脂多糖信号受体CD14、TLR4在人牙髓组织和牙髓成纤维细胞中的表达

目的研究人牙髓组织和牙髓成纤维细胞中脂多糖（LPS）信号受体CD14、TLR4的表达特点,探讨牙髓炎症组织中LPS的信号转导途径.方法采用免疫组化染色法观察健康和炎症牙髓组织中CD14、TLR4的表达情况;应用直接免疫荧光标记法,采用流式细胞术检测体外培养人牙髓成纤维细胞在LPS刺激前后的CD14、TLR4阳性细胞率和细胞表面平均荧光强度.结果正常牙髓组织中未见CD14、TLR4阳性细胞;炎症牙髓组织中可见大量CD14、TLR4阳性细胞,CD14、TLR4阳性细胞率差异无统计学意义（P＞0.05）.牙髓成纤维细胞经LPS刺激后,TLR4平均荧光表达强度和TLR4阳性细胞率均显著增高（P＜0.05）,而CD14在LPS刺激前、后均无表达.结论炎症牙髓组织中CD14、TLR4的阳性表达,提示LPS可能通过CD14、TLR4信号受体在牙髓炎症组织中发挥作用,而牙髓成纤维细胞在LPS刺激后仅表达TLR4,表明LPS可能通过TLR4对牙髓成纤维细胞发挥作用.     

CD14、TLR4在大鼠尖周炎症组织中的表达

目的:探讨CD14、TLR4在尖周组织中的分布和表达,为阐明CD14、TLR4在LPS引起的炎症反应中的作用机制提供实验依据.方法:分别将Pe、Pg菌液封入大鼠髓腔建立大鼠尖周炎症模型,并以髓腔自然开放和开髓后无菌坏死作为对照.免疫组织化学染色观察CD14、TLR4在尖周组织的分布情况.结果:4组的尖周炎症组织中均可见大量CD14、TLR4表达阳性细胞,以单核巨噬细胞为主.结论:LPS信号受体CD14和TLR4 可能参与大鼠LPS介导的尖周炎症反应.     

人正常及炎症牙髓中炎症信号受体TLR4表达的研究

目的:研究人正常及炎症牙髓组织中炎症信号受体TLR4的表达特点,探讨其在牙髓和根尖周组织炎症中的可能作用。方法:采用激光共聚焦扫描显微镜和免疫荧光技术,检测正常及炎症牙髓TLR4的表达情况。结果:正常牙髓组织中未发现TLR4免疫阳性细胞,炎症牙髓组织病灶处TLR4表达强阳性。正常组平均荧光强度为28.80±2.57,炎症组为258.12±24.51(P<0.001)。结论:与正常牙髓组织相比,炎症牙髓组织TLR4表达显著增强,提示其可能参与牙髓组织炎症过程。     

炎症信号受体TLR4在大鼠正常牙髓和牙周组织中的分布及意义

目的:研究大鼠正常牙髓和牙周组织中炎症信号受体TLR4表达特点,探讨其在健康牙髓及牙周组织天然免疫防御中的可能作用.方法:采用石蜡及冰冻切片和免疫组化技术检测大鼠健康牙髓及牙周组织中TLR4的表达情况.结果:正常牙髓组织中成牙本质细胞、血管内皮细胞及参与先天免疫的巨噬细胞、树突状细胞、中性粒细胞等细胞TLR4免疫反应阳性,牙髓成纤维细胞不表达TLR4.TLR4表达于牙周结缔组织,可见与上皮层临近的固有层结缔组织强阳性染色.结论:TLR4可能参与了牙髓牙周组织的先天免疫反应,在牙髓牙周组织抵御外来病原微生物入侵中可能起重要作用.     

CD14、TLR4在人炎症牙髓组织中的表达

目的 探讨CD14、TLR4在人牙髓组织中的表达,为阐明CD14、TLB4在LPS引起的炎症反应中的作用机制提供实验依据.方法 通过免疫组织化学染色和流式细胞术观察正常、深龋和牙髓炎牙髓组织中CD14、TLB4表达情况.结果 免疫组化染色观察正常组牙髓组织中未见CD14、TLR4阳性表达细胞;深龋组中可见CD14、TLB4表达阳性细胞,多为中性粒细胞阳性染色;慢性牙髓炎组可见大量CD14、TLR4表达阳性细胞,多为单核细胞、淋巴细胞阳性染色.流式细胞术检测到CD14、TLB4阳性细胞率随牙髓组织炎症的加重而增大;深龋组和牙髓炎组中TLB4的平均荧光强度比CD14高.结论 在牙髓炎症发展过程中,CD14、TLB4阳性细胞表达率随牙髓组织炎症的加重而增大,与炎症程度呈正相关.同时检测到在深龋组和牙髓炎组中TLR4的平均荧光强度比CD14高,可以推测LPS主要通过TLR4进行信号转导.     

根尖周炎动物模型中CD14、TLR4的表达

目的研究大鼠根尖周炎炎症组织中脂多糖炎症信号受体CD14、Toll样受体4（Toll．1ikereceptor4，TLR4）的表达特点，探讨根尖周炎中脂多糖的信号转导途径。方法建立大鼠磨牙内毒素根尖周炎模型，采用免疫组化染色观察根尖周炎炎症组织中CD14、TLR4的表达情况，并计算CD14、TLR4的阳性细胞率。结果正常根尖周组织中未发现CD14和TLR4免疫阳性细胞，根尖周炎症组织中CD14和TLR4表达阳性，CD14、TLR4阳性细胞率差异无统计学意义（P〉0．05）。结论与正常根尖周组织相比，炎症根尖周组织中CD14和TLR4的表达显著增强，CD14和TLR4的表达量差异无统计学意义，提示脂多糖可能通过CD14、TLR4信号受体在根尖周炎症中发挥作用。     

IL-34在慢性根尖周病损组织中的表达及临床意义

目的:比较慢性根尖周病损组织和健康牙周膜组织中白细胞介素34(Interleukin-34,IL-34)的表达水平,探讨IL-34在慢性根尖周炎发病中的作用.方法:收集25例诊断为慢性根尖周炎患牙的根尖周组织作为病例组,22例因正畸拔除的健康牙的牙周膜作为对照组,应用实时荧光定量PCR(real-time PCR,RT-PCR)检测IL-34mRNA的表达:应用免疫组织化学法检测IL-34蛋白的表达,采用图像分析软件检测病例组中IL-34蛋白的表达水平.采用SPSS13.0软件包对数据进行统计学分析.结果:IL-34mRNA在慢性根尖周炎中的表达水平(3.53±3.07)显著高于对照组(1.07±0.76),IL-34蛋白在慢性根尖周病损组织中阳性表达,主要定位于淋巴细胞、浆细胞及巨噬细胞中,且病例组中IL-34蛋白表达水平显著高于对照组(P＜0.01).结论:IL-34可能与慢性根尖周炎症密切相关.     

CD14在牙髓和牙龈组织中的免疫组化定位

目的：检测CD14在牙髓和牙龈组织的分布，探讨CD14在牙髓和牙龈炎症过程中的可能作用。方法：组织切片技术制备人牙髓和牙龈组织标本，用免疫组织化学方法检测CD14在健康和炎症牙髓、牙龈组织的分布。结果：健康牙髓没有发现CD14免疫阳性细胞，炎性牙髓组织病灶处炎细胞呈阳性反应；健康牙龈上皮棘层细胞有微弱的阳性反应，炎性牙龈上皮棘细胞层强阳性。结论：CD14参与了牙髓和牙龈组织的炎症反应过程；特别是牙龈上皮棘细胞层CD14免疫阳性，说明棘细胞层可能参与了牙龈炎免疫病理过程。     

TLR4在人健康和深龋牙髓组织中的定位表达研究

目的 比较人健康及深龋牙齿牙髓的组织形态及Toll样受体4(Toll like receptor 4,TLR4)在两种牙髓组织中的定位表达,以明确TLR4是否参与牙髓的天然免疫反应过程.方法 正常及深龋牙齿脱矿后行HE染色,观察牙本质及牙髓组织基本形态,正常及深龋牙髓组织行免疫组织化学染色,观察TLR4的定位表达情况,进行免疫组织化学染色评分分析.结果 HE染色显示正常牙齿牙本质小管结构完整、分布均匀,牙髓组织内细胞、血管和纤维结缔组织完整清晰.深龋牙齿部分牙本质小管遭到破坏,牙髓组织结构完整,与正常组牙髓组织相比较未见明显改变.免疫组织化学染色显示正常与深龋牙髓组织中TLR4阳性染色均表达于成牙本质细胞层及血管周围组织,但深龋牙髓组织阳性染色明显增强.免疫组织化学染色评分结果正常组为1.25±0.46,深龋组为2.10±0.74,2组差异具有统计学意义(t=2.833,P=0.012).结论 TLR4在正常及深龋牙髓中均有表达且深龋牙髓表达更多,提示TLR4参与了深龋牙髓组织的天然免疫反应.     

血小板衍化生长因子受体在人根尖周肉芽组织中表达的意义

目的 :观察血小板衍化生长因子 (platelet derivedgrowthfactor ,PDGF)受体在人根尖周肉芽组织中的表达和分布情况 ,探讨PDGF在牙周组织中的作用。方法 :制备人根尖周肉芽组织标本 15例 ,采用免疫组织化学ABC方法观察PDGFβ受体在人根尖周肉芽组织中的表达与分布情况。 结果 :在人根尖周肉芽组织中的成纤维细胞、毛细血管内皮细胞、浆细胞和巨噬细胞中能表达PDGFβ受体。 结论 :PDGF有可能在参与慢性根尖周病变的免疫病理过程并在根尖周病的修复过程中起重要的作用。     

Polymorphism of the CD14 and TLR4 Genes and Post-treatment Apical Periodontitis

Introduction

This study investigated the association of CD14 -260C>T and TLR4 +896A>G gene polymorphisms with post-treatment apical periodontitis in Brazilian individuals.

Methods

The study population consisted of 41 patients with post-treatment apical periodontitis and 42 individuals with root canal–treated teeth exhibiting healed/healing periradicular tissues (controls). All teeth had apical periodontitis lesions at the time of treatment, which was completed at least 1 year previously. Saliva was collected from the participants; DNA was extracted and used for CD14 and TLR4 genotyping using the polymerase chain reaction–restriction fragment length polymorphism approach and a real-time polymerase chain reaction TaqMan assay (Applied Biosystems, Foster City, CA), respectively.

Results

No specific genotype or allele of the CD14 and TLR4 genes or any combination thereof was positively associated with post-treatment apical periodontitis (P > .05).

Conclusions

Data from the present study suggest that polymorphisms in the CD14 and TLR4 genes do not influence the response to endodontic treatment of teeth with apical periodontitis.     

内毒素对人牙周膜细胞Toll样受体2和Toll样受体4表达的影响

目的:观察内毒素(LPS)刺激后人牙周膜细胞Toll样受体2和Toll样受体4的表达,探讨牙周膜细胞作为免疫活性细胞在牙周炎局部免疫反应中的作用。方法:组织块法原代培养人牙周膜细胞,经来源鉴定后,取生长良好的第4代细胞用LPS进行刺激,分别于刺激0、4、8、12、24 h,运用免疫细胞化学染色法检测TLR2和TLR4在牙周膜细胞中的表达;利用多功能真彩色细胞分析管理系统进行图像分析,对TLR2和TLR4进行免疫组化评分(IHS)。结果:无LPS刺激的对照组(刺激O h)TLR2和TLR4的表达均为弱阳性,加入LPS后,染色增强,且随着刺激时间的延长而增加(P<0.05),LPS刺激后各时间点TLP2和TLR4的IHS分值与对照组相比差异均有统计学意义(P<0.05)。结论:内毒素的刺激可以使牙周膜细胞TLR2和TLR4表达增多,提示牙周膜细胞参与牙周局部免疫反应。     

Effects of CD14 receptors on tissue reactions induced by local injection of two gram-negative bacterial lipopolysaccharides

Lipopolysaccharide (LPS) was recognized by CD14, which may be an important mediator in the deleterious effects of LPS on the periodontal destruction. To investigate the roles of CD14 molecules on LPS-induced soft tissue inflammation and bone destruction, the tissues of CD14-deficient mice were examined histopathologically following a local injection of either Salmonella minnesota or Porphyromonas gingivalis LPS. In the first group, 12 mice received a local injection of 500 microg of purified P. gingivalis LPS and six mice were injected with saline to the calvaria as controls. In the second group 13 mice were injected subcutaneously on the laterally abdominal skin with 50 microg of S. minnesota LPS and three mice were injected with PBS. Mice were sacrificed at day 5. After histological preparation, the tissue sections of calvaria and soft tissue specimen were stained with tartrate-resistant acid phosphatase (TRAP) marker for osteoclast and macrophage. The soft tissue sections were also stained with hematoxylin & eosin (H&E). Resorption surface and osteoclast index were measured to quantify bone resorption. Necrotic area and inflammatory cell numbers were estimated to assess the situation of local inflammation. Our results indicated that LPS-induced bone resorption is inhibited in CD14-deficient mice. An increase in the number of total inflammatory cells was noticed in both CD14-deficient mice and wild-type mice; however, the cell numbers were less in CD14-deficient mice than those in wild-type mice (two- to three-fold decrease). Therefore, we conclude that the LPS-stimulated bone resorption is mainly via CD14 receptor but the LPS-induced soft tissue inflammation appears to be partially dependent on the receptor.     

TLR2, TLR3, TLR4 and CD14 gene polymorphisms associated with oral lichen planus risk

The aim of this study was to assess whether polymorphisms in toll‐like receptor (TLR) and cluster of differentiation 14 (CD14) genes are associated with oral lichen planus (OLP) risk and clinical course of the disease. The study group consisted of 101 patients with confirmed OLP and 104 healthy blood donors without systemic or oral mucosal diseases. Single nucleotide polymorphisms of TLR2 (rs3804099), TLR3 (rs3775291 and rs5743312), TLR4 (rs4986790 and rs4986791), and CD14 (rs2569190) genes were genotyped using real‐time PCR or PCR‐restriction fragment length polymorphism (PCR‐RFLP). The rs5743312 TLR3 gene polymorphism was associated with increased OLP risk in comparison with the wild type genotype (OR = 15.984, P = 0.011). No association with OLP risk was observed for the polymorphisms studied in TLR2, TLR4 and CD14 genes or for the rs3775291 polymorphism of the TLR3 gene. The polymorphisms of the TLR3 gene were in linkage disequilibrium (D′ = 1, r² = 0.1). Identified haplotypes were not associated with the risk of OLP. The findings of the current study suggest that the TT genotype of the rs5743312 TLR3 gene polymorphism may play a significant role in the aetiology of OLP.     

Expression of CD14, CD16 and CD45RA on monocytes from periodontitis patients

BACKGROUND AND OBJECTIVE: Peripheral blood monocytes are a heterogeneous population, with phenotypes that change on activation or differentiation. Most of the monocytes express lipopolysaccharide (LPS) receptor, CD14 intensely, and do not express Fc gamma receptor III, CD16 (CD14++CD16- monocytes). But monocytes expressing CD16 with reduced CD14 (CD14+CD16+ monocytes) increase in inflammatory diseases as well as sepsis and bacteremia in hemodialysis patients. CD45RA is expressed on activated monocytes, and is regarded as an activation marker of peripheral blood monocytes. The purpose of this study was to determine the phenotypic and functional alteration of monocytes in periodontitis patients. METHODS: Peripheral blood was collected from 33 aggressive periodontitis patients (22 females, 11 males), 55 chronic periodontitis patients (35 females, 20 males) and 30 healthy subjects (16 females, 14 males), and the expression of CD14, CD16 and CD45RA on monocytes was determined using flow cytometry. The production of interleukin-6 (IL-6) by CD16+ and CD16- monocytes stimulated with LPS from Escherichia coli and Actinobacillus actinomycetemcomitans was also examined using flow cytometry. RESULTS: The percentage of CD14+CD16+ monocytes was significantly increased in chronic periodontitis patients. Percentage of monocytes expressing CD45RA was significantly increased in aggressive periodontitis patients compared to healthy subjects. CD16+ and CD16- monocytes produced IL-6 in response to LPS from E. coli and A. actinomycetemcomitans, and the percentage of IL-6 producing cells was higher in CD16+ monocytes than CD16- monocytes, suggesting that CD14+CD16+ monocytes represent a hyper-reactive phenotype. CONCLUSIONS: The present study demonstrated that CD14+CD16+ monocytes and CD45RA+ monocytes were increased in chronic and aggressive periodontitis, respectively. These findings suggest that alteration of monocytes in periodontitis patients could be evaluated by monitoring the surface expression of CD14, CD16 and CD45RA on monocytes.