Department of Infectious Diseases, Peking University First Hospital, No.8 Xishiku Street, Xicheng District, Beijing, 100034, China.
Peking University Ditan Teaching Hospital, Beijing 100015, China; Beijing; Key Laboratory of Emerging Infectious Diseases, Beijing, 100015, China.
Respir Res. 2019 Jul 22;20(1):163. doi: 10.1186/s12931-019-1102-2.
Pulmonary fibrosis is a progressive and irreversible disease for which therapeutic options are currently limited. A recent in vivo study showed that tenofovir, a nucleotide analogue reverse transcriptase inhibitor, had direct antifibrotic effects on skin and liver fibrosis. Another study in vitro revealed that NS5ATP9 inhibited the activation of human hepatic stellate cells. Because of the similarity of fibrotic diseases, we hypothesized that tenofovir alafenamide fumarate (TAF), the prodrug of tenofovir, and NS5ATP9, is related to and plays a role in the suppression of pulmonary fibrosis.
We investigated the influence of NS5ATP9 on fibrosis in vitro. Human lung fibroblasts (HFL1) were transfected with short interfering RNAs or overexpression plasmids of NS5ATP9 before stimulation by human recombinant transforming growth factor-β1. The effect of TAF was evaluated in a bleomycin-induced fibrosis murine model. Male C57BL/6 mice were treated with bleomycin on day 0 by intratracheal injection and intragastrically administered TAF or vehicle. Left lung sections were fixed for histological analysis, while homogenates of the right lung sections and HFL1 cells were analyzed by western blotting and quantitative reverse transcription polymerase chain reaction.
NS5ATP9 suppressed the activation of lung fibroblasts. Upregulation of collagen type 3 (α 1 chain) and α-smooth muscle actin was observed in HFL1 cells when NS5ATP9 was silenced, and vice-versa. TAF also showed anti-fibrotic effects in mice, as demonstrated by histological analysis of fibrosis and expression of extracellular matrix components in the lung sections. Additionally, TAF inhibited transforming growth factor-β1 and phosphorylated-Smad3 synthesis in HFL1 cells and the murine model, which was accompanied by upregulation of NS5ATP9.
Our results suggest that NS5ATP9 forms a negative feedback pathway in pulmonary fibrosis and TAF has anti-fibrotic properties as it upregulates the expression level of NS5ATP9. As TAF has been shown to be safe and well-tolerated in humans, TAF and NS5ATP9 may be useful for developing novel therapeutics for pulmonary fibrosis.
肺纤维化是一种进行性和不可逆转的疾病，目前治疗选择有限。最近的一项体内研究表明，核苷酸类似物逆转录酶抑制剂替诺福韦对皮肤和肝脏纤维化具有直接的抗纤维化作用。另一项体外研究表明，NS5ATP9 抑制人肝星状细胞的激活。由于纤维化疾病的相似性，我们假设替诺福韦前药富马酸替诺福韦二吡呋酯（TAF）和 NS5ATP9 相关，并在抑制肺纤维化中发挥作用。
我们研究了 NS5ATP9 对体外纤维化的影响。在人重组转化生长因子-β1 刺激前，用人肺成纤维细胞（HFL1）的短发夹 RNA 或过表达质粒转染 NS5ATP9。在博来霉素诱导的纤维化小鼠模型中评估 TAF 的作用。雄性 C57BL/6 小鼠在第 0 天通过气管内注射和胃内给予 TAF 或载体处理博来霉素。将左肺切片固定进行组织学分析，同时分析右肺切片和 HFL1 细胞的匀浆进行 Western blot 和定量逆转录聚合酶链反应。
NS5ATP9 抑制肺成纤维细胞的激活。当 NS5ATP9 被沉默时，在 HFL1 细胞中观察到胶原蛋白 3（α1 链）和α-平滑肌肌动蛋白的上调，反之亦然。TAF 在小鼠中也表现出抗纤维化作用，如纤维化的组织学分析和肺组织中细胞外基质成分的表达所示。此外，TAF 抑制 HFL1 细胞和小鼠模型中转化生长因子-β1 和磷酸化-Smad3 的合成，同时上调 NS5ATP9。
我们的结果表明，NS5ATP9 在肺纤维化中形成负反馈途径，TAF 具有抗纤维化特性，因为它上调 NS5ATP9 的表达水平。由于 TAF 已被证明在人体中是安全且耐受良好的，因此 TAF 和 NS5ATP9 可能有助于开发治疗肺纤维化的新疗法。
