Objective To explore the mechanism of Danggui Sini Decoction in treating knee osteoarthritis(KOA)by using network pharmacology,combined with in vitro experimental verification.Methods The active components of Danggui Sini Decoction were retrieved and screened through TCMSP,CNKI and PubMed databases,and their corresponding targets were predicted using the SwissTarget Prediction platform.KOA related targets were retrieved from GeneCards and OMIM databases.Intersection of drug targets and KOA targets was obtained,and key components,core targets,and their related biological mechanisms were identified through network topology analysis,GO and KEGG pathway enrichment analysis.Molecular docking was used to verify the degree of binding between key components and core targets.Danggui Sini Decoction containing serum was prepared,and an in vitro KOA model was constructed by inducing rat articular chondrocytes with lipopolysaccharide.The CCK-8 method was used to screen for the optimal concentration of drug containing serum intervention,fluorescent probes were used to detect intracellular ROS content,immunofluorescence was used to detect NF-κB p65 nuclear translocation,RT-qPCR was used to detect the expressions of IL-6,IL-1β and TNF-α mRNA,Western blot was used to detect the expression of Toll-like receptor 4(TLR4),myeloid differentiation factor 88(MyD88),NF-κB p65 and p-NF-κB p65 proteins.Results The results of network pharmacology analysis indicated that the treatment of KOA with Danggui Sini Decoction may involve regulating Toll like receptors,MAPK,TNF,apoptosis and other inflammatory and aging signaling pathways,compounds such as quercetin,kaempferol and glycyrrhizin may play important roles.Core targets included IL6,IL1B,TNF,TLR4,NFKB1,JUN,MAPK1,RELA.In vitro experiments showed that compared with the blank group,the ROS content in chondrocytes of the model group significantly increased(P<0.01),NF-κB p65 protein was significantly nuclear translocation(P<0.01),and mRNA expressions of IL-6,IL-1β,TNF-α mRNA and protein expressions of TLR4,MyD88 and p-NF-κB p65 significantly increased(P<0.01);compared with the model group,the intervention of Danggui Sini Decoction containing serum could significantly reduce intracellular ROS content(P<0.05),inhibit NF-κB p65 nuclear translocation(P<0.01),and reduce the expression of inflammatory factor mRNA and related proteins(P<0.05,P<0.01).Conclusion Danggui Sini Decoction can significantly reduce the inflammatory response of rat articular chondrocytes induced by lipopolysaccharide and exert therapeutic effects on KOA.Its mechanism may be related to the inhibition of TLR4/MyD88/NF-κB pathway.

Objective To explore the mechanism of Danggui Sini Decoction in treating knee osteoarthritis(KOA)by using network pharmacology,combined with in vitro experimental verification.Methods The active components of Danggui Sini Decoction were retrieved and screened through TCMSP,CNKI and PubMed databases,and their corresponding targets were predicted using the SwissTarget Prediction platform.KOA related targets were retrieved from GeneCards and OMIM databases.Intersection of drug targets and KOA targets was obtained,and key components,core targets,and their related biological mechanisms were identified through network topology analysis,GO and KEGG pathway enrichment analysis.Molecular docking was used to verify the degree of binding between key components and core targets.Danggui Sini Decoction containing serum was prepared,and an in vitro KOA model was constructed by inducing rat articular chondrocytes with lipopolysaccharide.The CCK-8 method was used to screen for the optimal concentration of drug containing serum intervention,fluorescent probes were used to detect intracellular ROS content,immunofluorescence was used to detect NF-κB p65 nuclear translocation,RT-qPCR was used to detect the expressions of IL-6,IL-1β and TNF-α mRNA,Western blot was used to detect the expression of Toll-like receptor 4(TLR4),myeloid differentiation factor 88(MyD88),NF-κB p65 and p-NF-κB p65 proteins.Results The results of network pharmacology analysis indicated that the treatment of KOA with Danggui Sini Decoction may involve regulating Toll like receptors,MAPK,TNF,apoptosis and other inflammatory and aging signaling pathways,compounds such as quercetin,kaempferol and glycyrrhizin may play important roles.Core targets included IL6,IL1B,TNF,TLR4,NFKB1,JUN,MAPK1,RELA.In vitro experiments showed that compared with the blank group,the ROS content in chondrocytes of the model group significantly increased(P<0.01),NF-κB p65 protein was significantly nuclear translocation(P<0.01),and mRNA expressions of IL-6,IL-1β,TNF-α mRNA and protein expressions of TLR4,MyD88 and p-NF-κB p65 significantly increased(P<0.01);compared with the model group,the intervention of Danggui Sini Decoction containing serum could significantly reduce intracellular ROS content(P<0.05),inhibit NF-κB p65 nuclear translocation(P<0.01),and reduce the expression of inflammatory factor mRNA and related proteins(P<0.05,P<0.01).Conclusion Danggui Sini Decoction can significantly reduce the inflammatory response of rat articular chondrocytes induced by lipopolysaccharide and exert therapeutic effects on KOA.Its mechanism may be related to the inhibition of TLR4/MyD88/NF-κB pathway.

Objective To conduct hazard vulnerability analysis(HVA)in a blood bank,aimed to identify high-risk e-vents and optimize emergency management measures.Methods The risk event evaluation index system was established by referring to the Kaiser model and the situation of Guangzhou Blood Center,and risk events were ranked by risk matrix and Borda count.Results The top five events with the highest risk values identified by Kaiser model were information system e-mergencies(39.61％),extreme weather(38.03％),major public sentiment(37.86％),public health events(37.37％)and policy changes(37.24％).The results of risk matrix and Borda count revealed 1 extremely high-risk indicator as information system emergency,5 high-risk indicators with the highest risk being major public sentiment,11 medium-risk indicators with the highest risk being major medical disputes and 1 low-risk indicator as external fires.Conclusion Conducting HVA in combination with the actual situation of blood banks can effectively identify high-risk events and provide theoretical basis for improving emergency management measures.

0.05). The generation of ROS increased obviously after the cells were incubated with them for 12 h, and the increase of ROS reached the peak treated for 24 h. The levels of ?? m were time-dependently decreased after treating with four compounds for 12, 24, and 48 h. Conclusion The grouth of both MDR KBv200 cells and parental drug-sensitive KB cells were inhibited to the treatment of four anthraquinone derivative in vitro. The mechanism of their effects is associated with the increase of the cellular ROS level and the decrease of ?? m.

AIM We found in experiment that the relation of cell number-OD in MTS cytotoxicity assay is non-linearity. In this paper we probe into the relations of cell number-OD and drug dose-inhibition rate. METHODS Cells HL-60 and Raji were used in this experiment. Assay on the relation of cell number-OD: Cells of different concentrations were seeded into 96 well plates. OD was read at 490 nm after incubating with MTS for 1～5 hour. The regression curves were estimated from the data with the software SPSS 11.0. Cytotoxicity assay: Cells were seeded into 96 well plate and incubated with drug of different doses for 72 hours. OD was read at 490 nm after incubating with MTS for 3 hour. The regression curves were estimated from the data with the software SPSS 11.0. RESULTS The cubic curves fit well the relations of cell number-OD of the 2 strains of cells at 5 time points and the coefficient R2 of these cubic curves is 0.997～1.000, greater than the R2 0.938～0.993 of linear model. For the relations of dose-inhibition rate, the cubic curves also have best fitness and their R2 is 0.998～1.000 greater than the R2 0.948～0.987 of linear model of logarithm dose-probit. CONCLUSION For the relations of cell number-OD in MTS cytotoxicity assay, the fitness of cubic curves are better than the linear model. A more accurate IC 50 would be obtained by the cubic curve equation than by probit regression that is in common use.