To study the molecular interplay between TLRs and complement representing ancient danger-sensing mechanisms, we examined the regulation of the C3a/anaphylatoxin C3a receptor (C3aR) axis in normal human epidermal keratinocytes (NHEKs) by treatment with different TLR ligands. Protein staining followed by flow cytometry revealed highly constitutive intracellular expression levels of the C3aR in NHEKs. Stimulation with Poly I:C up-regulated C3aR mRNA and intra- and extracellular expression in NHEKs which showed functional relevance by up-regulating CXCL10 and down-regulating C3 expression in response to C3a. mRNA and protein levels of C3 and protease cathepsin L (CTSL) that can cleave C3 were up-regulated by the TLR3 ligand Poly I:C. Enhanced intracellular expression levels of the biologically active C3 fragment (C3a), in response to TLR3 stimulation were also detectable in NHEKs. Cathelicidin antimicrobial peptide LL-37 potentiated Poly I:C-induced C3aR, C3, and CTSL up-regulation. In conclusion, we point to a role of TLR3 to promote up-regulation of C3aR, C3, and CTSL expression levels and generation of C3a. Our data provide evidence that local generation and activation of complement components as described for T cells or myeloid cells represent a scenario which may take place in a similar way in NHEKs.
J Innate Immun

中文翻译：

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C3a 及其受体 C3aR 在正常人表皮角质形成细胞中可检测到，并通过 TLR3 和 LL37 进行差异调节

为了研究 TLR 和代表古老危险感知机制的补体之间的分子相互作用，我们通过用不同的 TLR 配体处理，检查了正常人表皮角质形成细胞 (NHEK) 中 C3a/过敏毒素 C3a 受体 (C3aR) 轴的调节。蛋白质染色和流式细胞术揭示了 NHEK 中 C3aR 的高度组成性细胞内表达水平。用 Poly I:C 刺激可上调 NHEK 中的 C3aR mRNA 和细胞内和细胞外表达，其通过响应 C3a 上调 CXCL10 和下调 C3 表达显示出功能相关性。C3 的 mRNA 和蛋白质水平以及可以切割 C3 的蛋白酶组织蛋白酶 L (CTSL) 被 TLR3 配体 Poly I:C 上调。提高生物活性 C3 片段 (C3a) 的细胞内表达水平，在 NHEK 中也可检测到对 TLR3 刺激的反应。Cathelicidin 抗菌肽 LL-37 增强了 Poly I:C 诱导的 C3aR、C3 和 CTSL 上调。总之，我们指出 TLR3 在促进 C3aR、C3 和 CTSL 表达水平的上调和 C3a 生成方面的作用。我们的数据提供的证据表明，如 T 细胞或骨髓细胞所述的补体成分的局部生成和激活代表了一种可能在 NHEK 中以类似方式发生的情况。
J先天免疫