This article was aimed to study the preparation process of glycyrrhetinic acid (GA)-tanshinone IIA (TSN)-salvianolic acid B (SalB) compound liposomes with 3-succinic-30-stearyl glycyrrhetinic acid (18-GA-Suc) which is one of amphiphilicglycyrrhetinic acid derivatives as targeting molecule. The structure of the targeting molecule was validated by 1H-NMR and 13C-NMR methods. The feed ratio of 18-GA-Suc was optimized through single factor test and the incorporation ratio of 18-GA-Suc was determined by low-speed centrifugation. Meanwhile, physicochemi-cal properties between Suc-GTS-Lip and GTS-Lip were compared. In vitro release studies of three components in Suc-GTS-Lip were conducted by equilibrium dialysis method. The results showed that the optimum conditions were when the feed ratio of 18-GA-Suc was 10%lipid liposomal membrane (mol·mol-1). It revealed that the incorpora-tion ratio of 18-GA-Suc was 96.58%, and the encapsulation efficiencies of GA, TSN, and SalB were about 86.15%, 81.70%, and 91.05%, respectively. In addition, the Suc-GTS-Lip was spherical and uniformly dispersed with parti-cle size of 128.7 nm and zeta potential of-15.5 mV. The release model of GA and TSN was fitted well with Higuchi equation, while SalB was fitted well with Hixon-crowell equation. It was concluded that Glycyrrhetinic acid deriva-tives (18-GA-Suc) can be successfully expressed in the liposome membrane, and the optimal preparation method of Suc-GTS-Lip was stable. All three components encapsulated into liposomes had sustained-release effects, which laid a good foundation for its further study about liver-targeting.

ObjectiveTo investigate the therapeutic effect of Dayuanyin on acute lung injury induced by H1N1 infection and decipher the potential mechanism. MethodThe constituents in Dayuanyin were analyzed by ultra-high performance liquid chromatography-quadrupole-exactive orbitrap mass spectrometry （UHPLC-Q-Exactive Orbitrap MS）. Forty-eight female BALB/c mice were randomized into normal， model， oseltamivir （19.5 mg·kg^-1）， and low-， medium-， and high-dose （2.73， 5.46， 10.92 g·kg^-1） Dayuanyin groups. The normal and model groups were administrated with deionized water by gavage， and the other groups were administrated with the corresponding drugs by gavage. On day 3 of drug administration， the normal group received nasal inhalation of normal saline， and the other groups were inoculated intranasally with A/RP/8/34 （H1N1） for the modeling of influenza virus infection. Mice were administrated with drugs continuously for 7 days and weighed daily. Sampling was performed 12 h after the last administration， and the lung tissue was weighed to calculate the lung index. Hematoxylin-eosin staining was performed to observe the pathological and morphological changes of the lung tissue and bronchi. The cytometric bead array （CBA） was used to measure the serum levels of interferon-gamma （IFN-γ）， C-X-C motif ligand 1 （CXCL1）， tumor necrosis factor-alpha （TNF-α）， chemokine ligand 2 （CCL2）， interleukin-12p70 （IL-12p70）， chemokine ligand 5 （CCL5）， interleukin-1β （IL-1β）， chemokine （C-X-C motif） ligand 10 （CXCL10）， granulocyte-macrophage colony-stimulating factor （GM-CSF）， interleukin-10 （IL-10）， interferon-beta （IFN-β）， interferon-alpha （IFN-α）， and interleukin-6 （IL-6）. According to the results of mass spectrometry and network pharmacology， we analyzed the mechanism of Dayuanyin in treating acute lung injury caused by H1N1. The protein levels of extracellular signal-regulated kinase 1/2 （ERK1/2）， p38 mitogen-activated protein kinase （p38 MAPK）， nuclear factor-kappa B （NF-κB）， and their phosphorylated forms were determined by Western blot. The mRNA levels of myeloid differentiation factor 88 （MyD88）， Toll-like receptor 3 （TLR3）， Toll-like receptor 7 （TLR7）， and Toll-like receptor 8 （TLR8） in the lung tissue were measured by Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）. ResultA total of 57 compounds， including paeoniflorin and baicalein， were detected in Dayuanyin. Compared with the normal group， the model group showed decreased body weight （P<0.01）， lung edema and hemorrhage， increased lung index （P<0.01）， and elevated levels of IFN-γ， IL-12p70， CCL5， IL-1β， CXCL10， GM-CSF， IFN-β， and IL-6 （P<0.01）. Compared with the model group， Dayuanyin attenuated alveolar wall thickening， capillary congestion， and immune cell infiltration， reduced the alterations in body weight and lung index （P<0.01）， and down-regulated the protein levels of IFN-γ， IL-12p70， CCL5， IL-1β， CXCL10， GM-CSF， IFN-β， and IL-6 （P<0.01）. A total of 57 key genes were predicted by network pharmacological analysis， of which the MAPK signaling pathway was the main target signaling pathway. Compared with the normal group， the model group showed up-regulation in the protein levels of phosphorylation （p）-ERK1/2， p-p38 MAPK， and p-NF-κB （P<0.01） and the mRNA levels of TLR7， TLR8， MyD88， and TLR3 （P<0.05， P<0.01）. Compared with the model group， Dayuanyin lowered the phosphorylation levels of ERK1/2， p38 MAPK， and NF-κB p65 in a dose-dependent manner （P<0.01） and down-regulated the mRNA levels of TLR3， TLR7， TLR8， and MyD88 （P<0.01）. ConclusionDayuanyin can prevent and control H1N1 infection-induced acute lung injury by inhibiting the TLR/MAPK/NF-κB signaling pathway.