个体差异：来源于临床健康个体、牙龈炎和轻度牙周炎患者的HGEC在作用前后EMMPRIN和MMPs mRNA的表达无显著差异,而来源于重度牙周炎患者的HGEC在Pg LPS刺激后mRNA的水平显著上调（P〈0.05）。结论：EMMPRIN可能参与牙周组织正常的生理改建;在炎症状态下EMMPRIN可能作为MMPs的诱导因子参与了对牙周组织的破坏,且与牙周炎的严重程度存在一定相关性。     

组胺及组胺受体1在大鼠牙周炎发病中表达差异的研究

目的：研究组胺及组胺受体1（HR1）在牙周炎发病机制中的作用。方法：将40只雄性SD大鼠随机分为实验组和对照组（每组20只）,其中实验组采用正畸丝结扎法建立牙周炎模型,对照组不作任何处理;建模8周后,分别采用ELISA法检测各组龈沟液中组胺含量,荧光定量PCR法检测各组牙龈组织中HR1 mRNA的表达水平。结果：牙周炎组龈沟液中的组胺浓度（35．47±3．908）μg／L 高于正常对照组（19．77± 3．832）μg／L（P＜0．05）;牙周炎组牙龈组织中的HR1－mRNA表达水平低于正常对照组（P＜0．05）。结论：组胺及HR1对牙周炎的发病均有一定调控作用。     

大鼠慢性牙周炎与IgA肾病复合模型血清中MMP-3的表达及意义

目的：研究大鼠慢性牙周炎与IgA肾病（IgAN）复合模型血清中基质金属蛋白酶-3（MMP-3）的水平,探讨慢性牙周炎与IgAN之间的可能发病机制。方法：40只SD大鼠随机分成4组,每组10只,分别为空白对照组、慢性牙周炎组、IgAN组和慢性牙周炎＋IgAN复合组。第12周末处死动物分别检测血清中MMP-3、血肌酐（Scr）,24h尿蛋白,大鼠牙龈附着丧失（AL）及出血指数（SBI）结果,观察牙周和。肾脏组织病理学改变。结果：慢性牙周炎＋IgAN复合组分别与空白对照组、慢性牙周炎组、IgAN组相比,血清MMP-3、Scr含量均显著增高（P〈0．05）,血清MMP-3与24h尿蛋白定量、AL的多元线性回归方程：24h尿蛋白-1．822＋0．206×MMP-3,R2—0．691（P〈0．05）AL-26．316＋1．577×MMP-3,R2—0．549（P〈0．05）。结论：血清MMP-3的水平在慢性牙周炎与IgAN复合组中明显增加,可能与两种疾病存在一定的关系,为研究二者发病机制提供了一个方向。     

硝酸盐对牙龈局部炎症程度影响的实验研究

目的检测分析牙周组织局部滴注硝酸盐后,一氧化氮（NO）含量变化与局部炎症发生和发展的关系,初步探讨硝酸盐在牙周炎症预防与治疗中的作用。方法建立大鼠实验性牙周炎动物模型,将70只健康SD大鼠随机分为正常对照组（N组）,牙龈炎组（P组）和药物对照组（PP组）,其中牙龈炎组又分为1000uM／L硝酸钾（KNO3）滴注组和非KNO3滴注组,所有动物分别于术后1、4周处死,采用酶法检测实验动物牙龈组织中NO2^-和NO3^-的含量,并用肉眼和组织切片观察牙龈组织中炎症的变化情况。结果正常对照组与牙龈炎组中未滴注1000uM／LKNO3溶液的牙龈组织中NO含量有高度显著性差异（P〈0．01）,在牙龈炎组中,牙龈局部给予KNO3溶液滴注与未给予KNO3溶液滴注的牙龈组织中NO含量无显著性差异（P〉0．01）,药物对照组与正常对照组牙龈组织中NO含量有高度显著性差异（P〈0．01）,药物对照组（PP组）与P组未滴注KNO3溶液的牙龈组织中NO含量有显著性差异（P〈0．01）。结论牙周局部给予硝酸盐滴注后能够增加NO的产生,提高局部NO的浓度,从而改善微循环,增强牙周局部抵抗能力,减轻和预防牙周炎症的发生。     

实验性牙周炎大鼠体内外周血及牙周组织中Th17/Treg的检测

目的初步探讨实验性牙周炎大鼠外周血及炎症牙周组织中辅助性T细胞17（Th17）和调节性T细胞（Treg）的表达。 方法采用丝线结扎并牙龈卟啉单胞菌菌液涂抹法建立SD大鼠牙周炎模型,采集外周血、龈沟液以及颌骨样本。体视显微镜及苏木精-伊红染色法观察牙周组织病理改变,流式细胞术检测外周血Th17细胞与Treg细胞的比例,酶联免疫吸附法检测外周血及龈沟液中白细胞介素17（IL-17）、IL-10、转化生长因子β（TGF-β）表达水平,实时荧光定量聚合酶链反应（PCR）法检测牙周组织中RORγt、Foxp3 mRNA的表达。应用SPSS 22.0软件独立样本t检验及方差分析对数据进行统计学分析。 结果实验大鼠牙周组织病理改变符合牙周炎病理改变特征,牙槽骨吸收明显。实验组外周血Th17（F=30.88,P=0.00）、Treg细胞（F=147.03,P=0.00）比例以及Th17/Treg比率较对照组均显著上升（F=219.53,P=0.00）;实验组牙周组织中RORγt（F=35.61,P=0.04）和Foxp3 mRNA（F=216.60,P=0.01）表达较对照组表达显著上升;RORγt/Foxp3 mRNA比率较对照组升高,但差异无统计学意义（F=0.63,P=0.44）;实验组外周血血清中IL-10浓度较对照组显著上升（F=6.41,P=0.02）,而IL-17（F=0.08,P=0.78）、TGF-β（F=3.48,P=0.06）浓度与对照组差异无统计学意义;实验组龈沟液中IL-17（F=9.03,P=0.01）较对照组显著上升;IL-10（F=5.85,P=0.08）及TGF-β含量（F=9.28,P=0.06）含量较对照组升高,但差异无统计学意义。 结论实验性牙周炎大鼠外周血及牙周炎症组织中Th17与Treg表达均显著上调,可能与牙周炎症组织病理改变以及相关全身系统性疾病存在相关性。     

牙周炎患者牙龈组织中IL-21表达的研究

目的：研究不同类型牙周炎患者牙龈组织中IL-21基因的表达,探讨其在牙周炎发病中的作用。方法：选择慢性牙周炎患者12例,侵袭性牙周炎8例,健康对照组8例,采用实时定量PCR方法定量检测IL-21 mRNA在牙龈组织表达情况。结果：慢性牙周炎、侵袭性牙周炎、健康对照组牙龈组织中均有IL-21 mRNA的表达,慢性牙周炎组和侵袭性牙周炎组IL-21 mRNA相对表达量分别为0.000534±0.000504和0.00602±0.000137,显著高于健康对照组0.000161±0.000352,差异有统计学意义(P<0.05)。慢性牙周炎和侵袭性牙周炎牙龈组织IL-21mRNA表达没有显著性差异（P>0.05）。结论：IL-21在慢性牙周炎发病机制中可能发挥重要作用。     

慢性牙周炎患者牙龈组织中IL-6、IL-34和M-CSFR的表达及临床意义

目的: 比较慢性牙周炎患者牙龈组织和健康牙龈组织中IL-6、IL-34及M-CSFR的表达水平,探讨IL-34 在慢性牙周炎发病中的作用。方法: 收集8例诊断为轻度慢性牙周炎患牙的牙龈组织和8例诊断为重度慢性牙周炎患牙的牙龈组织作为病例组,8例冠延长手术切除的牙龈组织作为对照组,应用实时荧光定量PCR检测IL-6、IL-34及M-CSFR mRNA的表达;应用蛋白免疫印迹检测IL-6、IL-34及M-CSFR蛋白的表达。使用GraphPad Prism 6.0对结果进行统计学分析。结果: IL-6、IL-34、M-CSFR mRNA及蛋白在慢性牙周炎牙龈组织中的表达水平均显著高于正常牙龈组织中的表达（P<0.05）。结论: IL-6、IL-34及M-CSFR的表达可能与慢性牙周炎密切相关。     

Evidence of the presence of T helper type 17 cells in chronic lesions of human periodontal disease

Introduction:  Periodontal disease is a chronic inflammation of the attachment structures of the teeth, triggered by potentially hazardous microorganisms and the consequent immune-inflammatory responses. In humans, the T helper type 17 (Th17) lineage, characterized by interleukin-17 (IL-17) production, develops under transforming growth factor-β (TGF-β), IL-1β, and IL-6 signaling, while its pool is maintained by IL-23. Although this subset of cells has been implicated in various autoimmune, inflammatory, and bone-destructive conditions, the exact role of T lymphocytes in chronic periodontitis is still controversial. Therefore, in this study we investigated the presence of Th17 cells in human periodontal disease.
Methods:  Gingival and alveolar bone samples from healthy patients and patients with chronic periodontitis were collected and used for the subsequent assays. The messenger RNA expression for the cytokines IL-17, TGF-β, IL-1β, IL-6, and IL-23 in gingiva or IL-17 and receptor activator for nuclear factor-κB ligand in alveolar bone was evaluated by real-time polymerase chain reaction. The production of IL-17, TGF-β, IL-1β, IL-6, and IL-23 proteins was evaluated by immunohistochemistry and the presence of Th17 cells in the inflamed gingiva was confirmed by immunofluorescence confocal microscopy for CD4 and IL-17 colocalization.
Results:  Our data demonstrated elevated levels of IL-17, TGF-β, IL-1β, IL-6, and IL-23 messenger RNA and protein in diseased tissues as well as the presence of Th17 cells in gingiva from patients with periodontitis. Moreover, IL-17 and the bone resorption factor RANKL were abundantly expressed in the alveolar bone of diseased patients, in contrast to low detection in controls.
Conclusion:  These results provided strong evidence for the presence of Th17 cells in the sites of chronic inflammation in human periodontal disease.     

新疆木垒县哈萨克族慢性牙周炎患者血清与唾液中IL-6、IL-17的表达及相关性分析

目的:研究哈萨克族慢性牙周炎(chronic periodontitis,CP)患者血清和唾液中白细胞介素17(interleukin 17, IL17 )、白细胞介素6(interleukin 6, IL6 ) 的活性表达及相关性分析。方法:选择哈萨克族慢性牙周炎患者192例,健康人100例,记录临床牙周检查指标,采集非刺激性全唾液及外周血液各5 mL。采用抗体夹心ELISA法测定CP患者与健康人群唾液及血液中IL-17、IL-6总量和浓度。结果:牙周炎患者的唾液IL-6、IL-17浓度要显著高于血清IL-6、IL-17。在唾液中IL-6与IL-17彼此呈中度正相关性,且二者与牙周临床指标均为高度正相关关系。结论:唾液IL-6、IL-17在哈萨克族慢性牙周炎患者的表达水平明显增高,且与病情严重程度相关,可作为萨克族慢性牙周炎患者的诊断标志物,对二者的相关性可从基因学方面进行下一步研究。     

CXCL16及其受体CXCR6在中重度牙周炎患者牙龈组织中的表达

目的 观察趋化因子CXCL16及其受体CXCR6在中重度牙周炎牙龈组织和健康牙龈组织中的表达及相互关系。方法收集辽宁省人民医院口腔科门诊就诊和住院患者的牙龈组织79例,其中健康牙龈组40例,中、重度牙周炎组39例,年龄35~74岁。采用免疫组织化学染色法观察牙龈组织中CXCL16和CXCR6的阳性细胞百分率和染色强度。采用SPSS15.0软件包对数据进行统计学分析。结果与健康牙龈组相比, CXCL16在中重度牙周炎组中的表达显著升高（P<0.05）,CXCR6在中重度牙周炎组中的表达较健康牙龈组显著升高（P<0.05）。经相关性分析,CXCLl6与CXCR6在中重度牙周炎患者牙龈组织中的表达呈正相关（r=0.580,P<0.05）;而在健康牙龈组织中,两者的表达无相关性。结论趋化因子CXCLl6及其受体CXCR6可能与牙周炎的发病有关,两者在牙周炎的发病过程中可能共同发挥作用。     

Increased Expression of Interleukin (IL)‐35 and IL‐17, But Not IL‐27, in Gingival Tissues With Chronic Periodontitis

Background: Interleukin (IL)‐35 plays an important role in immune regulation through the suppression of effector T‐cell populations, including T‐helper 17 (Th17) cells. Although Th17 cells and IL‐17 are involved in the pathogenesis of periodontitis, the level of IL‐35 in inflamed periodontal tissues is unclear. Here, IL‐35, IL‐17, and IL‐27 production/expression in gingival crevicular fluid (GCF) and human gingival tissue were investigated. Methods: GCF samples were collected from buccal (mesial, center, and distal) sites of teeth from patients with chronic periodontitis (CP) and healthy controls and were analyzed by enzyme‐linked immunosorbent assay for IL‐35 (periodontitis, n = 36; healthy, n = 30) and IL‐17 (periodontitis, n = 16; healthy, n = 13). Gingival tissue, including sulcus/pocket epithelium and underlying connective tissue, was collected from an additional 10 healthy participants and 10 patients with CP and were analyzed by quantitative polymerase chain reaction (qPCR) for Epstein Barr virus‐induced gene 3 (EBI3), IL12A, and IL17A. IL27p28 was also tested by qPCR. Results: IL‐35 and IL‐17 were significantly higher in GCF from patients with periodontitis than healthy participants (P <0.01, P <0.05, respectively). In both healthy participants and those with periodontitis, positive correlations were found among IL‐35 and probing depth and clinical attachment level (CAL) as well as between IL‐17 and CAL. EBI3, IL12A (components of IL‐35), and IL17A messenger RNA expression levels were significantly higher in inflamed gingival tissue than in healthy control tissues (P <0.05). IL27p28 was not detected in any sample, suggesting that IL‐27 is not produced in large quantities in periodontal tissue. Conclusion: IL‐35 and IL‐17, but not IL‐27, may play important roles in the pathogenesis of periodontitis.     

小鼠牙周炎模型牙周组织中MicroRNA表达谱研究

目的：建立小鼠牙周炎模型牙周组织和正常小鼠牙龈组织microRNA表达差异谱,为探讨microRNA在牙周炎发病过程中是否起到可能的调控作用提供前期研究基础。方法：小鼠牙周炎牙周组织和正常小鼠牙龈组织各3例,Trizol法抽提细胞总RNA。采用基因芯片技术,将总RNA与microRNA芯片杂交,GenePixProV6．0软件进行数据分析。结果：与正常小鼠牙龈组织相比,牙周炎小鼠牙周组织中表达上调超过2倍的microRNA有4个,分别为mmu—miR-126,mmu—miR-32,mmu—miR-147,mmu—miR-155。其中牙周炎小鼠牙周组织中mmu—miR-155表达量明显提高,是正常小鼠牙龈组织的5．2倍。结论：筛选出的差异表达microRNA可能参与了牙周炎的发病过程,为探讨microRNA分子在牙周炎中的可能作用提供了途径。     

Porphyromonas gingivalis antigen preferentially stimulates T cells to express IL-17 but not receptor activator of NF-kappaB ligand in vitro

Although T cells have been implicated in the pathogenesis and are considered to be central to both their progression and control of chronic inflammatory periodontal diseases, the precise contribution of T cells to tissue destruction has not been fully clarified. Recently, interleukin (IL)-17 and receptor activator of Nuclear factor kappaB NF-kappaB ligand (RANKL) have received much attention as a result of their proinflammatory and bone metabolic roles, respectively. We therefore investigated the effect of outer membrane protein (OMP) from Porphyromonas gingivalis (P. gingivalis) on the expression of IL-17 and RANKL in peripheral blood mononuclear cells (PBMCs) and compared these between gingivitis and periodontitis, which are representative of stable and progressive lesions, respectively. The in situ expression of these molecules was also examined. P. gingivalis OMP stimulated PBMCs to express IL-17 at both the mRNA and protein level. Although the mean expression of mRNA was not different between the two groups, the mean level of IL-17 in the culture supernatants was higher in gingivitis patients than in periodontitis patients. However, the frequency of IL-17-positive samples was higher in the periodontitis patients. This stimulatory effect was not evident for RANKL expression in either periodontitis or gingivitis patients. In gingival tissue samples, IL-17 mRNA was detected in gingivitis more frequently than in periodontitis. The expression of RANKL mRNA was much lower than that of IL-17 in terms of both level and frequency. These results suggest that IL-17 but not RANKL may be involved in the pathogenesis of periodontal diseases. However, there may be negative regulatory mechanisms for IL-17 in gingivitis.     

慢性牙周炎病人基础治疗前后Th17型细胞因子及RORC的表达变化

目的:研究慢性牙周炎(CP)病人牙周基础治疗前后龈沟液(GCF)中的Th17型细胞因子及其外周血特异性转录因子RORC的表达变化规律。方法:选择CP病人30例,使用ELISA检测牙周基础治疗前后GCF中Th17型细胞因子(IL-17、IL-21)的水平变化,并利用荧光定量PCR检测牙周基础治疗前后外周血CD4+T细胞中特异性转录因子RORC的表达水平。结果:牙周基础治疗后龈沟液中的Th17型细胞因子IL-17、IL-21水平以及外周血CD4+T细胞中特异性转录因子RORC表达水平均较治疗前显著下降,差异均有统计学意义(P<0.05)。结论:Th17型细胞因子在慢性牙周炎发生发展中发挥重要的致炎作用。