Luteolin targets the AGE-RAGE signaling to mitigate inflammation and ferroptosis in chronic atrophic gastritis

Abstract

Chronic atrophic gastritis (CAG) is a chronic inflammatory disease and precancerous lesion in stomach cancer. Abnormal activation cellular ferroptosis further damages gastric tissue, which is susceptible to inflammation. Luteolin has powerful anti-inflammatory and regulatory potential for cellular ferroptosis. We aimed to clarify the involvement of luteolin in inflammation and ferroptosis during CAG. Luteolin targets were searched to identify intersecting genes in the chronic atrophic gastritis disease database. The AGE-RAGE pathway is a potential target of luteolin for the treatment of chronic atrophic gastritis and a binding site between luteolin and RAGE was predicted through a computer simulation of molecular docking. We established a CAG rat model using N-methyl-N-nitro-N-nitroguanidine. The therapeutic effect of luteolin on CAG was detected using western blotting, qPCR, hematoxylin and eosin staining, lipid oxidation (MDA), and Fe²⁺ assays. Luteolin inhibited the AGE-RAGE signaling pathway and reduced the inflammatory response in gastric tissues. Additionally, luteolin downregulated the concentration of (MDA) and Fe²⁺, and CAG downregulated the expression levels of ACSL4 and NOX1 and upregulated the expression levels of FIH1 and GPX4 ferroptosis-related proteins, thus inhibiting the ferroptosis of gastric tissue cells, which had a therapeutic effect on CAG.

Keywords: AGE-RAGE signaling pathway; chronic atrophic gastritis; ferroptosis; luteolin.

Figure 1

Screening of potential therapeutic targets for luteolin. (A) Drug targets and CAG differential genes were intersected to draw a Wayne diagram. (B) Heat map of intersecting gene clustering. (C) GO functional enrichment analysis. (D) KEGG functional enrichment analysis of intersecting genes. (E) Intersecting gene plot of the PPI network diagrams. CAG, chronic atrophic gastritis; GO, gene ontology; KEGG, Kyoto Encyclopedia of Genes and Genomes; PPI, protein-protein interaction.

Figure 2

Luteolin has a binding site with AGRE. (A) 2D and 3D molecular structures and relative molecular mass of luteolin. (B) Schematic of luteolin docking with AGRE.

Figure 3

Luteolin has a therapeutic effect on rat CAG in vivo. (A) CAG animal modeling and treatment flowchart. (B) Top view of the rat gastric tissue anatomy. (C) Body weight curves of the rats. (D) HE staining of rat gastric tissues. CAG, chronic atrophic gastritis; HE, hematoxylin and eosin.

Figure 4

Luteolin can reduce the inflammatory response in the gastric tissue of CAG rats. (A) Expression of IL-1 β and TNF-α in the CAG dataset. (B) Changes in Il-6, Il-10, Tnf-α, and Il-1β levels in rat serum were determined by ELISA experiments. (C) The level of mRNA changes in Il-6, Il-10, TNF-a, and Il-1b after luteolin treatment was determined by qPCR.

Figure 5

Luteolin inhibits ferroptosis in rat gastric tissue. (A) Changes in the levels of Fe²⁺ and MDA in gastric tissues of luteolin-treated rats. (B) Changes in the mRNA expression of FIH1, COX2, ACSL4, GPX4, and NOX1 were detected using qPCR. (C) Western blot analysis of the protein expression levels of FIH1, COX2, ACSL4, GPX4, and NOX1.

Figure 6

Luteolin inhibits the activity of the AGE-RAGE signaling pathway. (A) RAGE expression in the CAG disease database. (B) Western blot. The protein expression levels of AGE and RAGE were determined. (C) Western blot assay was used to detect the protein expression levels of NF-κB p65 and p-NF-κB p65.