已知 Toll 样受体 4 (TLR4)-骨髓分化因子 2 (MD-2) 复合物在炎症中发挥作用。阻断 MD-2 可以抑制炎症过程。然而，MD-2 抑制剂（包括 MAC28、L6H21 和 2i-10）对发炎的人牙髓细胞 (HDPC) 的实际作用从未得到检验。本研究旨在确定这 3 种化合物对脂多糖 (LPS) 处理的 HDPC 的细胞活力、炎症和成骨/成牙分化的药理作用。HDPC 用 10 μM MAC28、L6H21 或 2i-10 预处理 2 小时然后加入 20 μg/mL LPS 或载体 24 小时。使用 3-(4,5-二甲基噻唑-2-基)-2,5-二苯基溴化四唑 (MTT) 测定法评估细胞活力。使用定量实时聚合酶链反应 (qRT-PCR) 和蛋白质印迹分析测定蛋白质 TLR4、MD-2、肿瘤坏死因子 α (TNF-α) 和白细胞介素 6 (IL-6) 的 mRNA 和表达。使用 qRT-PCR 和茜素红染色研究骨/牙源性分化。LPS 不会改变细胞活力，但显着增加 HDPC 中 TLR4、MD-2、TNF-α 和 IL-6 的表达水平，同时骨/牙源性分化与媒介物治疗组相比，能力显着下降。在 LPS 处理的 HDPC 中，MAC28、L6H21 和 2i-10 预处理可减少炎症，并将骨/牙源性分化恢复到与媒介物处理组相似的水平。MAC28、L6H21 和 2i-10 通过下调LPS 处理的 HDPC 中 TLR4-MD-2 信号传导和恢复骨/牙源性分化。这些 MD-2 抑制剂在未来的研究中可被视为治疗牙髓炎症的潜在治疗补充剂。© 2023 中华民国牙科科学协会。 Elsevier B.V. 的出版服务

The toll-like receptor 4 (TLR4)-myeloid differentiation factor 2 (MD-2) complex is known to have a role in inflammation. Blocking MD-2 can suppress inflammatory process. However, the actual action of MD-2 inhibitors, including MAC28, L6H21, and 2i-10, on the inflamed human dental pulp cells (HDPCs) has never been examined. This study aims to determine the pharmacological effects of these 3 compounds on cell viability, inflammation, and osteo/odontogenic differentiation of lipopolysaccharide (LPS)-treated HDPCs.HDPCs were pretreated with 10 μM of MAC28, L6H21, or 2i-10 for 2 h followed by either 20 μg/mL LPS or vehicle for 24 h. Cell viability was assessed using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The mRNA and expression of the proteins TLR4, MD-2, tumor necrosis factor alpha (TNF-α), and interleukin 6 (IL-6) were determined using quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot analysis. Osteo/odontogenic differentiation was investigated using qRT-PCR and Alizarin Red staining.LPS did not alter cell viability but significantly increased the expression levels of TLR4, MD-2, TNF-α, and IL-6 in HDPCs while the osteo/odontogenic differentiation ability decreased significantly when compared to the vehicle-treated group. MAC28, L6H21, and 2i-10-pretreatment in LPS-treated HDPCs reduced inflammation and restored osteo/odontogenic differentiation to similar levels as the vehicle-treated group.MAC28, L6H21, and 2i-10 exhibited equal efficacy in attenuating inflammation through downregulation of TLR4-MD-2 signaling and restored osteo/odontogenic differentiation in LPS-treated HDPCs. These MD-2 inhibitors could be considered as the potential therapeutic supplement for curing inflammation of dental pulp in future studies.© 2023 Association for Dental Sciences of the Republic of China. Publishing services by Elsevier B.V.