川芎嗪通过增加 miR-193a 的表达抑制 PI3K/Akt/mTOR 和 hedgehog 信号通路，从而增加肺细胞自噬，改善百草枯诱导的肺纤维化。
Ligustrazin increases lung cell autophagy and ameliorates paraquat-induced pulmonary fibrosis by inhibiting PI3K/Akt/mTOR and hedgehog signalling via increasing miR-193a expression.
机构信息
Department of Emergency, the First Affiliated Hospital of Kunming Medical University, 295 Xichang Road, Kunming, 650032, Yunan, People's Republic of China.
Emergency Intensive Care Unit, the Second Affiliated Hospital of Kunming Medical University, Kunming, 650106, China.
出版信息
BMC Pulm Med. 2019 Feb 11;19(1):35. doi: 10.1186/s12890-019-0799-5.
BACKGROUND
Reactive oxygen species (ROS) levels largely determine pulmonary fibrosis. Antioxidants have been found to ameliorate lung fibrosis after long-term paraquat (PQ) exposure. The effects of antioxidants, however, on the signalling pathways involved in PQ-induced lung fibrosis have not yet been investigated sufficiently. Here, we examined the impacts of ligustrazin on lung fibrosis, in particular ROS-related autophagy and pro-fibrotic signalling pathways, using a murine model of PQ-induced lung fibrosis.
METHODS
We explored the effects of microRNA-193 (miR-193a) on Hedgehog (Hh) and PI3K/Akt/mTOR signalling and oxidative stress in lung tissues. Levels of miR-193a, protein kinase B (Akt), phosphoinositide 3-Kinase (PI3K), ceclin1, mammalian target of rapamycin (mTOR), sonic hedgehog (SHH), myosin-like Bcl2 interacting protein (LC3), smoothened (Smo), and glioma-associated oncogene-1 (Gli-1) mRNAs were determined with quantitative real-time PCR. Protein levels of PI3K, p-mTOR, p-Akt, SHH, beclin1, gGli-1, LC3, smo, transforming growth factor-β1 (TGF-β1), mothers against DPP homologue-2 (Smad2), connective tissue growth factor (CTGF), collagen I, collagen III, α-smooth muscle actin (α-SMA) nuclear factor erythroid 2p45-related factor-2 (Nrf2), and p-Smad2 were detected by western blotting. In addition, α-SMA, malondialdehyde, ROS, superoxide dismutase (SOD), oxidised and reduced glutathione, hydroxyproline, and overall collagen levels were identified in lung tissues using immunohistochemistry.
RESULTS
Long-term PQ exposure blocked miR-193a expression, reduced PI3K/Akt/mTOR signalling, increased oxidative stress, inhibited autophagy, increased Hh signalling, and facilitated the formation of pulmonary fibrosis. Ligustrazin blocked PI3K/Akt/mTOR and Hh signalling as well as reduced oxidative stress via increasing miR-193a expression and autophagy, all of which reduced pulmonary fibrosis. These effects of ligustrazin were accompanied by reduced TGF-β1, CTGF, and Collagen I and III expression.
CONCLUSIONS
Ligustrazin blocked PQ-induced PI3K/Akt/mTOR and Hh signalling by increasing miR-193a expression, thereby attenuating PQ-induced lung fibrosis.
背景
活性氧（ROS）水平在很大程度上决定了肺纤维化。抗氧化剂已被发现可改善长期百草枯（PQ）暴露后的肺纤维化。然而，抗氧化剂对百草枯诱导的肺纤维化相关信号通路的影响尚未得到充分研究。在这里，我们使用百草枯诱导的肺纤维化小鼠模型，研究了川芎嗪对肺纤维化、特别是 ROS 相关自噬和促纤维化信号通路的影响。
方法
我们探讨了 microRNA-193（miR-193a）对 Hedgehog（Hh）和 PI3K/Akt/mTOR 信号以及肺组织氧化应激的影响。采用实时定量 PCR 检测 miR-193a、蛋白激酶 B（Akt）、磷酸肌醇 3-激酶（PI3K）、ceclin1、雷帕霉素靶蛋白（mTOR）、sonic hedgehog（SHH）、肌球蛋白样 Bcl2 相互作用蛋白（LC3）、smoothened（Smo）和 Glioma-associated oncogene-1（Gli-1）mRNA 的水平。Western blot 检测 PI3K、p-mTOR、p-Akt、SHH、beclin1、gGli-1、LC3、 smo、转化生长因子-β1（TGF-β1）、Mothers against DPP homolog 2（Smad2）、结缔组织生长因子（CTGF）、胶原 I、胶原 III、α-平滑肌肌动蛋白（α-SMA）核因子红细胞 2p45 相关因子-2（Nrf2）和 p-Smad2 的蛋白水平。此外，采用免疫组织化学法检测肺组织中α-SMA、丙二醛、ROS、超氧化物歧化酶（SOD）、氧化型和还原型谷胱甘肽、羟脯氨酸和总胶原水平。
结果
长期 PQ 暴露抑制 miR-193a 表达，降低 PI3K/Akt/mTOR 信号，增加氧化应激，抑制自噬，增加 Hh 信号，促进肺纤维化形成。川芎嗪通过增加 miR-193a 表达和自噬来阻断 PI3K/Akt/mTOR 和 Hh 信号，并减少氧化应激，从而减少肺纤维化。这些川芎嗪的作用伴随着 TGF-β1、CTGF 和胶原 I 和 III 表达的减少。
结论
川芎嗪通过增加 miR-193a 表达阻断 PQ 诱导的 PI3K/Akt/mTOR 和 Hh 信号，从而减轻 PQ 诱导的肺纤维化。
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