Baicalein suppresses inflammation and attenuates acute lung injury by inhibiting glycolysis via HIF‑1α signaling

Abstract

Baicalein, a flavonoid monomer compound isolated from the dried root of the traditional Chinese herb Scutellaria baicalensis, has several pharmacological activities, such as anti‑inflammatory, anti‑angiogenic, antitumor, antimicrobial and antiviral properties. Acute lung injury (ALI) is characterized by injury of the alveolar epithelium and capillary endothelium, which results in decreased lung volume, decreased lung compliance, ventilation/perfusion mismatch, intrapulmonary edema, alveolar edema and even acute hypoxemic respiratory failure. The present study aimed to investigate the effects of baicalein on lung injury and inflammation. Bioinformatics analysis using network pharmacology predicted that the hypoxia inducible factor‑1α (HIF‑1α) and glycolysis signaling pathways were involved in the mechanism underlying the therapeutic effects of baicalein. Further in vitro and in vivo experiments, such as immunohistochemistry, immunofluorescence and PCR, verified that baicalein could inhibit HIF‑1α signaling, thus suppressing glycolysis, and improving inflammatory responses and ALI. Taken together, the results of the present study suggested that the anti‑inflammatory effects of baicalein on treating ALI were associated with its ability to suppress glycolysis via the HIF‑1α signaling pathway.

Keywords: HIF‑1α signaling; acute lung injury; baicalein; glycolysis; inflammation.

Figure 1.

Baicalein ameliorates lung pathological damage in mice with lipopolysaccharide-induced acute lung injury. (A) 2D and 3D structure of baicalein. (B) H&E staining images of lung tissues (magnification, ×200). The arrows represent inflammatory cell infiltration and thickening of the alveolar wall. (C) Inflammatory (alveolitis) score of the H&E staining images. (D) Lung wet/dry weight ratio of mice. (E) Number of total cells and total protein in the BALF of mice. Data are presented as the mean ± SEM, n=6. *P<0.05, **P<0.01. BALF, bronchoalveolar lavage fluid; DEX, dexamethasone; H, high; H&E, hematoxylin and eosin; L, low.

Figure 2.

Network analysis of the baicalein-related targets and ALI-related genes. (A) Venn diagram showing 248 overlapping targets between baicalein-related targets and ALI-related genes. (B) Protein-protein interaction network of the top 100 overlapping targets. (C) KEGG pathway enrichment analysis of the overlapping targets. (D) GO analysis of the overlapping targets. GO terms and KEGG pathways with P<0.05 were considered significantly enriched. ALI, acute lung injury; BP, biological process; CC, cellular component; GO, Gene Ontology; KEGG, Kyoto Encyclopedia of Genes and Genomes; MF, molecular function.

Figure 3.

Baicalein inhibits the inflammatory response in LPS-induced macrophages and in mice with LPS-induced acute lung injury. Levels of IL-6 and TNF-α in the (A) serum and (B) BALF of mice. (C) Effects of baicalein at various concentrations on MH-S cell viability. Levels of IL-6 and TNF-α were assessed using (D) enzyme-linked immunosorbent assay and (E) quantitative PCR in MH-S cells. Data are presented as the mean ± SEM. **P<0.01. BALF, bronchoalveolar lavage fluid; DEX, dexamethasone; H, high; L, low; LPS, lipopolysaccharide; TNF-α, tumor necrosis factor-α.

Figure 4.

Baicalein inhibits glycolysis in LPS-induced macrophages and in the lung tissues of mice with LPS-induced acute lung injury. Immunohistochemical staining images of (A) HK2, (B) PFK1 and (C) PKM2 (magnification, ×200) and (D) IOD values in the lungs. (E) Lactate content in the lung tissues. (F) mRNA expression levels of HK2, PFK1 and PKM2 in macrophages. (G) Lactate content in macrophages. Data are presented as the mean ± SEM. *P<0.05, **P<0.01. DEX, dexamethasone; H, high; HK2, hexokinase 2; IOD, integrated optical density; L, low; LPS, lipopolysaccharide; PFK1, phosphofructokinase-1; PKM2, pyruvate kinase M2.

Figure 5.

Baicalein suppresses HIF-1α signaling in LPS-induced macrophages and in mice with LPS-induced ALI. (A) Immunofluorescence staining images (magnification, ×200) of HIF-1α in lung tissues. (B) Immunohistochemical staining images (magnification, ×200) and (C) IOD values of HIF-1α in the lungs. (D) mRNA expression levels of HIF-1α in LPS-induced macrophages. (E) Molecular docking of baicalein and HIF-1α. Data are presented as the mean ± SEM. *P<0.05, **P<0.01. DEX, dexamethasone; H, high; HIF-1α, hypoxia inducible factor-1α; IOD, integrated optical density; L, low; LPS, lipopolysaccharide.

Figure 6.

Diagram of the molecular mechanism underlying the therapeutic effects of baicalein on ALI. This picture was drawn by Figdraw (https://www.figdraw.com/; ID: PIPRId8c78). ALI, acute lung injury; HIF-1α, hypoxia inducible factor-1α; HK2, hexokinase 2; PFK1, phosphofructokinase-1; TNF-α, tumor necrosis factor-α.