Department of Microbiology and Immunology, Graduate School of Medicine, Hokkaido University, Sapporo, Japan.
PLoS One. 2010 Sep 2;5(9):e12550. doi: 10.1371/journal.pone.0012550.
Natural killer (NK) cells are lymphocyte effectors that are activated to control certain microbial infections and tumors. Many NK-activating and regulating receptors are involved in regulating NK cell function. In addition, activation of naïve NK cells is fundamentally triggered by cytokines or myeloid dendritic cells (mDC) in various modes. In this study, we synthesized 16 S-[2,3-bis(palmitoyl)propyl]cysteine (Pam2Cys) lipopeptides with sequences designed from lipoproteins of Staphylococcus aureus, and assessed their functional properties using mouse (C57BL/6) bone marrow-derived DC (BMDC) and NK cells. NK cell activation was evaluated by three criteria: IFN-gamma production, up-regulation of NK activation markers and cytokines, and NK target (B16D8 cell) cytotoxicity. The diacylated lipopeptides acted as TLR2 ligands, inducing up-regulation of CD25/CD69/CD86, IL-6, and IL-12p40, which represent maturation of BMDC. Strikingly, the Pam2Cys lipopeptides induced mouse NK cell activation based on these criteria. Cell-cell contact by Pam2Cys peptide-stimulated BMDC and NK cells rather than soluble mediators released by stimulated BMDC induced activation of NK cells. For most lipopeptides, the BMDC TLR2/MyD88 pathway was responsible for driving NK activation, while some slightly induced direct activation of NK cells via the TLR2/MyD88 pathway in NK cells. The potential for NK activation was critically regulated by the peptide primary sequence. Hydrophobic or proline-containing sequences proximal to the N-terminal lipid moiety interfered with the ability of lipopeptides to induce BMDC-mediated NK activation. This mode of NK activation is distinctly different from that induced by polyI:C, which is closely associated with type I IFN-inducing pathways of BMDC. These results imply that the MyD88 pathway of BMDC governs an alternative NK-activating pathway in which the peptide sequence of TLR2-agonistic lipopeptides critically affects the potential for NK activation.
自然杀伤 (NK) 细胞是被激活以控制某些微生物感染和肿瘤的淋巴细胞效应物。许多 NK 激活和调节受体参与调节 NK 细胞功能。此外，在各种模式下，幼稚 NK 细胞的激活基本上是由细胞因子或髓样树突状细胞 (mDC) 触发的。在这项研究中，我们合成了 16S-[2,3-双(棕榈酰)丙基]半胱氨酸 (Pam2Cys) 脂肽，其序列是根据金黄色葡萄球菌脂蛋白设计的，并使用小鼠 (C57BL/6) 骨髓来源的 DC (BMDC) 和 NK 细胞评估它们的功能特性。通过三种标准评估 NK 细胞的激活：IFN-γ产生、NK 激活标志物和细胞因子的上调以及 NK 靶标 (B16D8 细胞) 的细胞毒性。二酰化脂肽作为 TLR2 配体起作用，诱导 CD25/CD69/CD86、IL-6 和 IL-12p40 的上调，这代表 BMDC 的成熟。引人注目的是，Pam2Cys 脂肽根据这些标准诱导小鼠 NK 细胞的激活。由 Pam2Cys 肽刺激的 BMDC 和 NK 细胞之间的细胞-细胞接触而不是由刺激的 BMDC 释放的可溶性介质诱导 NK 细胞的激活。对于大多数脂肽，BMDC TLR2/MyD88 途径负责驱动 NK 激活，而一些脂肽通过 TLR2/MyD88 途径在 NK 细胞中略微诱导 NK 细胞的直接激活。NK 激活的潜力受到肽一级序列的严格调节。靠近 N 端脂质部分的疏水性或脯氨酸含量的序列干扰了脂肽诱导 BMDC 介导的 NK 激活的能力。这种 NK 激活模式与与 BMDC 的 I 型 IFN 诱导途径密切相关的聚肌胞苷酸诱导的模式明显不同。这些结果表明，BMDC 的 MyD88 途径控制着一种替代的 NK 激活途径，其中 TLR2 激动性脂肽的肽序列对 NK 激活的潜力有重要影响。
