ObjectiveBased on traditional quality evaluation of Gardeniae Fructus（GF） recorded in historical materia medica， this study systematically compared the quality differences between wild and cultivated GF from morphological characteristics， microscopic features， and contents of primary and secondary metabolites. MethodsVernier calipers and analytical balances were used to measure the length， diameter and individual fruit weight of wild and cultivated GF， and the aspect ratio was calculated. A colorimeter was used to determine the chromaticity value of wild and cultivated GF， and the paraffin sections of them were prepared by safranin-fast green staining and examined under an optical microscope to observe their microstructure. Subsequently， the contents of water-soluble and alcohol-soluble extracts of wild and cultivated GF were detected by hot immersion method under the general rule 2201 in volume Ⅳ of the 2020 edition of the Pharmacopoeia of the People's Republic of China， the starch content was measured by anthrone colorimetric method， the content of total polysaccharides was determined by phenol-sulfuric acid colorimetric method， the sucrose content was determined by high performance liquid chromatography coupled with evaporative light scattering detection（HPLC-ELSD）， and the contents of representative components in them were measured by ultra-performance liquid chromatography（UPLC）. Finally， correlation analysis was conducted between quality traits and phenotypic traits， combined with multivariate statistical analysis methods such as principal component analysis（PCA） and orthogonal partial least squares-discriminant analysis（OPLS-DA）， key differential components between wild and cultivated GF were screened. ResultsIn terms of traits， the wild GF fruits were smaller， exhibiting reddish yellow or brownish red hues with significant variation between batches. While the cultivated GF fruits are larger， displaying deeper orange-red or brownish red. The diameter and individual fruit weight of cultivated GF were significantly greater than those of wild GF， while the blue-yellow value（b^*） of wild GF was significantly higher than that of cultivated GF. In the microstructure， the mesocarp of wild GF contained numerous scattered calcium oxalate cluster crystals， while the endocarp contained stone cell class round， polygonal or tangential prolongation， undeveloped seeds were visible within the fruit. In contrast， the mesocarp of cultivated GF contained few calcium oxalate cluster crystals， or some batches exhibited extremely numerous cluster crystals. The stone cells in the endocarp were predominantly round-like， with the innermost layer arranged in a grid pattern. Seeds were basically mature， and only a few immature seeds existed in some batches. Regarding primary metabolite content， wild GF exhibited significantly higher total polysaccharide level than cultivated GF（P<0.01）. In category-specific component content， wild GF exhibited significantly higher levels of total flavonoids and total polyphenols compared to cultivated GF（P<0.01）. Analysis of 12 secondary metabolites revealed that wild GF exhibited significantly higher levels of Shanzhiside， deacetyl asperulosidic acid methyl ester， gardenoside and chlorogenic acid compared to cultivated GF（P<0.01）. Conversely， the contents of genipin 1-gentiobioside， geniposide and genipin were significantly lower in wild GF（P<0.01）. ConclusionThere are significant differences between wild and cultivated GF in terms of traits， microstructure， and contents of primary and secondary metabolites. At present， the quality evaluation system of cultivated GF remains incomplete， and this study provides a reference for guiding the production of high-quality GF medicinal materials.

Aim To investigate the therapeutic effects of cinnamaldehyde on synovial lesions in mice with knee osteoarthritis(KOA)and its regulatory mecha-nism in the phagocytic function of synovial macropha-ges.Methods In the animal experiments,mouse ser-um and synovial tissue were extracted.HE staining was used to evaluate the inflammatory cell infiltration in the synovial tissue.ELISA was employed to detect the lev-els of inflammatory factors such as interleukins in the serum.Western blot was used to detect the expression of Ras homolog family member A(RhoA),Rho-associ-ated protein kinase 1(ROCK1),myosin light chain(MLC),and p-MLC proteins in the synovial tissue.RT-qPCR was utilized to detect the expression of in-flammatory factors and pathway-related mRNA in the synovial tissue.TUNEL staining was used to detect ap-optosis in the synovial tissue.In the cellular experi-ments,after the intervention,RAW267.4 cells were subjected to Western blot and RT-qPCR for the detec-tion of the aforementioned indicators,and confocal mi-croscopy was used to assess phagocytic function.Re-sults After cinnamaldehyde intervention,the synovial inflammatory infiltration was significantly reduced,the protein and mRNA expression of the RhoA/ROCK1/MLC signaling pathway was markedly downregulated,the fluorescence intensity of TUNEL staining signifi-cantly decreased,and the phagocytic function of macro-phages was enhanced.Conclusion Cinnamaldehyde can inhibit RhoA/Rock1/MLC signaling pathway,en-hance macrophage burial,improve synovial inflamma-tion,and delay the progression of KO A mice.

With the completion of the "Human Genome Project" and the smooth progress of the "Herbal Genome Project", the research wave of RNAomics is gradually advancing, opening the research gateway for the modernization of traditional Chinese medicine (TCM) and initiating the post-genome era of medicinal plant RNA research. Therefore, this article proposes for the first time the concept of HerbRNomes, which involves constructing databases of medicinal plant, medicinal fungus, and medicinal animal RNA at different stages, from different origins, and in different organs. This research aims to explore the role of HerbRNA in self-genetic information transmission, functional regulation, as well as cross-species regulation functional mechanisms and key technologies. It also investigates application scenarios, providing a theoretical basis and research ideas for the resistance of TCM or medicinal plants to adversity and stress, molecular assistant breeding, and the development of small nucleic acid drugs. This article reviews recent research progress in elucidating the molecular mechanisms of the transmission and expression of genetic information, self-regulation and cross-species regulation of herbs at the RNA level, along with key technologies. It proposes a development strategy for small nucleic acid drugs based on HerbRNomes, providing theoretical support and guidance for the modernization of TCM based on HerbRNomes research.

Aim To investigate the therapeutic effects of cinnamaldehyde on synovial lesions in mice with knee osteoarthritis(KOA)and its regulatory mecha-nism in the phagocytic function of synovial macropha-ges.Methods In the animal experiments,mouse ser-um and synovial tissue were extracted.HE staining was used to evaluate the inflammatory cell infiltration in the synovial tissue.ELISA was employed to detect the lev-els of inflammatory factors such as interleukins in the serum.Western blot was used to detect the expression of Ras homolog family member A(RhoA),Rho-associ-ated protein kinase 1(ROCK1),myosin light chain(MLC),and p-MLC proteins in the synovial tissue.RT-qPCR was utilized to detect the expression of in-flammatory factors and pathway-related mRNA in the synovial tissue.TUNEL staining was used to detect ap-optosis in the synovial tissue.In the cellular experi-ments,after the intervention,RAW267.4 cells were subjected to Western blot and RT-qPCR for the detec-tion of the aforementioned indicators,and confocal mi-croscopy was used to assess phagocytic function.Re-sults After cinnamaldehyde intervention,the synovial inflammatory infiltration was significantly reduced,the protein and mRNA expression of the RhoA/ROCK1/MLC signaling pathway was markedly downregulated,the fluorescence intensity of TUNEL staining signifi-cantly decreased,and the phagocytic function of macro-phages was enhanced.Conclusion Cinnamaldehyde can inhibit RhoA/Rock1/MLC signaling pathway,en-hance macrophage burial,improve synovial inflamma-tion,and delay the progression of KO A mice.

A strategy combining a tailored database and high-throughput activity screening that discover bioactive metabolites derived from Magnoliae Officinalis Cortex(MOC)was developed and implemented to rapidly profile and discover bioactive metabolites in vivo derived from traditional Chinese medicine(TCM).The strategy possessed four characteristics:1)The tailored database consisted of metabolites derived from big data-originated reference compound,metabolites predicted in silico,and MOC chemical profile-based pseudomolecular ions.2)When profiling MOC-derived metabolites in vivo,attentions were paid not only to prototypes of MOC compounds and metabolites directly derived from MOC compounds,as reported by most papers,but also to isomerized metabolites and the degradation products of MOC compounds as well as their derived metabolites.3)Metabolite traceability was performed,especially to distinguish isomeric prototypes-derived metabolites,prototypes of MOC compounds as well as phase Ⅰ metabolites derived from other MOC compounds.4)Molecular docking was utilized for high-throughput activity screening and molecular dynamic simulation as well as zebrafish model were used for verification.Using this strategy,134 metabolites were swiftly characterized after the oral administration of MOC to rats,and several metabolites were reported for the first time.Furthermore,17 potential active metabolites were discovered by targeting the motilin,dopamine D2,and the serotonin type 4(5-HT4)receptors,and part bioactivities were verified using molecular dynamic simulation and a zebrafish constipation model.This study extends the application of mass spectrometry(MS)to rapidly profile TCM-derived metabolites in vivo,which will help pharmacologists rapidly discover potent metabolites from a complex matrix.

A strategy combining a tailored database and high-throughput activity screening that discover bioactive metabolites derived from Magnoliae Officinalis Cortex (MOC) was developed and implemented to rapidly profile and discover bioactive metabolites in vivo derived from traditional Chinese medicine (TCM). The strategy possessed four characteristics: 1) The tailored database consisted of metabolites derived from big data-originated reference compound, metabolites predicted in silico, and MOC chemical profile-based pseudomolecular ions. 2) When profiling MOC-derived metabolites in vivo, attentions were paid not only to prototypes of MOC compounds and metabolites directly derived from MOC compounds, as reported by most papers, but also to isomerized metabolites and the degradation products of MOC compounds as well as their derived metabolites. 3) Metabolite traceability was performed, especially to distinguish isomeric prototypes-derived metabolites, prototypes of MOC compounds as well as phase I metabolites derived from other MOC compounds. 4) Molecular docking was utilized for high-throughput activity screening and molecular dynamic simulation as well as zebrafish model were used for verification. Using this strategy, 134 metabolites were swiftly characterized after the oral administration of MOC to rats, and several metabolites were reported for the first time. Furthermore, 17 potential active metabolites were discovered by targeting the motilin, dopamine D2, and the serotonin type 4 (5-HT₄) receptors, and part bioactivities were verified using molecular dynamic simulation and a zebrafish constipation model. This study extends the application of mass spectrometry (MS) to rapidly profile TCM-derived metabolites in vivo, which will help pharmacologists rapidly discover potent metabolites from a complex matrix.

Aim To verify the mechanism of action of Carthami Flos-Lycopodii Herba in treating KOA carti-lage inflammation based on network pharmacology and in vitro and in vivo experiments.Methods The effec-tive ingredients of Carthami Flos-Lycopodii Herba were screened through the database,the core targets of"drug disease"were analyzed,and pathway enrichment analy-sis and molecular docking verification were conducted.Experimental verification:Primary chondrocytes were extracted from mice and divided into the control group,IL-1β group,treatment group,and treatment+TLR4 agonist group.CCK-8 method was used to screen the optimal intervention concentration of Carthami Flos-Ly-copodii Herba.ELISA was used to detect the content of inflammatory factors in chondrocytes.Western blot was employed to detect the protein expression related to cellular pathways.Subsequently,a KOA mouse model was constructed using the DMM method.After admin-istration,the knee joint injury of mice was evaluated u-sing safranin O-green staining.ELISA was used to de-tect the levels of inflammatory factors in serum.West-ern blot was employed to detect collagen Ⅱ,MMP13,Aggrecan,and apoptosis related protein expression in cartilage tissue.TUNEL staining was used to detect the apoptosis rate of cells.Results A total of 26 active ingredients of Carthami Flos-Lycopodii Herba were screened,as well as 123 potential targets for treating KOA.The enrichment analysis results indicated that it mainly involved mechanisms such as Toll like receptors and cell apoptosis.The experimental results showed that Carthami Flos-Lycopodii Herba alleviated the in-flammatory response of chondrocytes and affected the expression of pathway related proteins.Compared with KOA mice,safflower stretched muscle grass could im-prove cartilage damage and reduce the concentration of serum inflammatory factors,regulate the expression of collagen Ⅱ,MMP13,Aggrecan,and apoptosis related proteins in cartilage tissue,and reduce the fluorescence intensity of TUNEL staining in the tissue.Conclusions Carthami Flos-Lycopodii Herba can improve KOA cartilage inflammation,and its mechanism may be relat-ed to the TLR4/MyD88/NF-κB signaling pathway.

Aim To verify the mechanism of action of Carthami Flos-Lycopodii Herba in treating KOA carti-lage inflammation based on network pharmacology and in vitro and in vivo experiments.Methods The effec-tive ingredients of Carthami Flos-Lycopodii Herba were screened through the database,the core targets of"drug disease"were analyzed,and pathway enrichment analy-sis and molecular docking verification were conducted.Experimental verification:Primary chondrocytes were extracted from mice and divided into the control group,IL-1β group,treatment group,and treatment+TLR4 agonist group.CCK-8 method was used to screen the optimal intervention concentration of Carthami Flos-Ly-copodii Herba.ELISA was used to detect the content of inflammatory factors in chondrocytes.Western blot was employed to detect the protein expression related to cellular pathways.Subsequently,a KOA mouse model was constructed using the DMM method.After admin-istration,the knee joint injury of mice was evaluated u-sing safranin O-green staining.ELISA was used to de-tect the levels of inflammatory factors in serum.West-ern blot was employed to detect collagen Ⅱ,MMP13,Aggrecan,and apoptosis related protein expression in cartilage tissue.TUNEL staining was used to detect the apoptosis rate of cells.Results A total of 26 active ingredients of Carthami Flos-Lycopodii Herba were screened,as well as 123 potential targets for treating KOA.The enrichment analysis results indicated that it mainly involved mechanisms such as Toll like receptors and cell apoptosis.The experimental results showed that Carthami Flos-Lycopodii Herba alleviated the in-flammatory response of chondrocytes and affected the expression of pathway related proteins.Compared with KOA mice,safflower stretched muscle grass could im-prove cartilage damage and reduce the concentration of serum inflammatory factors,regulate the expression of collagen Ⅱ,MMP13,Aggrecan,and apoptosis related proteins in cartilage tissue,and reduce the fluorescence intensity of TUNEL staining in the tissue.Conclusions Carthami Flos-Lycopodii Herba can improve KOA cartilage inflammation,and its mechanism may be relat-ed to the TLR4/MyD88/NF-κB signaling pathway.

Objective:To investigate the effects and mechanisms of glycyrrhetinic acid(GA)on the proliferation,migration,invasion,and epithelial-mesenchymal transition(EMT)of hepatocellular carcinoma SMMC-7721 cells by regulating the β-catenin/transcription factor 4(TCF4)signaling pathway.Methods:HCC cells SMMC-7721 were randomly separated into the control,L-GA,M-GA,H-GA groups(50,100,200 μmol/L GA)and the GA+LiCl group(200 μmol/L GA+40 μmol/L β-catenin activator LiCl).The effects of GA on the proliferation,migration,invasion and apoptosis of SMMC-7721 cells were detected by plate cloning,Transwell experiment and flow cytometry.WB was applied to detect the effects of GA on the expressions of β-catenin/TCF4 signaling pathway proteins(β-catenin、TCF4)and EMT(E-cadherin,vimentin)-related proteins in SMMC-7721 cells.The model of SMMC-7721 cell transplanted tumor in nude mice was established to observe the effects of GA on the growth of transplanted tumor and the expressions of β-catenin and TCF4 proteins.Results:Compared with the control group,SMMC-7721 cell clonal number,migration and invasion numbers,β-catenin,TCF4 and vmentin protein expressions in the L-GA,M-GA and H-GA groups decreased significantly(all P<0.05),while the apoptosis rate and the expression of E-cadherin protein increased(all P<0.05).Compared with the H-GA group,the numbers of clones,migration and invasion,β-catenin,TCF4 and vimentin protein expressions in the GA+LiCl group increased significantly(all P<0.05),while the apoptosis rate and E-cadherin protein expression decreased significantly(all P<0.05).Tumor-bearing nude mouse model experiment showed that the tumor weight and volume and the positive rates of β-catenin and TCF4 proteins in the GA group were significantly lower than those in the control group(all P<0.05).Conclusion:GA can significantly inhibit the proliferation,migration,invasion,and the EMT process of SMMC-7721 cells,thereby inhibiting the progression of HCC.Its mechanism may be achieved by inhibiting the β-catenin/TCF4 signaling pathway.

Nitazoxanide is an FDA-approved antiprotozoal drug. Our previous study found that oral administration of nitazoxanide inhibited Western diet (WD)-induced hepatic steatosis in ApoE^-/- mice. However, the specific mechanism remains to be elucidated. In the present study, we performed an untargeted metabolomics approach to reveal the effect of nitazoxanide on the liver metabolic profiles in WD-fed ApoE^-/- mice, and carried out the cellular experiments to elucidate the underlying mechanisms. UPLC-MS-based untargeted metabolomics analysis was used to investigate the effect of nitazoxanide on global metabolite changes in liver tissues. The differential metabolites were screened for enrichment analysis and pathway analysis. Hepatocytes were treated with tizoxanide, the metabolite of nitazoxanide, to investigate the underlying mechanism based on the findings in metabolomics study. The improvement of liver lipid metabolism disorders by nitazoxanide treatment in WD-fed ApoE^-/- mice was mainly through regulating glycerophospholipid metabolism, D-glutamine and glutamate metabolism, glutathione metabolism, and arginine biosynthesis metabolism. Tizoxanide, the active metabolite of nitazoxanide, increased glutathione (GSH) contents and glutamate-cysteine ligase catalytic subunit (Gcl-c) and glutathione reductase (Gsr) mRNA expressions in HepG2 cells. Tizoxanide increased cystathionine β-synthase (CBS) and phosphatidylethanolamine N-methyltransferase (PEMT) protein levels, inhibited lipid accumulation in hepatocytes induced by free fatty acid (FFA). Tizoxanide increased S-adenosyl-L-homocysteine hydrolase (SAHH) protein levels in HepG2 cells and mouse primary liver cells stimulated with free fatty acid (FFA). Tizoxanide increased N-acetyl glutamate synthase (Nags) and carbamoylphosphate synthetase 1 (Cps1) mRNA expressions in HepG2 cells. In conclusion, nitazoxanide improves WD-induced hepatic steatosis in ApoE^-/- mice and the underlying mechanisms include increasing CBS expression, GSH content, PEMT protein expression, Nags and Cps1 mRNA expression in hepatocytes.