Objective To construct plvx-cyclooxygenase-2(COX-2)-DsRed and establish breast cancer cell line MCF7 which overexpressed COX-2, to explore the effect of COX-2 on breast cancer cell.Methods The full-length COX-2 PCR product was obtained by total COX-2 PCR primers and COX-2 cDNA vector.After the PCR product and lentiviral vector plvx-DsRed-Monomer-N1 were cut simultaneously by restriction enzyme BamH1 and Xholl, they were connected and sequenced, to get lentiviral vector plvx-COX-2-DsRed.After selected by puromycin, overexpressed COX-2 breast cancer cell line MCF7-plvx-COX-2-DsRed was obtained.The stable cell line was verified by real time PCR and Western blot.The differences of proliferation ability between stable cell line and normal one were compared by colony formation test and Western blot.Results The lentiviral vector plvx-COX-2-DsRed and stable cell line MCF7-plvx-COX-2-DsRed after selecting were obtained.COX-2 expression level of the stable cell line was 75.29 times as high as that of MCF7, and 64.91 times as high as that of cell line MCF7-plvx-DsRed-Monomer-N1 by PCR assay (P ＜ 0.05), which was consistent with the results of Western blot and microscope photo.MTT results showed that cell line MCF7-plvx-COX-2-DsRed had grown faster than cell line MCF7 and MCF7-plvx-DsRed-Monomer-N1 from the 2nd day (P ＜ 0.05), which was accordant with colony formation assay.MCF7-plvx-COX-2-DsRed cell line had higher c-myc expression and lower β-catenin expression than MCF7 cell and cell line MCF7-plvx-DsRed-Monomer-N1 detected by Western blot relative quantification (P ＜ 0.05).Conclusion The plvx-COX-2-DsRed lentiviral vector and cell line MCF7-plvx-COX-2-DsRed are successfully constructed.COX-2 can increase proliferation of MCF7 cells through up-regulating the expression of c-myc.

Objective To observe the effect of Wenxiao Decoction in treating the patients with carotid atherosclerostic (CAS) plaque and assess the safety. Methods Sixty patients with CAS plaque were randomly allocated to treatment group (33 cases) and control group (27 cases). The control group was given routine therapy, and the treatment group was given Wenxiao Decoction additionally. And they were further divided into half-year treatment group and half-year control group, 1-year treatment group and 1-year control group. Carotid artery color Doppler ultrasound and hepatorenal function were examined before and after treatment. Results Compared with the control groups after therapy, the average numbers of plaques, maximal plaque thickness, Crouse plaque integral were decreased both in 1-year and half-year treatment groups (P<0.05, P<0.01). The plaque stability of 1-year treatment group was better than that of half-year treatment group. The average numbers of plaques, plaques size and Crouse plaque integral increased in control groups after routine therapy (P<0.05, P<0.01). Bilateral common carotid artery diameter, intima-medial thickness (IMT) and hepatorenal function had no statistical difference between before and after treatment both in treatment and control groups (P>0.05). Conclusion Wenxiao Decoction is effective and safe in treatment of CAS plaque, and plaque stability is more obvious with the extension of treatment time.

Objective To investigate ACEI ( benazepril ) and tranilast exert renoprotective properties in diabetic nephropathy( DN) through the inhibition of thioredoxin( Trx) . Methods Forty male SD rats were randomly divided into normal control group,model control group,tranilast group and benazepril group (n=10 each).Normal control group was fed with normal diet. Other groups were fed with high-glucose high-fat diet to make DN models. Rats in model control, tranilast, and benazepril groups were fed with normal diet,400 mg??kg-1??d-1 tranilast plus normal diet,and 10 mg??kg-1??d-1 benazepril plus normal diet,respectively,via oral gavage for 12 weeks.The 24-hour proteinuria,blood glucose(BG),blood urea nitrogen (BUN),serum creatinine ( Scr) and renal pathology changes were detected. Expression of Trx was measured by Western-blot. Results The 24 h urine protein, BG, BUN, Scr, kidney/body weight, and glomerular sclerosis index were significantly decreased in tranilast group and benazepril group,as compaired with model control group ( P<0.05) ,but there was no statistical difference between the two drug groups (P>0.05).Both tranilast and benazepril can reduce renal pathological changes,and can increase the expression of Trx of DN rats, but benazepril had a more significant effect on increasing Trx expression. Conclusion Both tranilast and benazepril have renoprotective function in DN, and benazepril is more effective than tranilast in delaying the progression of diabetic nephropathy by increasing Trx expression and decreasomg oxidative stress.

BACKGROUND:Semen cuscutae has the effect of tonifying the liver and kidney system and benefiting the essence.The main pathogenesis of osteoarthritis is deficiency of the liver and kidneys.Therefore,it is hypothesized that there is a link between semen cuscutae and osteoarthritis. OBJECTIVE:To explore the potential relationship between osteoarthritis and semen cuscutae and validate the mechanism of semen cuscutae based on the network pharmacology and molecular docking analysis. METHODS:First,the active ingredients and targets of semen cuscutae were screened in TCMSP,and the genes related to osteoarthritis were collected in the disease databases GeneCard's,OMIM and TTD.The intersected genes were taken and then subjected to a series of analyses and screened for hub genes.Through the enrichment analysis of hub genes,the pathway of semen cuscutae acting on osteoarthritis was selected.The role of hub genes was verified by molecular docking.Therefore,the appropriate active ingredients of semen cuscutae were selected for experimental verification. RESULTS AND CONCLUSION:There were 11 active ingredients of semen cuscutae,66 intersection target genes of semen cuscutae and osteoarthritis,and 12 hub genes,including tumor necrosis factor,interleukin 1B,TP53,RAC-alpha serine/threonine protein kinase(AKT1),vascular endothelial growth factor A,matrix metalloproteinase 9,prostaglandin peroxidase 2,cystatinase 3,epidermal growth factor,peroxisome proliferator-activated receptor gamma,interleukin 10,vascular cell adhesion factor 1.After the enrichment analysis of the hub genes,the classical inflammatory pathway,nuclear factor-κB signaling pathway,was selected for subsequent validation of semen cuscutae to alleviate osteoarthritic inflammation.Through the results obtained after molecular docking of each active ingredient and the hub gene of the pathway prostaglandin peroxidase 2,sesamin with the highest affinity was selected for subsequent cell experiments,and the experimental results confirmed that sesamin,the active ingredient of semen cuscutae,could reduce the expression of cyclooxygenase 2 by inhibiting the nuclear factor-κB signaling pathway induced by interleukin-1β.To conclude,sesamin,the active ingredient of semen cuscutae,reduces the expression of cyclooxygenase 2 by inhibiting the nuclear factor-κB signaling pathway induced by interleukin-1β,thereby improving inflammation in osteoarthritis and expanding the therapeutic effect of semen cuscutae in osteoarthritis.

Objective:To explore the immune and angiogenesis-related genes in aortic valve stenosis(AS)and potential therapeutic targets, based on bioinformatics and machine learning analysis.Methods:AS data sets from the Gene Expression Omnibus(GEO), immune-related genes from the ImmPort database, and angiogenesis-related genes from the Genecards database and MsigDB were downloaded and combined to determine differentially expressed immune and angiogenesis-related genes(DEGs).Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)functional enrichment analyses were performed.Protein-protein interaction(PPI)was analyzed by using STRING database.The key biomarkers were identified by two machine learning methods including Least Absolute Shrinkage Selection Operator(LASSO)and Support Vector Machine Recursive Feature Elimination(SVM-RFE), validated in training data set and verification data set by receiver operating characteristic curve(ROC), and analyzed by Gene Set Enrichment Analysis(GSEA).The subtypes of immune infiltrating cells were analyzed by CIBERSORT.Based on starBase, miRDB, miRWalk and hTFtarget databases, the mRNA-miRNA-TF was constructed.Finally, Potential therapeutic targets and drugs were analyzed through the CTD database.Results:A total of 90 DEGs related to AS, immune, and angiogenesis were obtained.Enrichment analysis found that DEIRGs were mainly related to immune regulation and cell cycle regulation, such as "leukocyte migration" , "cell chemotaxis" and "cytokine-cytokine receptor interaction" .84 related proteins and 548 interactions were obtained by PPI analysis.Two key biomarkers SecretograninⅡ(SCG2)and Tenascin-C(TNC)were identified by machine learning, which showed high diagnostic value for AS by ROC.SCG2 and TNC are mainly involved in the immune regulation by Enrichment analysis.The infiltration level of macrophage M0 in AS group was significantly higher than that in control group by CIBERSORT analysis.The correlation between macrophage M0 and macrophage M2 and SCG2 was the highest.879 mRNA-miRNA-TF, 253 potential therapeutic agents and 299 relationships were obtained.Conclusions:The key biomarkers, immune characteristics and potential therapeutic targets obtained from the research play a vital role in exploring the pathophysiological progress and new therapeutic strategies of AS.

ObjectiveTo evaluate the intervention effect of meteorological risk forecasting service on acute onset and medical expenses of chronic obstructive pulmonary disease（COPD） patients， and to provide scientific basis for the establishment of health management model for chronic obstructive pulmonary disease（COPD） patients. MethodsStudy subjects were recruited from chronic obstructive pulmonary patients aged ≥40 in Pudong New Area. Propensity score matching method was used to determine the intervention group and the control group. The control group received regular health education and follow-up management， and the intervention group was provided with meteorological and environmental risk forecasting services through WeChat， mobile phone short message service（SMS）and telephone. Finally， a total of2 589 subjects were included in the analysis， including 1 300 in the intervention group and 1 289 in the control group. General demographic data， past medical history and family history of COPD， COPD related knowledge and practice survey， COPD related symptom assessment， acute onset， health service utilization and medical expenses before and after intervention were collected through questionnaire survey. The differences of acute attack， health service utilization and related medical expenses between the two groups before and after intervention were compared to evaluate the intervention effect. ResultsIn terms of acute attacks， after intervention， the incidence of acute attacks in the intervention group was lower than that before intervention（χ²=52.901， P<0.001）， and the incidence of acute attacks in the groups with different intervention methods was lower than that before intervention （P<0.001）. WeChat had the best effect， decreasing the incidence by 14.4%， followed by mobile phone SMS SMS decreasing by 12.3%. In terms of utilization of health services， the outpatient rate due to acute attack was lower in the intervention group after intervention than that before intervention （χ²=7.129， P=0.008）， and the outpatient rate due to acute attack was lower in the subjects who received the forecast service through mobile phone SMS than that before intervention （χ²=4.675， P<0.001）. In terms of medical expenses， there was no significant difference between control group and intervention group with different intervention methods before intervention （P>0.05）. After intervention， the difference between the control group and the intervention group with different intervention methods was statistically significant （H=11.864， P<0.05）. The results of multiple comparisons showed that compared with the control group， the average annual medical expenses of patients receiving mobile phone SMS and telephone forecasting services after intervention were lower than those of the control group， and the difference was statistically significant （P<0.05）. ConclusionMeteorological risk forecasting service can reduce the acute onset of COPD， reduce the rate of consultation and medical expenses due to acute onset， and provide scientific basis for the basic COPD health management model.

Hepatic ischemia/reperfusion injury (HIRI) is a serious complication that occurs following shock and/or liver surgery. Gut microbiota and their metabolites are key upstream modulators of development of liver injury. Herein, we investigated the potential contribution of gut microbes to HIRI. Ischemia/reperfusion surgery was performed to establish a murine model of HIRI. 16S rRNA gene sequencing and metabolomics were used for microbial analysis. Transcriptomics and proteomics analysis were employed to study the host cell responses. Our results establish HIRI was significantly increased when surgery occurred in the evening (ZT12, 20:00) when compared with the morning (ZT0, 08:00); however, antibiotic pretreatment reduced this diurnal variation. The abundance of a microbial metabolite 3,4-dihydroxyphenylpropionic acid was significantly higher in ZT0 when compared with ZT12 in the gut and this compound significantly protected mice against HIRI. Furthermore, 3,4-dihydroxyphenylpropionic acid suppressed the macrophage pro-inflammatory response in vivo and in vitro. This metabolite inhibits histone deacetylase activity by reducing its phosphorylation. Histone deacetylase inhibition suppressed macrophage pro-inflammatory activation and diminished the diurnal variation of HIRI. Our findings revealed a novel protective microbial metabolite against HIRI in mice. The potential underlying mechanism was at least in part, via 3,4-dihydroxyphenylpropionic acid-dependent immune regulation and histone deacetylase (HDAC) inhibition in macrophages.