To explore the mechanisms of action of fructus broussonetiae regulating phosphoinositide 3-kinase/ protein kinase B (PI3K/Akt) in diabetic kidney disease (DKD).

The traditional Chinese medicine systems pharmacology database and analysis platform (TCMSP), the human gene database (GeneCards), online mendelian inheritance in man (OMIM) and therapeutic target database (TTD) were used to screen the active ingredients and targets. The protein-protein interaction (PPI) network and topology analysis were constructed and enriched using gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG), and molecular docking and molecular dynamics (MD) simulations were used to verify the interactions between the core ingredients and targets. Mouse podocyte cells 5 were divided into control group (without any treatment), model group (treated with 30 mmol·L^-1 glucose for 48 h), mannitol group (treated with 44.5 mmol·L^-1 mannitol for 48 h on the basis of the control group), experimental group (treated with 2.5, 5.0, 15.0, 30.0 and 45.0 μmol·L^-1 luteolin for 24 h, on the basis of the model group) and positive drug group (treated with 100 μmol·L^-1 captopril for 24 h on the basis of the model group). Cell viability were detected by cell counting kit-8 (CCK-8), apoptosis were detected by flow cytometry, levels of inflammatory factors interleukin-1β (IL-1β), IL-6 and tumour necrosis factor (TNF-a) were measured by enzyme-linked immunosorbent assay (ELISA), and B-cell lymphoma-2 gene (Bcl-2), Bcl-2-related X protein (Bax), cysteine aspartate specific proteinase-3 (Caspase-3), heat shock protein 90-α (HSP90AA1), phosphorylated protein kinase B (p-Akt) and Akt protein expression levels were detected by Weatern blot.

A total of 42 intersection targets of fructus broussonetiae and DKD were identified, and 13 core targets were screened out, including protein kinase B1 (Akt1), prostaglandin-endoperoxide synthase 2 (PTGS2), tumour protein 53 (TP53) and HSP90AA1, etc. The GO/KEGG enrichment analysis showed that the mechanism of action was involved in the regulation of PI3K/Akt signalling pathway as well as the oxidative stress, apoptosis, etc. The molecular docking and kinetic simulations confirmed that HSP90AA1 formed the most stable complex with luteolin (binding free energy -34.08 kcal·mol^-1), therefore, luteolin were selected for subsequent cellular experiments. CCK-8 showed that the effect of luteolin entered into a plateau phase when the concentration reached 30.0 μmol·L^-1, and this concentration was used for the subsequent experiments. The total apoptosis rates of the control group, model group, mannitol group, experimental group (30.0 μmol·L^-1) and positive drug group were (6.23±0.40)%, (35.47±3.08)%, (6.42±0.52)%, (11.20±1.46)% and (10.78±1.06)%, respectively; the IL-6 levels of the cells in the control, model and experimental groups (30.0 μmol·L^-1) were (29.52±2.21), (91.12±6.14) and (36.48±3.65) pg·mL^-1, respectively; IL-1β levels were (12.68±1.17), (47.42±3.49) and (18.08±1.42) pg·mL^-1, respectively; and TNF-α levels were (23.15±1.81), (79.35±7.01) and (29.62±2.17) pg·mL^-1, respectively; the relative expression levels of Bcl-2 protein were 1.02±0.11, 0.45±0.04 and 0.92±0.07, respectively; the relative expression levels of Bax protein were 1.01±0.09, 2.18±0.17 and 1.12±0.11, respectively; the relative expression levels of Caspase-3 protein relative expression levels were 1.03±0.08, 2.32±0.17 and 1.31±0.14, respectively. The relative expression levels of HSP90AA1 protein were 1.00±0.07, 1.96±0.12 and 1.08±0.10, respectively; the relative expression levels of p-Akt/Akt were 1.00±0.07, 2.25±0.07 and 1.11±0.06, respectively. The above indexes in the model group were statistically significant when compared with those in the control group, and when compared with those in the experimental and model groups (P<0.001).

The active component luteolin of fructus broussonetiae may reduce the expression of HSP90AA1, inhibit the activation of PI3K/Akt signaling pathway, and reduce the inflammatory response, thus achieving the purpose of treating DKD.