Center of Excellence in Regenerative Dentistry, Faculty of Dentistry, Chulalongkorn University, Bangkok, Thailand.
Oral Biology Graduate Program, Faculty of Dentistry, Chulalongkorn University, Bangkok, Thailand.
J Periodontol. 2020 Sep;91(9):1225-1236. doi: 10.1002/JPER.19-0551. Epub 2020 Feb 14.
Toll-like receptors (TLR) are a group of receptors that play roles in the innate immune system. Human periodontal ligament cells (hPDL cells) express several TLRs, including TLR3, a nucleotide sensing receptor that recognizes double-stranded RNA from viral infection. However, its role in hPDL cells is unclear. The aim of this study was to investigate the responses of hPDL cells in terms of immunomodulation after TLR3 engagement.
HPDL cells were treated with various doses of poly I:C, a TLR3 activator. The expression of interferon-gamma (IFNγ), indoleamine 2,3 dioxygenase (IDO), and human leukocyte antigen G (HLA-G) was determined. Chemical inhibitors and small interfering RNA (siRNA) were used to confirm the role of TLR3. Coculture with human peripheral blood mononuclear cells (PBMCs) with poly I:C-activated hPDL cells was performed.
Endosomal TLR3 in hPDL cells was observed by immunocytochemistry. Addition of poly I:C significantly enhanced the expression and secretion of IFNγ, IDO, and HLA-G. Knockdown of TLR3 using siRNA decreased the poly I:C-induced expression of these three molecules. Bafilomycin-A, an inhibitor of auto-phagosome and lysosome fusion, inhibited poly I:C-induced IDO and HLA-G expression, whereas cycloheximide and a TLR3-neutralizing antibody had no effect. In co-culture experiments, poly I:C-activated hPDL cells inhibited PBMCs proliferation and increased mRNA expression of forkhead box P3 (FOXP3), a transcription factor which is a marker of regulatory T cells.
Our findings indicated that TLR3 engagement of hPDL cells induced immunosuppressive properties of these cells. Because immunosuppressive properties play an important role in tissue healing and regeneration, activation of TLR3 may help to attenuate tissue destruction by limiting the inflammatory process and perhaps initiate the healing and regeneration process of the periodontium.
Toll 样受体（TLR）是一组在固有免疫系统中发挥作用的受体。人牙周韧带细胞（hPDL 细胞）表达多种 TLR，包括 TLR3，这是一种核苷酸感应受体，可识别病毒感染的双链 RNA。然而，其在 hPDL 细胞中的作用尚不清楚。本研究旨在探讨 TLR3 激活后 hPDL 细胞在免疫调节方面的反应。
用不同剂量的 poly I:C（TLR3 激活剂）处理 hPDL 细胞。测定干扰素-γ（IFNγ）、吲哚胺 2,3 双加氧酶（IDO）和人类白细胞抗原 G（HLA-G）的表达。使用化学抑制剂和小干扰 RNA（siRNA）来确认 TLR3 的作用。用 poly I:C 激活的 hPDL 细胞与人外周血单核细胞（PBMC）共培养。
免疫细胞化学观察到 hPDL 细胞内体 TLR3。添加 poly I:C 可显著增强 IFNγ、IDO 和 HLA-G 的表达和分泌。用 siRNA 敲低 TLR3 可降低 poly I:C 诱导的这三种分子的表达。自噬体和溶酶体融合的抑制剂巴弗洛霉素-A 抑制 poly I:C 诱导的 IDO 和 HLA-G 表达，而环己酰亚胺和 TLR3 中和抗体则没有作用。在共培养实验中，poly I:C 激活的 hPDL 细胞抑制 PBMC 增殖，并增加叉头框 P3（FOXP3）的 mRNA 表达，FOXP3 是调节性 T 细胞的标志物。
我们的研究结果表明，TLR3 激活 hPDL 细胞可诱导这些细胞的免疫抑制特性。由于免疫抑制特性在组织愈合和再生中起着重要作用，因此激活 TLR3 可能有助于通过限制炎症过程来减轻组织破坏，并可能启动牙周组织的愈合和再生过程。
