Although titanate nanofibers (TiNFs) and titanate nanotubes (TiNTs) have been proposed as relatively biocompatible nanomaterials (NMs), there is currently lacking of systemic studies which investigated the toxicity of TiNFs and TiNTs to endothelium. In this study, we developed endothelial monolayer model by using cell culture inserts, and systemically investigated the toxicity of TiNFs and TiNTs by RNA-seq, with a focus on Kruppel-like factor (KLF)-mediated effects, since KLF are transcription factors (TF) involved in the regulation of vascular biology. It was shown that NMs did not significantly induce cytotoxicity despite substantial internalization. However, the expression of many KLF was altered, and Western blot further confirmed that NMs down-regulated KLF2 proteins. Ingenuity pathway analysis (IPA) revealed that NMs altered the expression of KLF2-targed genes, typically the genes involved in inflammatory responses. KLF2-related Gene Ontology (GO) terms and Kyoto Encyclopedia of Gene and Genomes (KEGG) pathways were also altered, and it should be noticed that NMs altered GO terms and KEGG pathways related with endothelial NO synthase (eNOS). This study further verified that NMs decreased intracellular NO and eNOS proteins. All the observed effects were more obvious for TiNFs compared with TiNTs. Combined, this study showed that TiNFs or TiNTs were non-cytotoxic to endothelial monolayer model, but TiNFs and more modestly TiNTs decreased KLF2 leading to decreased eNOS proteins and NO production. Our data may provide novel understanding about the toxicity of TiNFs as well as other Ti-based NMs to endothelium.

尽管已经提出钛酸酯纳米纤维（TiNFs）和钛酸酯纳米管（TiNTs）作为相对生物相容性纳米材料（NMs），但是目前缺乏系统的研究来研究TiNFs和TiNTs对内皮的毒性。在这项研究中，我们通过使用细胞培养插入物建立了内皮单层模型，并通过RNA-seq系统研究了TiNFs和TiNTs的毒性，并重点研究了Kruppel样因子（KLF）介导的作用，因为KLF是转录因子（ TF）参与了血管生物学的调控。结果表明，尽管大量内在化，NMs并没有明显诱导细胞毒性。但是，许多KLF的表达发生了变化，Western印迹进一步证实了NMs下调了KLF2蛋白。机敏途径分析（IPA）显示，NMs改变了KLF2靶基因的表达，该基因通常是与炎症反应有关的基因。KLF2相关基因本体论（GO）术语和《京都议定书》的基因和基因组百科全书（KEGG）途径也发生了变化，应注意的是，NMs改变了与内皮一氧化氮合酶（eNOS）有关的GO术语和KEGG途径。该研究进一步证实了NMs减少了细胞内NO和eNOS蛋白。与TiNTs相比，TiNFs的所有观察到的效果都更加明显。综合起来，这项研究表明，TiNFs或TiNTs对内皮单层模型无细胞毒性，但TiNFs和更适度的TiNTs降低KLF2导致eNOS蛋白和NO生成减少。我们的数据可能提供有关TiNFs和其他基于Ti的NMs对内皮毒性的新颖理解。