MicroRNAs Regulate Cytokine Responses in Gingival Epithelial Cells
MicroRNAs Regulate Cytokine Responses in Gingival Epithelial Cells
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DOI：
10.1128/iai.00263-16
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发表时间：
2016-12-01
影响因子：
3.1
通讯作者：
Papapanou, Panos N.
中科院分区：
文献类型：
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作者：
Chen, Steven C. Y.;Constantinides, Christos;Papapanou, Panos N.
microRNAs（miRNAs）是参与牙周病病理生理过程的重要调控因子。为了扩大我们早期对临床健康和牙周炎影响的牙龈组织之间特异性miRNA表达差异的观察，我们在功能丧失实验中使用miRNA抑制剂（海绵）来研究特异性miRNA参与口袋上皮来源的端粒酶永生化人牙龈角质形成细胞（TIGKs）对微生物感染的反应。我们构建了5种上皮表达的miRNAs（miR-126、miR-141、miR-155、miR-210和miR-1246）的稳定敲低（KD）细胞系，并使用微阵列分析、定量PCR（qPCR）、酶联免疫吸附测定（ELISA）和Western印迹测定来评估它们对牙周病原体感染的反应。miR-126 KD细胞在热灭活野生型牙龈卟啉单胞菌、活牙龈卟啉单胞菌蛋白酶缺陷突变体KDP 128和活伴放线菌聚集杆菌刺激后，在mRNA和蛋白质水平上显示出比对照更低的白细胞介素8（IL-8）和CXCL 1表达。相反，用相同的生物体感染miR-155 KD和miR-210 KD细胞导致更高的IL-8和CXCL 1 mRNA和蛋白表达。这些作用似乎受NF-κ B调节，如NF-κ B系统组分的转录和/或磷酸化状态改变所示。在对miR-126 KD细胞的感染的响应中观察到减少的嗜中性粒细胞样HL-60细胞趋化活性，表明miR-126在免疫应答中起重要作用。我们的研究结果表明，特定的miRNA调节炎症细胞因子在人类牙龈上皮细胞的表达，以响应微生物感染。
MicroRNAs (miRNAs) have been established as key regulators of various biological processes with possible involvement in the pathobiology of periodontal disease. Expanding our earlier observations of substantial differential expression of specific miRNAs between clinically healthy and periodontitis-affected gingival tissues, we used miRNA inhibitors (sponges) in loss-of-function experiments to investigate the involvement of specific miRNAs in the response of pocket epithelium-derived, telomerase-immortalized human gingival keratinocytes (TIGKs) to microbial infection. We constructed stable knockdown (KD) cell lines for five epithelium-expressed miRNAs (miR-126, miR-141, miR-155, miR-210, and miR-1246) and assessed their response to infection with periodontal pathogens using microarray analysis, quantitative PCR (qPCR), enzyme-linked immunosorbent assay (ELISA), and Western blot assay. miR-126 KD cells showed lower expression of interleukin 8 (IL-8) and CXCL1, both on the mRNA and protein levels, than did controls upon stimulation by heat-killed wild-type Porphyromonas gingivalis, live P. gingivalis protease-deficient mutant KDP128, and live Aggregatibacter actinomycetemcomitans. In contrast, infection of miR-155 KD and miR-210 KD cells with the same organisms resulted in higher IL-8 and CXCL1 mRNA and protein expression. These effects appeared to be regulated by NF-kappa B, as suggested by altered transcription and/or phosphorylation status of components of the NF-kappa B system. Reduced neutrophil-like HL-60 cell chemotactic activity was observed in response to infection of miR-126 KD cells, indicating that miR-126 plays an important role in immune responses. Our findings indicate that specific miRNAs regulate the expression of inflammatory cytokines in human gingival epithelial cells in response to microbial infection.