Advanced glycation end-products increase IL-6 and ICAM-1 expression via RAGE, MAPK and NF-κB pathways in human gingival fibroblasts
Advanced glycation end-products increase IL-6 and ICAM-1 expression via RAGE, MAPK and NF-κB pathways in human gingival fibroblasts
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DOI：
10.1111/jre.12518
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发表时间：
2018-06-01
影响因子：
3.5
通讯作者：
Kido, J.
中科院分区：
文献类型：
--
作者：
Nonaka, K.;Kajiura, Y.;Kido, J.
背景和目的：糖尿病(DM)是牙周病的危险因素，可加重牙周炎的发病过程。晚期糖基化终末产物(AGEs)会导致糖尿病并发症，与血糖控制水平相关，并且在牙周炎和糖尿病患者的牙周组织中积累更多的AGEs。本研究通过观察AGEs对人牙周成纤维细胞(HGFs)炎症相关因子表达的影响，探讨AGEs对糖尿病相关牙周炎的影响。检测细胞活力，并从AGE处理的细胞中分离出RNA和蛋白质组分。采用逆转录-聚合酶链式反应、实时定量聚合酶链式反应和酶联免疫吸附试验检测细胞因子IL-6、细胞间黏附分子-1(ICAM-1)和糖基化终产物受体(RAGE)的表达，并用2，7-二氯二乙酸荧光素试剂盒检测细胞内活性氧的活性。用荧光试剂标记的人单核细胞(THP-1)与经AGEs和IL-6 siRNA处理的HGFs共同培养，检测THP-1细胞与HGFs的黏附活性。用重组人IL-6、RAGE和IL-6的siRNAs以及MAPK和NF-B的抑制剂处理HGFs，观察AGEs对HGFs IL-6和ICAM-1表达的影响。结果：AGEs促进HGFRAGE、IL-6、ICAM-1mRNA和蛋白的表达，增强细胞内活性氧的活性，促进THP-1细胞与HGFs的黏附，但对细胞存活率无明显影响。重组人IL-6促进HGFICAM-1的表达，RAGE和IL-6的siRNAs抑制AGE诱导的IL6和ICAM1mRNA的表达，IL-6siRNA抑制AGE诱导的THP-1细胞黏附。结论：AGEs通过RAGE、MAPK和NF-kappaB途径促进HGFIL-6和ICAM-1的表达，可能加重牙周疾病的发生发展。
Background and Objectives: Diabetes mellitus (DM) is a risk factor for periodontal diseases and may exacerbate the progression of the pathogenesis of periodontitis. Advanced glycation end-products (AGEs) cause DM complications relative to levels of glycemic control and larger amounts accumulate in the periodontal tissues of patients with periodontitis and DM. In the present study, we investigated the effects of AGEs on the expression of inflammation-related factors in human gingival fibroblasts (HGFs) to elucidate the impact of AGEs on DM-associated periodontitis.Material and Methods: HGFs were cultured with or without AGEs. Cell viability was examined, and RNA and protein fractions were isolated from AGE-treated cells. The expression of interleukin (IL)-6, intercellular adhesion molecule-1 (ICAM-1), and the receptor for AGE (RAGE) was investigated using reverse transcription-polymerase chain reaction, quantitative real-time polymerase chain reaction and enzyme-linked immunosorbent assay, and reactive oxygen species activity was measured using a kit with 2,7-dichlorofluorescin diacetate. Human monocytic cells (THP-1) labeled with a fluorescent reagent were co-cultured with HGFs treated with AGEs and IL-6 siRNA, and the adhesive activity of THP-1 cells to HGFs was assessed. The expression of IL-6 and ICAM-1 was examined when HGFs were pretreated with recombinant human IL-6, the siRNAs of RAGE and IL-6, and inhibitors of MAPK and NF-B, and then cultured with and without AGEs. The phosphorylation of MAPK and NF-B was assessed using western blotting.Results: AGEs increased the mRNA and protein expressions of RAGE, IL-6, ICAM-1 and reactive oxygen species activity in HGFs, and promoted the adhesion of THP-1 cells to HGFs, but had no effect on cell viability until 72hours. Recombinant human IL-6 increased ICAM-1 expression in HGFs, while the siRNAs of RAGE and IL-6 inhibited AGE-induced IL6 and ICAM1 mRNA expression, and IL-6 siRNA depressed AGE-induced THP-1 cell adhesion. AGEs increased the phosphorylation of p38 and ERK MAPKs, p65 NF-kappa B and IB, while inhibitors of p38, ERK MAPKs and NF-kappa B significantly decreased AGE-induced IL-6 and ICAM-1 expression.Conclusion: AGEs increase IL-6 and ICAM-1 expression via the RAGE, MAPK and NF-kappa B pathways in HGFs and may exacerbate the progression of the pathogenesis of periodontal diseases.