Objective To investigate the protective effect and mechanism of small RNAs (sRNA) derived from Xuanfei Baidu Formula (宣肺败毒方, XFBD) targeting angiotensin-converting enzyme (ACE) in lipopolysaccharide (LPS)-induced acute lung injury (ALI) in mice. Methods The modified CTAB method was employed to extract sRNA from XFBD and a library was constructed, followed by prediction and screening of sRNAs targeting ACE. The targeting specificity was verified using a dual-luciferase reporter system, and sRNAs capable of suppressing ACE expression were screened in human pulmonary microvascular endothelial cells (HPMEC). The effects of sRNA on angiotensin Ⅱ (Ang II) generation, inhibitor of inhibitor of nuclear factor-κB α (IκBα) and inflammatory cytokine expressions were examined by ELISA, Western blotting and qRT-PCR. An LPS-induced ALI mice model was established, control group, model group, captopril (10 mg/kg) group, XFBD (9.2 g/kg) group, NC-sRNA (10 nmol/animal) group, ACE-sRNA-1 (10 nmol/animal) group and ACE-sRNA-26 (10 nmol/animal) group were set up, with six mice in each group. Hematoxylin-eosin (HE) staining and Micro CT were used to evaluate the pathological and imaging changes of lung tissue; The number of white blood cells and neutrophils in peripheral blood, as well as the total protein concentration, white blood cells, neutrophils and lymphocytes numbers in bronchoalveolar lavage fluid (BALF) were detected; Immunohistochemistry was used to detect the expressions of vascular endothelial-cadherin (VE-cadherin) and intercellular adhesion molecule-1 (ICAM-1) in lung tissue; Western blotting and qRT-PCR were used to detect the expressions of ACE-Ang Ⅱ-Ang Ⅱ type 1 receptor (AT1R) pathway, IκBα and inflammatory factors in lung tissue; ELISA was used to detect the levels of Ang II and inflammatory factors in serum. Results A total of 50 potential ACE-targeting sRNAs were screened from XFBD, with 26 sRNAs validated to exhibit targeting effects, among which 12 sRNAs significantly inhibited ACE expression in HPMEC cells (P < 0.05, 0.01, 0.001). Seven sRNAs significantly suppressed Ang II generation, with ACE-sRNA-1/26 demonstrating the most potent effect (P < 0.001), inhibited IκBα protein and inflammatory factor expressions (P < 0.01, 0.001). In animal experiments, mice in model group had severe lung injury (P < 0.001); Compared with model group, ACE-sRNA-1 and ACE-sRNA-26 significantly improved lung injury in mice (P < 0.05, 0.01, 0.001), up-regulated VE-cadherin expression in lung tissue (P < 0.001), down-regulated ICAM-1, ACE, AT1R, IκBα and inflammatory factor expressions in lung tissue (P < 0.05, 0.01, 0.001), and reduced Ang II and inflammatory factor levels in serum (P < 0.05, 0.01, 0.001). Conclusion ACE-sRNA-1 and ACE-sRNA-26 derived from XFBD could target the inhibition of ACE expression and activity, alleviate pulmonary endothelial inflammation and barrier damage, and improve LPS-induced ALI in mice. The mechanism may be related to the regulation of ACE-Ang Ⅱ-AT1R pathway.

angiotensin-converting enzyme  /  Xuanfei Baidu Formula  /  sRNA  /  acute lung injury  /  inflammation  /  naringin  /  polydatin  /  agnuside  /  amygdalin  /  glycyrrhizic acid  /  cornin  /  sinapine  /  verbascoside  /  liquiritin  /  ephedrine