尼古丁通过调控Toll样受体4抑制牙周膜干细胞的成骨分化能力

发布时间：2018-07-09 17:30

本文选题：尼古丁 + 牙周膜干细胞　；参考：《华西口腔医学杂志》2017年04期

【摘要】：目的探讨尼古丁对牙周膜干细胞(PDLSCs)增殖和成骨分化能力的调控作用,检测尼古丁能否通过调控Toll样受体4(TLR4)抑制PDLSCs的增殖及成骨分化能力。方法培养PDLSCs,采用流式细胞仪检测PDLSCs表面抗原标志,WST-1试剂盒检测不同浓度尼古丁刺激后PDLSCs增殖能力的改变。采用茜素红染色观察PDLSCs经不同浓度尼古丁刺激和成骨诱导后的矿化结节生产情况。运用实时定量聚合酶链反应(RT-PCR)和Western blot检测经尼古丁刺激后,PDLSCs成骨能力相关基因和蛋白的改变,以及尼古丁联合TLR4抑制剂TAK-242刺激后,PDLSCs成骨能力相关基因及蛋白的改变。结果培养的细胞表达间充质干细胞表面标志物CD90和CD105,证实为PDLSCs。与对照组相比,培养3 d后,尼古丁浓度为10-4 mol·L-1的PDLSCs的增殖能力受到明显抑制(P0.05);成骨诱导21 d后,10-4 mol·L-1尼古丁刺激组茜素红染色矿化结节明显减少。RT-PCR反应及Western blot显示:与对照组相比,尼古丁刺激组PDLSCs的碱性磷酸酶、骨钙素、Runt相关转录因子2的基因及蛋白表达量均降低(P0.05),加入TLR4抑制剂TAK-242后,尼古丁的抑制效应减弱。结论尼古丁可能通过TLR4信号通路抑制PDLSCs的增殖及成骨分化能力。
[Abstract]:Objective to investigate the effects of nicotine on the proliferation and osteogenic differentiation of periodontal ligament stem cells (PDLSCs), and to investigate whether nicotine can inhibit the proliferation and osteogenic differentiation of PDLSCs by regulating Toll-like receptor 4 (TLR4). Methods PDLSCs were cultured and the proliferation of PDLSCs was detected by flow cytometry. The mineralized nodules of PDLSCs stimulated by different concentrations of nicotine and induced by osteogenesis were observed by alizarin red staining. Real-time quantitative polymerase chain reaction (RT-PCR) and Western blot were used to detect the changes of osteoblast related genes and proteins in PDLSCs stimulated by nicotine and TLR4 inhibitor TAK-242. Results the cultured cells expressed CD90 and CD105, which were confirmed as PDLSCs. Compared with the control group, the cells were cultured for 3 days. The proliferation of PDLSCs with 10 ~ (-4) mol L ~ (-1) of nicotine was significantly inhibited (P0.05), and the mineralized nodules of alizarin red staining were significantly decreased in the group stimulated by nicotine at 10 ~ (-4) mol L ~ (-1) after osteogenesis (P0.05). The results of RT-PCR and Western blot showed that: compared with the control group, there was no significant difference between the two groups. The expression of alkaline phosphatase and osteocalcin Runt related transcription factor 2 gene and protein in PDLSCs were decreased in nicotine stimulation group (P0.05). The inhibitory effect of nicotine on PDLSCs was weakened by adding TLR4 inhibitor TAK-242. Conclusion nicotine may inhibit the proliferation and osteogenic differentiation of PDLSCs through TLR4 signaling pathway.
【作者单位】：重庆医科大学附属口腔医院牙体牙髓病科口腔疾病与生物医学重庆市重点实验室重庆市高校市级口腔生物医学工程重点实验室;
【基金】：国家自然科学基金面上项目(31571508,31371473) 重庆市第七批重点学科建设项目(牙体牙髓病学,2011年) 重庆市卫生局医学科研计划重点项目(2011-1-062) 重庆市渝北区科技项目[渝北财教(2011)33号]~~
【分类号】：R781.4

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