Del⁃1在人牙周膜成纤维细胞中的表达及相关炎症机制初探
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国家自然科学基金(81771074,81470749);江苏省高校自然科学研究重大项目(16KJA320001);江苏省高层次卫生人才“六个一工程”(LGY2016048);江苏省高校优势学科建设工程资助(2014-37)


Expression of Del⁃1 in human periodontal ligament cells and its related inflammatory mechanism
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    摘要:

    目的:检测Del-1蛋白在人牙周膜成纤维细胞中的表达情况,并观察内毒素(lipopolysaccharides,LPS)对人牙周膜成纤维细胞Del-1表达的影响,探讨其在牙周炎进展中的可能作用机制。方法:组织块法取人牙周膜成纤维细胞,用细胞免疫荧光、流式细胞术及体外管腔形成试验鉴别细胞来源,细胞免疫荧光检测Del-1蛋白在人牙周膜成纤维细胞中的表达情况;用不同浓度LPS刺激细胞,检测Del-1蛋白的表达水平,以Del-1减少最显著的LPS浓度为最佳刺激浓度,再用该浓度的LPS刺激细胞不同时间,检测Del-1及炎症因子IL-6的表达水平;用不同浓度的Del-1蛋白预刺激细胞1 h,再用最佳浓度的LPS刺激细胞24 h,检测炎症因子IL-6的表达情况;用最适宜浓度的Del-1蛋白预刺激细胞1 h,再用LPS刺激细胞10 min,检测IκBα的激活情况。结果:Del-1蛋白在人牙周膜成纤维细胞中有表达,定位于细胞的胞浆中;LPS刺激细胞时Del-1的表达水平降低,并随浓度的升高和处理时间的延长而下降,IL-6的表达与之相反;Del-1蛋白可下调LPS诱导的炎症因子IL-6的表达,抑制IκBα的磷酸化。结论:本研究首次发现Del-1可以在人牙周膜成纤维细胞中表达,且在LPS刺激人牙周膜成纤维细胞时表达量降低,Del-1蛋白可通过抑制NF-κB通路的激活,在牙周炎的发生发展中发挥拮抗作用。

    Abstract:

    Objective:To investigate whether Del-1 is expressed in human periodontal ligament cells(hPDLCs),and the effect of lipopolysaccharides(LPS)on the expression of Del-1 in hPDLCs,and to explore the potential mechanism of Del-1 in the development of periodontitis. Methods:HPDLCs were isolated and cultured by tissue block method,and immunofluorescence assay,flow cytometry and Matrigel assay were used to identify cells. Immunofluorescence was used to detect if Del-1 was expressed in hPDLCs. Then variable concentrations of LPS were used to stimulate hPDLCs,and the expression of Del-1 was detected. The optimal concentration of LPS was selected,based on the most significant decrease of Del-1. The optimal concentration was selected to be used in the time-dependent experiment. The expressions of Del-1 and IL-6 were respectively assayed. hPDLCs were pretreated with Del-1 of variable concentrations(0,0.05,0.5 and 5 μg/mL)for 1 h,and then stimulated with LPS for 24 h. The expressions of IL-6 were assayed. hPDLCs were pretreated with the optimal concentration of Del-1 for 1 h,and then stimulated with LPS for 10 min,for detecting the phosphorylation of IκBα. Results:Del-1 could be expressed in hPDLCs,locating in cytoplasm of hPDLCs. The expressions of Del-1 were significantly down-regulated by LPS in a dose-independent and time-independent manner,which were contrary to IL-6. Del-1 could down-regulate the expression of IL-6 which was induced by LPS and inhibit the phosphorylation of IκBα in hPDLCs. Conclusions:Our study discovered for the first time that Del-1 could be expressed in hPDLCs,and it was down-regulated by LPS in hPDLCs,Del-1 could suppress the phosphorylation of NF-κB,playing an antagonistic role in the development of periodontitis.

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王玉珍,陈 旭,吴偲偲,葛 舒,徐 艳,叶 宇. Del⁃1在人牙周膜成纤维细胞中的表达及相关炎症机制初探[J].南京医科大学学报(自然科学版),2018,(7):915-921,927

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  • 收稿日期:2018-02-25
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  • 在线发布日期: 2018-07-20
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