巴西天然蜂胶对牙龈卟啉单胞菌激活的巨噬细胞极化的调控及机制探讨

发布时间：2018-07-01 12:36

本文选题：慢性牙周炎 + 人巨噬细胞　；参考：《吉林大学》2014年硕士论文

【摘要】：目的与背景：慢性牙周炎是最常见的口腔慢性炎症性疾病之一,它不仅能够导致牙齿松动脱落，还与许多全身系统性疾病相关，包括心血管疾病，糖尿病等[1-3]。P.g是一种革兰氏阴性厌氧菌，它被认为是主要的牙周致病菌，其细胞壁成分内毒素P.g LPS是其致炎的主要毒力因子[4]，P.g LPS通过TLR2和TLR4两个表面受体激活了宿主免疫细胞巨噬细胞，细胞内IκB-α磷酸化后，NF-κB活化进核，诱导炎症相关基因转录，产生促炎因子并分泌，导致了牙周结缔组织损伤和牙槽骨吸收[5]。由于巨噬细胞的抗原呈递和强大的吞噬功能，使其在宿主免疫反应中具有重要的作用，近年来，巨噬细胞的极化现象已成为炎症及免疫研究领域的热点。在慢性牙周炎组织中，巨噬细胞趋化并集中于炎症部位，病原体的入侵将巨噬细胞的表面受体TLR激活，激活的巨噬细胞按功能分为经典激活的巨噬细胞（M1）和替代激活的巨噬细胞（M2），M1通过分泌iNOS，IL-1β，TNF-α等促炎因子，诱导促炎反应，具有强烈的细胞毒性和组织损伤性，在杀灭病原体的同时，促进了牙周结缔组织及骨组织的降解；M2则高表达IL-10，Arginase-1等抗炎因子，诱导抗炎反应，抑制破骨细胞的增殖，具有组织修复和重塑的作用[6-7]。所以，如何调节巨噬细胞使之替代激活，成为炎症反应时减少牙周组织吸收的关键[8-9]。巴西天然蜂胶，古罗马称之为“天然的抗菌素”，实验证明其具有明显的抗细胞氧化，抗炎症反应，促进免疫调节，无毒副作用等特点[10-12]。巴西天然蜂胶中的抗炎抗氧化成分具有血液相容性和细胞膜透过性，可有效的达到病损区发挥作用。本实验组已证明巴西天然蜂胶可抑制脑中巨噬细胞的炎症因子产生[13],因此本实验探讨巴西天然蜂胶是否能够调控巨噬细胞的极化，进而减少促炎因子的产生和分泌。实验方法： 1. HE染色及免疫组织化学法分析人慢性牙周炎牙龈组织中巨噬细胞表面受体及炎症因子的表达及分布。 2. Real-time PCR法检测P.g LPS诱导人巨噬细胞细胞系THP-1的极化。 3. Real-time PCR法及ELISA法检测巴西天然蜂胶对P.g LPS诱导THP-1极化的调控。 4. Western blot法检测巴西天然蜂胶对P.g LPS激活的NF-κB活化的抑制作用。 5.实验结果统计采用GraphPad Prism软件，单因素方差分析法分析。实验结果： 1.人慢性牙周炎牙龈组织中巨噬细胞大量浸润,表面受体TLR2和TLR4高表达，，巨噬细胞呈现M1极化。 2. P.g LPS诱导THP-1向M1极化。 3.巴西天然蜂胶显著抑制P.g LPS诱导的THP-1向M1极化。 4.巴西天然蜂胶抑制P.g LPS诱导的NF-κB活化。结论：证实了巴西天然蜂胶通过抑制P.g LPS激活的THP-1中NF-κB的活化，阻止了THP-1向M1极化。
[Abstract]:Objective and background: chronic periodontitis is one of the most common chronic oral inflammatory diseases. It can not only cause tooth loosening and loss, but also associated with many systemic diseases, including cardiovascular disease. Diabetes, et al. [1-3] .P.g is a gram-negative anaerobic bacterium that is thought to be the leading periodontal pathogen. The cell wall component, endotoxin P.g LPS, is the main virulence factor [4] P.g LPS, which activates the host immune cell macrophages through TLR2 and TLR4 surface receptors. After I 魏 B- 伪 phosphorylation in the host cells, the intracellular I 魏 B- 伪 phosphorylation activates into the nucleus and induces the transcription of inflammatory related genes. Proinflammatory factors are produced and secreted, leading to periodontal connective tissue injury and alveolar bone resorption [5]. Macrophages play an important role in host immune response due to their antigen presentation and strong phagocytosis. In recent years, the polarization of macrophages has become a hot topic in the field of inflammation and immune research. In chronic periodontitis, macrophages become chemotaxis and focus on the inflammatory site, and the invasion of pathogens activates the macrophage surface receptor TLR. Activated macrophages can be divided into classical activated macrophages (M1) and activated macrophages (M2) by secreting pro-inflammatory factors such as iNOS, IL-1 尾 and TNF- 伪. The activated macrophages have strong cytotoxicity and tissue damage. It promoted the degradation of periodontal connective tissue and bone tissue, and increased the expression of anti-inflammatory factors such as IL-10, Arginase-1, induced anti-inflammatory reaction, inhibited the proliferation of osteoclasts, and had the function of tissue repair and remodeling [6-7]. Therefore, how to regulate macrophage replacement activation is the key to reduce periodontal absorption in inflammatory response [8-9]. Brazilian natural propolis, known as "natural antimicrobial" in ancient Rome, has obvious characteristics of anti-cellular oxidation, anti-inflammatory reaction, promoting immune regulation and no toxic and side effects [10-12]. The anti-inflammatory and antioxidant components of natural propolis in Brazil have blood compatibility and membrane permeability, which can effectively reach the diseased area. It has been proved that Brazilian natural propolis can inhibit the production of inflammatory cytokines of macrophages in the brain [13]. Therefore, this experiment is to investigate whether the natural propolis can regulate the polarization of macrophages and thus reduce the production and secretion of proinflammatory factors. Experimental method: 1. The expression and distribution of macrophage surface receptors and inflammatory factors in gingival tissue of chronic periodontitis were analyzed by HE staining and immunohistochemistry. 2. Real-time PCR was used to detect the polarization of human macrophage cell line THP-1 induced by P.g LPS. Real-time PCR and Elisa were used to detect the regulation of natural propolis on the polarization of THP-1 induced by P.g LPS. 4. Western blot assay was used to detect the inhibitory effect of natural propolis on the activation of NF- 魏 B activated by P.g LPS. The results were analyzed by GraphPad Prism software and single factor variance analysis. Experimental results: 1. Macrophages infiltrated in gingival tissue of chronic periodontitis, the surface receptors TLR2 and TLR4 were highly expressed, macrophages showed M1 polarization. 2. P.g LPS induced THP-1 to M1 polarization. 3. Brazilian natural propolis significantly inhibited the polarization of THP-1 to M1 induced by P.g LPS. 4. Brazilian natural propolis inhibited the activation of NF- 魏 B induced by P.g LPS. Conclusion: Brazilian natural propolis inhibits the polarization of THP-1 to M1 by inhibiting the activation of NF- 魏 B in THP-1 activated by P.g LPS.
【学位授予单位】：吉林大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R781.42

【共引文献】