Department of Thoracic Surgery, The First Affiliated Hospital of Nanchang University, Nanchang, China.
Department of Anesthesiology, The First Affiliated Hospital of Nanchang University, Nanchang, China.
Transplantation. 2021 Mar 1;105(3):529-539. doi: 10.1097/TP.0000000000003435.
MicroRNA-145 (miR-145) has been shown to play a critical role in ischemia/reperfusion (I/R) injury; however, the expression and function of miR-145 in lung I/R injury have not been reported yet. This study aimed to elucidate the potential effects of miR-145 in lung I/R injury.
Lung I/R mice models and hypoxia/reoxygenation (H/R) pulmonary microvascular endothelial cell models were established. The expression of miR-145 and sirtuin 1 (SIRT1) was measured with reverse transcription-quantitative polymerase chain reaction and Western blot analysis in mouse lung tissue and cells. Artificial modulation of miR-145 and SIRT1 (downregulation) was done in I/R mice and H/R cells. Additionally, Pao2/FiO2 ratio, wet weight-to-dry weight ratio, and cell apoptosis in mouse lung tissues were determined by blood gas analyzer, electronic balance, and deoxyuridine triphosphate-biotin nick end-labeling assay, respectively. Autophagy marker Beclin 1 and LC3 expression, NF-κB acetylation levels, and autophagy bodies were detected in cell H/R and mouse I/R models by Western blot analysis. pulmonary microvascular endothelial cell apoptosis was detected with flow cytometry.
miR-145 was abundantly expressed in the lung tissue of mice and PMVECs following I/R injury. In addition, miR-145 directly targeted SIRT1, which led to significantly decreased Pao2/FiO2 ratio and increased wet weight-to-dry weight ratio, elevated acetylation levels and transcriptional activity of NF-κB, upregulated expressions of tumor necrosis factor-α, interleukins-6, and Beclin 1, autophagy bodies, cell apoptosis, as well as LC3-II/LC3I ratio.
In summary, miR-145 enhances autophagy and aggravates lung I/R injury by promoting NF-κB transcriptional activity via SIRT1 expression.
微小 RNA-145（miR-145）在缺血/再灌注（I/R）损伤中发挥着关键作用；然而，miR-145 在肺 I/R 损伤中的表达和功能尚未见报道。本研究旨在阐明 miR-145 在肺 I/R 损伤中的潜在作用。
建立了肺 I/R 小鼠模型和缺氧/复氧（H/R）肺微血管内皮细胞模型。采用逆转录定量聚合酶链反应和 Western blot 分析检测小鼠肺组织和细胞中 miR-145 和沉默调节蛋白 1（SIRT1）的表达。在 I/R 小鼠和 H/R 细胞中对 miR-145 和 SIRT1（下调）进行人工调节。此外，通过血气分析仪、电子天平、脱氧尿嘧啶三磷酸生物素末端标记法分别测定小鼠肺组织的 Pao2/FiO2 比值、湿重/干重比值和细胞凋亡。通过 Western blot 分析检测细胞 H/R 和小鼠 I/R 模型中的自噬标志物 Beclin 1 和 LC3 表达、NF-κB 乙酰化水平和自噬体。通过流式细胞术检测肺微血管内皮细胞凋亡。
miR-145 在 I/R 损伤后小鼠肺组织和 PMVECs 中大量表达。此外，miR-145 直接靶向 SIRT1，导致 Pao2/FiO2 比值显著降低，湿重/干重比值增加，NF-κB 乙酰化水平和转录活性升高，肿瘤坏死因子-α、白细胞介素-6 和 Beclin 1 表达上调，自噬体形成，细胞凋亡以及 LC3-II/LC3I 比值增加。
综上所述，miR-145 通过促进 SIRT1 表达增强自噬，并通过 NF-κB 转录活性加重肺 I/R 损伤。
