[Mechanism of Xuebijing Injection in ameliorating acute lung injury by promoting efferocytosis based on proteomics...

This study aims to elucidate the mechanism by which Xuebijing Injection ameliorates acute lung injury(ALI) through promoting efferocytosis and suppressing inflammatory responses based on proteomics and experimental validation. Forty male Sprague Dawley(SD) rats were randomly divided into a normal control group and a model group. An ALI rat model was established by intraperitoneal injection of lipopolysaccharide(LPS). Successfully modeled rats were then randomized into three groups: a model control group, an Xuebijing Injection(8 mL·kg~(-1)) group, and a dexamethasone(5 mg·kg~(-1)) group, with 10 rats in each group. Intraperitoneal injection was administered for three days. Lung edema was evaluated by the wet/dry weight(W/D) ratio of lung tissue. Inflammatory infiltration and injury in lung tissue were assessed by hematoxylin-eosin(HE) staining, and the level of inflammatory cytokines, including interleukin(IL)-17, IL-18, and IL-4, as well as the anti-inflammatory molecule transforming growth factor-β1(TGF-β1) in serum was measured by enzyme-linked immunosorbent assay(ELISA). Differentially expressed genes(DEGs) in lung tissue among the normal control group, the model control group, and the Xuebijing Injection group were analyzed by proteomic sequencing, and Gene Ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG) were utilized for pathway enrichment analysis. Apoptotic cells were quantified by terminal deoxynucleotidyl transferase dUTP nick end labeling(TUNEL) assay, and the expression of efferocytosis-related key factors in lung tissue including protein S1(ProS1), Mer proto-oncogene tyrosine kinase(MerTK), and Ras-related C3 botulinum toxin substrate 1(Rac1) was evaluated by quantitative polymerase chain reaction(qPCR), Western blot, and immunofluorescence. The results of the W/D ratio of lung tissue and pathological staining showed that lung edema, inflammatory infiltration, and injury in ALI rats were ameliorated after Xuebijing Injection intervention. ELISA demonstrated that Xuebijing Injection significantly increased the TGF-β1 level and decreased the levels of IL-17, IL-18, and IL-4. Proteomics and KEGG analysis revealed that DEGs associated with anti-ALI treatment by Xuebijing Injection were enriched in signaling pathways such as efferocytosis, apoptosis, and inflammatory response. The TUNEL staining results showed that Xuebijing Injection could significantly reduce the apoptosis rate of cells in lung tissue. qPCR, Western blot, and immunofluorescence showed that Xuebijing Injection could promote the expression of key factors ProS1, MerTK, and Rac1 involved in efferocytosis. Xuebijing Injection significantly alleviates inflammatory infiltration and injury in the lung tissue of ALI rats by enhancing efferocytosis in alveolar macrophages and promoting apoptotic cell clearance to suppress inflammation.