Puerarin has been reported as a potential agent for neuro-inflammatory disorders. However, there have been no reports of using puerarin for the treatment of depression based on Toll-like receptor 4 (TLR4)–mediated inflammatory injury. In this study, we evaluated the protective effects of puerarin on depression-like rats induced by a high-fat diet (HFD) combined with chronic unpredictable mild stress (CUMS). The mechanism was screened by lipidomics and molecular docking and confirmed by in vivo tests. Puerarin treatment significantly improved 1% sucrose preference and ameliorated depression-like behavior in the open-field test. The antidepressive effects of puerarin were associated with decreased pro-inflammatory cytokine production, including interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α), and increased anti-inflammatory cytokine levels (IL-10) in rat hippocampal tissues and plasma. Hematoxylin–eosin (H&E), immunofluorescence staining, and Western blotting results displayed that puerarin alleviated inflammatory injury by suppressing TLR4 expression and by repairing the intestine mucus barrier via enhancing the expression of claudin-1 and occludin. Non-targeted lipidomics analysis showed that the most significantly different metabolites modified by puerarin were phospholipids. Puerarin treatment–altered biomarkers were identified as PC (15:1/20:1), PE (15:1/16:1), and PI (18:2/20:1) in comparison with the HFD/CUMS group. Molecular docking modeling revealed that puerarin could bind with cytosolic phospholipase A2 (cPLA2) and cyclooxygenase-2 (COX-2), which play central roles in TLR4-mediated phospholipid metabolism. In vivo, puerarin treatment decreased the enzyme activities of cPLA2 and COX-2, resulting in lower production of prostaglandin E₂ (PGE₂) in hippocampal and intestinal tissues. In conclusion, puerarin treatment reverses HFD/CUMS-induced depression-like behavior by inhibiting TLR4-mediated intestine mucus barrier dysfunction and neuro-inflammatory damages via the TLR4/cPLA2/COX-2 pathway.

Evodiamine has gained wide interests recently because of its antitumor activities. However, a superior bioavailability is required to achieve better efficacy due to its poor water solubility. The aim of this study was to enhance the evodiamine’s aqueous solubility by preparing evodiamine/hydroxypropyl-β-cyclodextrin (EVO/HP-β-CD) inclusion complex, which is incorporated evodiamine into HP-β-CD, and compare the antitumor activities before and after inclusion with HP-β-CD in human hepatoma HepG2 cells. The EVO/HP-β-CD inclusion complexes were prepared by the kneading method and structurally characterized. P-glycoprotein ATPase assays firstly demonstrated that evodiamine was a substrate of P-glycoprotein, while HP-β-CD and EVO/HP-β-CD inclusion complexes inhibited P-glycoprotein by blocking P-glycoprotein ATPase activity. The EVO/HP-β-CD inclusion complexes may be a promising anticancer drug candidate without drug resistance. After given evodiamine or EVO/HP-β-CD inclusion complexes intervention, cell viability evaluation indicated that the half inhibition concentration of evodiamine and EVO/HP-β-CD inclusion complexes on HepG2 cells was 8.516 and 0.977 μM, respectively. The caspase-3 enzyme activity analysis and Annexin V/PI double-staining revealed that EVO/HP-β-CD inclusion complexes possessed better antitumor activities than evodiamine. Additionally, Hoechst 33258 staining and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling assay demonstrated that EVO/HP-β-CD inclusion complexes induced HepG2 cell apoptosis more effectively than evodiamine. The improved antitumor activities of evodiamine were attributed to the enhanced solubility and P-glycoprotein inhibition by HP-β-CD. These results are promising for the drug administration of EVO/HP-β-CD inclusion complexes to enhance the bioavailability of evodiamine in vivo.

Although, Dan-Lou prescription (DLP) is used for antagonizing check discomfort and heartache, the pharmacological mechanism has not been clearly illustrated. Our present study aimed to design inflammatory models induced by LPS in vivo and in vitro to investigate the anti-inflammation of DLP ethanol extract (EEDL) and the potential mechanisms. EEDL was prepared and then analyzed by high performance liquid chromatography (HPLC). Further, the anti-inflammatory effects of EEDL in vivo was evaluated by measuring inflammation-associated factors includingcytokines, chemokines and acute phase proteins in lipopolysaccharide (LPS)-induced mice serum and liver. The anti-inflammatory mechanism exploration of EEDL was performed in LPS-stimulated RAW 264.7 cells. Different effects of EEDL on nitric oxide (NO) and prostaglandin (PG)E2 secretion were investigated by Griess reagent method and enzyme-linked immunosorbent assay (ELISA) respectively. Then the mRNA and protein expression of inducible NO synthase (iNOS) and cyclooxygenase (COX)-2 were measured by real-time reverse-transcription polymerase chain reaction (RT-PCR), ELISA and Western blot. Other chemokines and acute phase proteins were determined by proteome profile array. Finally, the ELISA based transcription factor assay was applied to measure the DNA-binding activity of nuclear transcription factor (NF)-κB p65. Eight compounds from EEDL have been identified as gallic acid, salvianic acid, puerarin, daidzin, paeoniflorin, salvianolic acid B, cryptotanshinone, and tanshinone IIA, with amounts of 0.26, 9.84, 10.41, 2.55, 9.44, 3.82, 0.24 and 0.3 mg/kg, respectively. In vivo, EEDL administration antagonized the up-regulation of more than 17 kinds of cytokines, chemokines and acute phase proteins in LPS-treated mice serum, and inhibited LPS-induced IL-6 mRNA and protein expression in mice liver tissue. In vitro, LPS-induced NO and PGE2 over-productions were decreased by EEDL treatment. The mRNA and protein expression of iNOS, COX-2 and IL-6 were similarly inhibited by EEDL treatment, which might be attributed to decrease the DNA-binding activity of NF-κB p65. EEDL was valid for anti-inflammation and the potential molecular mechanisms might be due to the inhibition of of LPS-induced iNOS/NO, COX-2/PGE2 and cytokines expression by antagonizing the activation of NF-κB p65.