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Eldecalcitol Inhibits LPS-Induced NLRP3 Inflammasome-Dependent Pyroptosis in Human Gingival Fibroblasts by Activating the Nrf2/HO-1 Signaling Pathway
Drug Design, Development and Therapy ( IF 4.8 ) Pub Date : 2020-11-13 , DOI: 10.2147/dddt.s269223
Cancan Huang 1 , Chaotao Zhang 1 , Panpan Yang 1 , Rui Chao 1 , Ziqi Yue 1 , Congshan Li 1 , Jie Guo 1 , Minqi Li 1
Affiliation  

Purpose: Periodontitis is a major chronic oral disease that is accelerated by activation of the NLRP3 inflammasome and the resulting pyroptosis. According to recent studies, active vitamin D and its analogs have been reported to have great anti-inflammatory effects. However, the anti-inflammatory mechanism of a newly found vitamin D analog, eldecalcitol (ED-71), is still unclear. This study investigates whether ED-71 could protect human gingival fibroblasts (HGFs) from LPS-induced pyroptosis and, if so, determine its underlying mechanism.
Methods: After HGFs were treated with LPS alone or with LPS and ED-71, their viability was measured by CCK8 assay. The degrees of inflammation and pyroptosis were measured via LDH assay, H2O2 assay, fluorescent staining, flow cytometry, and Western blots. Intracellular ROS, Hoechst 33,342, and PI stains were assessed with a fluorescence microscope. ROS inhibitor NAC, NLRP3 inhibitor MCC950, and Nrf2 inhibitor ML385 were added to further clarify the mechanism.
Results: LPS induced cytotoxicity in HGFs, as shown by CCK8 assay. LPS also increased intracellular ROS, H2O2 levels, release of LDH, and expression of the pyroptosis-related proteins NLRP3, caspase-1, and IL-1β. NAC and MCC950 reduced LPS-induced NLRP3, caspase-1, and IL-1β. Pretreatment with ED-71 effectively inhibited the LPS-induced pyroptosis and was associated with activation of the Nrf2/HO-1 signaling pathway. This beneficial effect of ED-71 was suppressed by ML385.
Conclusion: This study demonstrates the therapeutic effect of ED-71 on LPS-induced NLRP3 inflammasome-dependent pyroptosis in HGFs and further reveals that ED-71 can inhibit pyroptosis by activating the Nrf2/HO-1 pathway. Our results thus suggest that ED-71 is a potential candidate for the treatment of periodontitis.

Keywords: HGFs, pyroptosis, NLRP3, ED-71, Nrf2/HO-1


中文翻译:

Eldecalcitol 通过激活 Nrf2/HO-1 信号通路抑制 LPS 诱导的 NLRP3 炎症小体依赖性人牙龈成纤维细胞焦亡

目的:牙周炎是一种主要的慢性口腔疾病,可通过激活 NLRP3 炎性体和由此产生的细胞焦亡而加速。根据最近的研究,据报道活性维生素 D 及其类似物具有很好的抗炎作用。然而,新发现的维生素 D 类似物 eldecalcitol (ED-71) 的抗炎机制仍不清楚。本研究调查 ED-71 是否可以保护人牙龈成纤维细胞 (HGF) 免受 LPS 诱导的细胞焦亡,如果可以,确定其潜在机制。
方法: HGFs 单独用 LPS 或用 LPS 和 ED-71 处理后,通过 CCK8 法测定其活力。炎症和焦亡的程度通过LDH测定、H 2 O 2测量测定、荧光染色、流式细胞术和蛋白质印迹。用荧光显微镜评估细胞内 ROS、Hoechst 33,342 和 PI 染色。添加了 ROS 抑制剂 NAC、NLRP3 抑制剂 MCC950 和 Nrf2 抑制剂 ML385 以进一步阐明机制。
结果: LPS 在 HGF 中诱导细胞毒性,如 CCK8 测定所示。LPS 还增加了细胞内 ROS、H 2 O 2水平、LDH 的释放以及细胞焦亡相关蛋白 NLRP3、caspase-1 和 IL-1β 的表达。NAC 和 MCC950 降低了 LPS 诱导的 NLRP3、caspase-1 和 IL-1β。ED-71 预处理可有效抑制 LPS 诱导的细胞焦亡,并与 Nrf2/HO-1 信号通路的激活有关。ED-71 的这种有益作用被 ML385 抑制。
结论:本研究证实了 ED-71 对 LPS 诱导的 NLRP3 炎症小体依赖性 HGFs 细胞焦亡的治疗作用,并进一步揭示 ED-71 可通过激活 Nrf2/HO-1 通路抑制细胞焦亡。因此,我们的结果表明 ED-71 是治疗牙周炎的潜在候选者。

关键词: HGFs,细胞焦亡,NLRP3,ED-71,Nrf2/HO-1
更新日期:2020-11-13
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