ObjectiveTo explore the mechanism of Yishen Qubi Tongluo Formula （益肾祛痹通络方， YQTF） in the treatment of rheumatoid arthritis（RA）. MethodsNetwork pharmacology was employed to retrieve and screen the active components and potential targets of YQTF as well as RA-related targets using databases including TCMSP， BATMAN， ETCM and GEO. The intersection of targets related to active components and RA-related targets was identified， and a protein-protein interaction （PPI） network was constructed. Gene Ontology （GO） functional enrichment analysis and Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment analysis were performed， and a drug-active component-common target network of YQTF in the treatment of RA was established. The core components of YQTF were molecularly docked with key targets. Human rheumatoid arthritis synovial fibroblast cell line MH7A was divided into blank group， model group， methotrexate group and YQTF group. The blank group was cultured with 10% fetal bovine serum， while the other three groups were stimulated with 10 μg/L of recombinant human tumor necrosis factor-α （TNF-α） for 24 h to establish the RA cell model. On this basis， the methotrexate group was treated with methotrexate suspension at a concentration of 20 μmol/L， and the YQTF group was treated with 10% YQTF-medicated serum. After 48 h of intervention， the levels of TNF-α and interleukin-17A（IL-17A）contents in cell supernatants were detected by enzyme-linked immunosorbent assay （ELISA）， and mRNA expressions of phosphatidylinositol 3-kinase（PI3K）， protein kinase B（AKT） and mammalian target of rapamycin（mTOR） were detected by real-time quantitative polymerase chain reaction （RT-qPCR）. ResultsNetwork pharmacological analysis identified 209 active components and 583 potential target genes of YQTF， as well as 818 RA-related targets. A total of 29 common targets were obtained from the intersection of drug-related targets and RA-related targets. Quercetin，β-sitosterol， kaempferol， stigmasterol and luteolin were the core active components of YQTF for the treatment of RA， while matrix metalloproteinase-9 （MMP9）， prostaglandin-endoperoxide synthase 2 （PTGS2）， Toll-like receptor 4 （TLR4）， tumor protein p53 （TP53） and transcription factor AP-1 subunit JUN were the key targets. The GO and KEGG pathway enrichment analysis showed that the involved biological processes and pathways were mainly associated with antioxidant responses， PI3K-AKT signaling pathway and Toll-like receptor （TLR） signaling pathway. Molecular docking results showed that MMP9 and PTGS2 exhibited high binding affinities with quercetin， β-sitosterol， kaempferol， stigmasterol and luteolin； TLR4 exhibited high binding activities with β-sitosterol， stigmasterol and luteolin； and TP53 showed high binding affinity with luteolin. The results of cell experiments showed that compared with the control group， the contents of TNF-α and IL-17A as well as the mRNA expressions of AKT and mTOR in the model group significantly increased （P＜0.05 or P＜0.01）. Compared with the model group， all the above indicators significantly decreased in the YQTF group， while the contents of TNF-α and the mRNA expression of AKT significantly decreased in the methotrexate group （P＜0.05 or P＜0.01）. ConclusionThe mechanism of YQTF in the treatment of RA may be associated with reducing inflammatory cytokine secretion and inhibiting the activation of the PI3K-AKT-mTOR signaling pathway.

The medicinal history of Zanthoxyli Radix is long,and it is recorded in the Shen Nong Ben Cao Jing under the name of Manjiao.Modern pharmacological research has proven that Zanthoxyli Radix has anti-tumor,antibacterial,antioxidant,and hemostatic effects.This article reviewed the pharmacological effects of Zanthoxyli Radix based on its functional indications.Based on the basic requirements of TCM quality markers(Q-markers),this article predicted and analyzed the Q-markers of Zanthoxyli Radix from the perspectives of plant phylogeny and chemical component specificity,chemical component and pharmacological correlation,and chemical component testability.It is proposed to select alkaloids,flavonoids,and lignans as the Q-markers for the general classification of Zanthoxyli Radix.The candidate components for Q-marker were identified,including chloramphenicol,white croaker alkaloid,magnolian alkaloid,taro alkaloid,vanillin,hesperidin,L-sesamin and L-asarone,providing a reference for further research on the quality standards of Zanthoxyli Radix.

OBJECTIVE:To study the purification technology of Sanhuang yishen formula. METHODS:Using retention rate and impurity rate of purified total polysaccharide,astragaloside and calycosin glucoside as index,the purification effects of water extraction and alcohol precipitation method (50%,60%,70% ethanol) and clarifying agent method (101 juice clarifying agent, ZTC natural clarifying agent,chitosan clarifying agent) were respectively detected to screen the purification method;orthogonal test was used to optimize the technology parameters(mass concentration of liquid,amount of clarifying agent and pH of liquid)by the optimized purification method,and the verification test was conducted. RESULTS:The purification was better when using chito-san as clarifying agent with comprehensive score of 98.62;the purified technology parameters were mass concentration of liquid 1 g/mL,1% chitosan solution amount of 2 mL/g,pH 5.1;the average value of retention rate and impurity rate of purified total poly-saccharide, astragaloside and calycosin glucoside in verification test were 79.56%(RSD=1.24%, n=3), 78.11%(RSD=0.97%,n=3),79.46%(RSD=1.03%,n=3)and 32.18%(RSD=1.16%,n=3),respectively. CONCLUSIONS:Using chito-san as clarifying agent shows good purification effect for Sanhuang yishen formula,which is simple. The optimized technology is stable and feasible.

OBJECTIVE:To optimize the extraction technology of Compound qima capsules. METHODS:With the blood pres-sure lowering of rats as index,pharmacological efficacy test was used to screen the preparation technology(A was whole herb de-coction;B was Gastrodia elata fine powder mixed with other decocted medical materials). The extraction technology was opti-mized by single factor and orthogonal test using the contents of astragaloside and isoflavone grape glycosides and the quality of sol-id as indexes,with added water,decoction time,decoction times as factors;and the verification test was carried out. RESULTS:Pharmacological efficacy test showed that antihypertensive effect of sample by technology B was superior. The optimal extraction condition of other medical materials of technology B was as follows as 12-fold water per time,decocting for 1.5 h,for 3 times. In verification test,average extraction rates of astragaloside and isoflavone grape glycosides were 64.02% and 51.97%,and average value of the quality of solid was 5.69 g(RSD≤1.92%,n=3). CONCLUSIONS:The optimized extraction technology is stable and feasible.

BACKGROUND: Compared with bone marrow-derived mesenchymal stem cells (MSCs),human umbilical cord blood (hUCB)-MSCs possess many advantages,including easy acquirement,low immunogenicity,able to tolerance a higher degree of HLA-matching inconsistency,and with higher purity.OBJECTIVE: To analyze the method and condition for in vitro isolation,purification,proliferation,and osteoblast and lipoblast differentiation of hUCB-MSCs.DESIGN,TIME AND SETTING: The present observational experiment was performed in the Key National Laboratory of Biological Treatment,Huaxi Hospital,Sichuan University (i.e.,Institute of Stem Cells and Tissue EngIneering) between September 2004 and November 2005.MATERIALS: Neonatal cord blood at gestational age 37 to 40 weeks.METHODS: hUCB-MSCs were collected in aseptic condition,isolated by density gradient centrifugation,sedimenting red cells with methylcellulose followed by density gradient centrifugation,or immunomagnetic beads negative technique of CD34+.Theisolated MSCs were used to carry on plastic adherent culture in L-DMEM +10% fetal bovine serum (FBS) or MesencultTM medium +10% FBS.The third passage of cells were used for surface antigen determination by flow cytometry and induced to differentiate towards osteoblasts and lipoblasts.MAIN OUTCOME MEASURES: ①Identification of hUCB-MSCs.②Confirmation of hUCB-MSC differentiation by alizarin red and oil red staining.RESULTS: The mononuclear cells isolated by sedimented and centrifuged way cultured in MesencultTM medium +10% FBS were most available.Obvious colonies appeared in the third passage.The cells obtained by only centrifugation in density gradient were hard to form colony,and those isolated by immunomagnetic beads were hard to culture.The surface antigens of these colony cells presented CD29,CD59,and CD71,hut did not express CD34,CD45,and HLA-DR,and so on.After alizarin red staining,osteoblast-differentiating colony cell cytoplasm exhibited mineralized matrices.After old red staining,lipoblast-differentiating colony cell cytoplasm was full of lipid vacuoles.CONCLUSION: The MSCs can be successfully isolated by sedimenting red cells with methylcellulose followed by centrifugation and cultured in MesencultTM medium +10% FBS.Obvious colony growth appears in the third passage.After induction,the MSCs can differentiate into osteoblasts and lipoblasts.