老年牙周炎牙龈组织Fas/Apo-1(CD95)抗原表达的免疫组织化学研究

目的 :观察慢性老年牙周炎牙龈组织中Fas/Apo - 1(CD95 )抗原表达和分布情况。方法 :采用免疫组织化学染色方法对 10例慢性老年牙周炎患者、10例慢性成人牙周炎患者、10例青少年牙周炎患者和 10例健康老年人牙龈组织中Fas/Apo - 1(CD95 )抗原阳性表达和分布情况进行了观察和比较。结果 :在慢性老年牙周炎组完整的牙龈鳞状上皮间桥、核周胞浆Fas/Apo - 1(CD95 )抗原阳性表达和结缔组织中淋巴细胞Fas/Apo - 1(CD95 )抗原阳性表达为 4组中最强 ;健康老年人组上皮角化层Fas/Apo - 1(CD95 )抗原阳性表达为 4组中最强 ;慢性成人牙周炎组和青少年牙周炎组结缔组织中淋巴细胞和吞噬细胞Fas/Apo - 1(CD95 )抗原阳性表达强于健康老年人组 ;慢性老年牙周炎组上皮细胞和上皮细胞的细胞间桥Fas/Apo - 1(CD95 )抗原阳性表达例数明显高于慢性成人牙周炎组和青少年牙周炎组 (P <0 .0 5 ) ;健康老年人组结缔组织中淋巴细胞和吞噬细胞Fas/Apo - 1(CD95 )抗原阳性表达例数明显低于其它 3组 (P <0 .0 5 )。结论 :由于感染性炎症和衰老的影响 ,①牙周炎患者和健康老年人牙龈组织中鳞状上皮凋亡细胞发生部位与身体其他器官鳞状上皮细胞凋亡发生部位不同 ;②在慢性老年牙周炎患者牙龈组织中上皮细胞和炎性细胞对细胞凋亡易感性     

老年人牙周炎牙龈组织中凋亡细胞免疫组织化学研究

目的 :观察老年人牙周炎牙龈组织中凋亡细胞分布特点。方法 :采用特异性凋亡细胞免疫组织化学染色法 (TUNEL染色 ) ,对 15例老年人牙周炎患者 ,10例慢性成人牙周炎患者 ,10例青少年牙周炎患者和 11例健康老年人牙龈组织中阳性凋亡细胞的分布及计数进行比较研究。结果 :老年人牙周炎组阳性凋亡细胞总例数明显高于健康老年人组和慢性成人牙周炎组 (P <0 .0 5 ) ;慢性成人牙周炎组和青少年牙周炎组阳性凋亡细胞总例数也明显高于健康老年人组 (P <0 .0 5 ) ;老年人牙周炎组阳性凋亡细胞计数明显高于慢性成人牙周炎组、青少年牙周炎组和健康老年人组 (P <0 .0 5 )。结论 :感染性炎症因素和衰老因素均能促进老年人牙周炎时牙龈组织细胞凋亡 ,这两个因素的双重协同作用造成了老年人牙周炎患者局部牙龈组织对刺激做出过强的细胞凋亡反应。     

慢性牙周炎患者牙龈组织内诱导型一氧化氮合酶表达的研究

目的:研究牙周健康者和慢性牙周炎患者牙龈组织中诱导型一氧化氮合酶的表达强度,探讨一氧化氮在牙周病发病过程中的作用.方法:选择牙周健康组、慢性牙周炎活动期组,慢性牙周炎静止期组各20例,采取免疫组织化学的方法染色,光镜下观察牙龈组织内诱导型一氧化氮合酶的表达强度.结果:慢性牙周炎时牙龈组织中诱导型一氧化氮合酶主要在鳞状上皮和间质组织的细胞胞浆中阳性表达,正常组表达强度弱于慢性牙周炎静止期组和活动期组,慢性牙周炎静止期组表达强度弱于慢性牙周炎活动期组.结论:一氧化氮参与了慢性牙周炎的发生和发展过程,牙龈组织中诱导型一氧化氮合酶的表达强度与慢性牙周炎的炎症程度密切相关.     

老年牙周炎患者牙龈组织NO水平研究

目的：观察老年牙周炎患者牙龈组织中NO水平变化并与健康老年人、慢性成人牙周炎患者、青少年牙周炎患者比较。方法：采用硝酸还原酶法测量老年牙周炎患者、健康老年人、慢性成人牙周炎患者和青少年牙周炎患者牙龈组织中NO含量并比较。结果：老年牙周炎组NO水平明显低于健康老年人组、慢性成人牙周炎组和青少年牙周炎组（P＜0．05）。结论：由于衰老和牙周炎症的双重作用,老年牙周炎患者牙龈组织生成和分泌NO明显减少,导致抗感染和免疫调节功能的改变,非特异性免疫功能降低。     

年龄对NK细胞在牙周炎龈组织中分布影响的免疫组化研究

目的：观察老年牙周炎牙龈组织中自然杀伤（NK）细胞的分布。方法：采用免疫组织化学方法对15例老年牙周炎患者10例慢性成人牙周炎患者牙龈组织中NK细胞、T淋巴细胞和B淋巴细胞的分布及阳性计数进行比较研究。结果：慢性成人牙周炎组NK阳性细胞计数明显高于老年牙周炎组（P＜0.01)，慢性成人牙周炎组NK阳性细胞表达率也高于老年牙周炎组。结论：由于年龄增加的影响，老年牙周炎患者局部牙龈组织中出向免疫调节功能的紊乱，NK细胞功能降低。     

糖尿病对大鼠牙周组织及诱导型一氧化氮合酶分布的影响

目的：观察糖尿病对大鼠牙周炎牙周组织形态结构的影响及诱导型一氧化氮合酶（iNOS）在牙龈组织中的分布。方法：建立大鼠糖尿病牙周炎模型,制作组织切片,进行HE及免疫组织化学染色,观察牙周组织形态结构的破坏情况及iNOS在牙龈组织中的分布。结果：糖尿病牙周炎组的结缔组织附着丧失量及牙槽骨高度丧失量均明显高于牙周炎组、糖尿病组及正常组。差异有显著性（P〈0．05）。糖尿病牙周炎组和糖尿病组的免疫组化染色均为阳性或强阳性。结论：糖尿病加重了牙周炎牙周组织的破坏程度。     

血管内皮生长因子在正常与牙周炎牙龈组织中的表达差异研究

目的 检测血管内皮生长因子(VEGF)在正常与牙周炎的牙龈组织中的表达差异。方法 采用免疫组化PV两步法,检测VEGF在20例健康成人与20例慢性中重度牙周炎患者的牙龈组织中的表达差异。结果 健康组牙龈组织VEGF表达局限于上皮的颗粒层与部分棘层,其下方的结缔组织呈弱阳性表达,实验组牙龈组织上皮全层及结缔组织均呈强阳性表达,实验组VEGF表达高于健康组,两组之间的差异具有统计学意义(上皮区P<0.01,结缔组织区P<0.05)。结论 VEGF可能参与了牙周炎牙周袋壁龈组织的病理改变过程,在牙周炎发生发展及修复机制中可能具有重要意义。     

细胞粘附分子ICAM-1在慢性牙周炎牙龈组织中白细胞的表达

目的　研究ICAM - 1在慢性牙周炎牙龈组织中白细胞的表达 ,探讨ICAM - 1在慢性牙周炎免疫病因机理中和作用。方法　应用抗ICAM - 1单克隆抗体 ,通过碱性磷酸酶 -抗碱性磷酸酶的免疫组织化学技术 ,对I CAM - 1在正常和慢性牙周炎的牙龈组织中白细胞的表达进行分析。结果　ICAM - 1阳性白细胞主要聚集于慢性牙周炎结合上皮和袋上皮下方结缔组织中 ,慢性牙周炎牙龈组织单位面积中表达ICAM - 1阳性的白细胞的比例显著高于正常牙龈组 (P <0 .0 1)。结论　ICAM - 1可能参与和调节慢性牙周炎牙龈组织中白细胞介导的免疫粘附过程。     

细胞粘附ICAM-1慢性牙周炎牙龈组织中白细胞的表达

目的研究ICAM-1在慢性牙周炎牙龈组织中白细胞的表达,探讨ICAM-1在慢性牙周炎免疫病因机理中和作用.方法应用抗ICAM-1单克隆抗体,通过碱性磷酸酶-抗碱性磷酸酶的免疫组织化学技术,对ICAM-1在正常和慢性牙周炎的牙龈组织中白细胞的表达进行分析.结果 ICAM-1阳性白细胞主要聚集于慢性牙周炎结合上皮和袋上皮下方结缔组织中,慢性牙周炎牙龈组织单位面积中表达ICAM-1阳性的白细胞的比例显著高于正常牙龈组(P＜0.01).结论 ICAM-1可能参与和调节慢性牙周炎牙龈组织中白细胞介导的免疫粘附过程.     

基质金属蛋白酶-2在人慢性牙周炎肥大细胞上表达的研究

目的观察基质金属蛋白酶-2(matrix metalloproteinase-2,MMP-2)在慢性牙周炎牙龈组织中肥大细胞上的表达,探讨MMP-2-tryptase双阳性肥大细胞(mast cells,MCs)在慢性牙周炎发病机制中的作用。方法将45例参试者依据慢性牙周炎的病变程度分成3组:健康对照组15例;轻度牙周炎组15例;重度牙周炎组15例。牙龈标本经10%福尔马林液固定48 h;制作牙龈组织连续切片,HE染色,光学显微镜下观察组织学改变;采用免疫荧光双染色法,荧光显微镜下观察牙龈组织中MMP-2-tryptase双阳性MCs的表达情况。结果与健康对照组相比,轻度和重度慢性牙周炎组牙龈组织MMP-2-tryptase双阳性MCs密度均显著升高(P<0.01);重度慢性牙周炎组牙龈组织MMP-2-tryptase双阳性MCs密度显著高于轻度牙周炎组(P<0.05)。结论慢性牙周炎牙龈组织MMP-2-tryptase双阳性MCs密度与牙周炎病变程度的趋势相一致,MMP-2-tryptase双阳性MCs在牙周炎的进展中可能有促进作用。     

Immunohistochemical localization of Toll‐like receptors 1–10 in periodontitis

Background/aim: In periodontitis, bacteria and pathogen‐associated molecular patterns are sensed by Toll‐like receptors (TLRs), which initiate intracellular signaling cascades that may lead to host inflammation. In this study, the expression and distribution of TLRs (TLR‐1 to TLR‐10) were immunohistochemically detected in gingival epithelium and connective tissue. Methods: Immunohistochemistry was used for the localization of TLRs in gingival tissue samples from 10 patients with chronic periodontitis and 10 healthy controls; these samples were obtained during routine periodontal flap operations and during extraction operations performed for retained wisdom teeth, respectively. For the evaluation, epithelial layers were stratified to basal, spinous, and superficial layers, and the percentages of TLR‐positive cells were determined. Results: Both healthy and periodontitis gingival tissues expressed all TLRs except TLR‐10. In patients with periodontitis, epithelial cells showed increased TLR expression towards the basal layer. Healthy controls showed more variable cellular TLR expression and distribution between the layers of epithelium. In the connective tissue, consistently higher TLR expression was found within the periodontitis group compared to the healthy group. Conclusions: For the first time, the cellular expression and distribution of TLR‐1 to TLR‐10 have been studied in periodontitis, indicating that TLR‐1 to TLR‐9 are differentially expressed both in connective tissue and epithelial layers. Except for TLR‐7 and TLR‐8, all the other TLRs showed statistically significant differences between patients with periodontitis and healthy controls, suggesting their involvement in the pathogenesis of periodontitis.     

Tissue localization of Actinobacillus actinomycetemcomitans in human periodontitis. I. Light, immunofluorescence and electron microscopic studies

Invasion of periodontal tissues by different bacterial morphotypes has been reported in human periodontitis; however, limited information is available as to prevalence, localization and the bacterial species involved. The present study determined prevalence and gingival localization of Actinobacillus (Haemophilus) actinomycetemcomitans in periodontal lesions of juvenile periodontitis patients. Thirty-five gingival biopsies were obtained from 12 juvenile periodontitis patients at the time of periodontal therapy. One additional control biopsy was obtained from each of two adult periodontally healthy subjects, one adult periodontitis patient and one periodontally healthy monkey (Macaca fosibolius). The biopsies were carefully processed to avoid mechanical introduction of bacteria into the tissues and were examined using light and electron microscopy. Rabbit antisera specific for the three A. actinomycetemcomitans serotypes were used for immunofluorescence microscopic localization of A. actinomycetemcomitans antigens in the gingival sections. Immunofluorescence microscopy showed A. actinomycetemcomitans specific antigens in the gingival tissues of 11 of the 12 juvenile patients examined. None of the control specimens showed evidence of A. actinomycetemcomitans antigens in the gingival connective tissue. One specimen from a periodontally healthy subject and the monkey biopsy, however, showed A. actinomycetemcomitans antigens in bacterial plaque on the surface of the crevicular epithelium. Transmission electron microscopic examination showed microcolonies of small gram-negative rods in the connective tissue, as well as single bacterial cells between collagen fibers and in areas of cell debris. In addition to these extracellular bacterial cells, evidence of bacterial cells was also found within gingival connective tissue phagocytic cells. The data from the present study suggest that the gingival tissue in juvenile periodontitis lesions harbors A. actinomycetemcomitans.     

慢性牙周炎龈组织中Smac和Bax的表达及其对细胞凋亡的作用

目的:通过检测Smac和Bax蛋白在慢性牙周炎龈组织中的表达,旨在探讨二者的相关性及其在细胞凋亡中与牙周炎发生、发展的关系。方法:慢性牙周炎测试组27人,健康对照组27人,利用光镜和透射电镜观察凋亡细胞形态结构,流式细胞仪测定线粒体膜电位。免疫组化检测Smac、Bax在龈组织不同区域的表达。结果:光镜观察慢性牙周炎龈组织中存在有凋亡细胞;电镜观察凋亡细胞中线粒体形态结构改变;慢性牙周炎龈组织线粒体膜电位降低,与健康组比较,两组间有显著性差异(P<0.05);慢性牙周炎组中Smac和Bax在上皮组织、结缔组织的表达增加,与健康组相比具有显著性差异(P<0.05);Smac、Bax二者的表达在慢性牙周炎龈组织具有明显正相关(P<0.05)。结论:慢性牙周炎龈组织中有凋亡细胞存在;Smac、Bax的表达在慢性牙周炎组中呈明显正相关,表明在细胞凋亡过程中起协同作用,这与牙周组织破坏有关。     

Immunohistochemical localization of CD1a and S100 in gingival tissues of healthy and chronic periodontitis subjects

Oral Diseases (2012) 18, 778-785 Objective: The aim of this study was to evaluate the presence and distribution of CD1a and S100 protein markers in states of gingival health and chronic periodontitis in human subjects. Materials and Methods: Gingival tissue samples were derived from 10 healthy and 10 chronic periodontitis-affected human subjects. The presence and distribution of CD1a and S100 protein was assessed using immunohistochemistry, and the cell types involved in their expression was determined. Results: The presence and distribution of CD1a was confined only to the gingival epithelium, whereas S100 was seen in the epithelium and connective tissue. However, increased expression of both CD1a and S100 protein was seen in periodontitis-affected gingival tissues compared with healthy gingiva. Immunohistochemistry demonstrated that CD1a- and S100-positive cells in the epithelium are Langerhans cells (LCs) and S100 positive cells in the connective tissue are dendritic cells (DCs). Conclusion: Our findings suggest the transition of CD1a-positive LCs to S100-positive DCs from epithelium to connective tissue in response to an antigenic challenge. Demonstration of increased number of S100-positive DCs in the gingival connective tissue in chronic periodontitis possibly suggests their involvement in bone resorption in addition to their antigen presentation property.     

Immunohistochemical localization of Toll-like receptors 1-10 in periodontitis

Background/aim:  In periodontitis, bacteria and pathogen-associated molecular patterns are sensed by Toll-like receptors (TLRs), which initiate intracellular signaling cascades that may lead to host inflammation. In this study, the expression and distribution of TLRs (TLR-1 to TLR-10) were immunohistochemically detected in gingival epithelium and connective tissue.
Methods:  Immunohistochemistry was used for the localization of TLRs in gingival tissue samples from 10 patients with chronic periodontitis and 10 healthy controls; these samples were obtained during routine periodontal flap operations and during extraction operations performed for retained wisdom teeth, respectively. For the evaluation, epithelial layers were stratified to basal, spinous, and superficial layers, and the percentages of TLR-positive cells were determined.
Results:  Both healthy and periodontitis gingival tissues expressed all TLRs except TLR-10. In patients with periodontitis, epithelial cells showed increased TLR expression towards the basal layer. Healthy controls showed more variable cellular TLR expression and distribution between the layers of epithelium. In the connective tissue, consistently higher TLR expression was found within the periodontitis group compared to the healthy group.
Conclusions:  For the first time, the cellular expression and distribution of TLR-1 to TLR-10 have been studied in periodontitis, indicating that TLR-1 to TLR-9 are differentially expressed both in connective tissue and epithelial layers. Except for TLR-7 and TLR-8, all the other TLRs showed statistically significant differences between patients with periodontitis and healthy controls, suggesting their involvement in the pathogenesis of periodontitis.     

Preparation and characterization of human gingival cells

A procedure was developed for the preparation of single cell suspensions from gingival and periodontal tissues using collagenase digestion followed by gentle mechanical disruption. The procedure was evaluated on rat gingival tissues. One-hour incubation was found to produce the greatest number of cells with the highest viability, and the largest recovery of mononuclear cells. Collagenase did not appear to effect recovery, viability or B cell surface markers on human peripheral blood cells. Human gingival and periodontal tissues were obtained from 35 patients: 20 with adult periodontitis; 8 with juvenile periodontitis; 7 with normal gingivae or chronic gingivitis. Scaling, followed by probing measurements and then surgery of the affected sites was routinely performed. In single cell suspensions from tissue obtained from surgery, significantly fewer mononuclear cells were recovered from the tissues of normal/chronic gingivitis patients than cells/mg of tissue recovered from adult periodontitis or juvenile periodontitis groups. The maximum contribution of blood mononuclear cells to the gingival cells averaged 0.5+0.2%. This method is useful for recovering gingival (periodontal) cells for phenotypic and functional studies.     

Immunohistochemical study of cathepsin G and medullasin in inflamed gingival tissues from periodontal patients

Cathepsin G and medullasin are 2 major serine proteinases associated with the granular fraction of polymorphonuclear leukocytes (PMNs). To know their possible involvement in the pathophysiological gingival connective tissue turnover, we have determined the distribution and localization of these 2 enzymes in inflamed gingival tissues from periodontal patients by immunohistochemistry with discriminating antibodies specific for each enzyme. The gingival connective tissues were obtained from periodontitis patients with various inflammatory conditions and control healthy subjects without any clinical signs of periodontal inflammation. In all gingival specimens examined, cathepsin G and medullasin were found mainly in neutrophil-like cells and partly in macrophage-like cells. No positive staining for both enzymes was obtained in endothelial cells and fibroblasts in every part of the gingival tissues. Immunoreactivity for each enzyme in the gingival tissues from the periodontitis group was stronger and greater in the intensity and frequency than that from the control group and appeared to be increased with the severity of the disease. In both groups, the number of immunoreactive cells for each enzyme was greater in the vicinity of pocket epithelium (zone I) than in the area of central connective tissue (zone II) or the area subjacent to the oral epithelium (zone III). While both enzymes in zones II and III were exclusively found in coarse granules, their stainings in zone I were not only coarse but also diffuse. These results strongly suggest that both enzymes may have some association with inflamed gingival tissue degradation.