Objective To optimize the extraction process of Euphorbia hirta L.by Central composite design/response surface methodology with ethanol by the ultrasonic extraction technology.Methods Based on the single factor experiment and central composite design-response surface methodology,the content of quercitroside was regarded as evaluation index,select ethanol concentration,extraction time,and solid-liquid ratio as three evaluation factors to achieve the purpose. Results The results indicated that the optimal extraction process was obtained as follows:10 times the amount of 60% ethanol under ultrasound for 2 times,50 min each;the average content of the quercitroside was 2.39 mg/g.Conclusion The method to optimize the extraction process is simple and reasonable.

Objective To evaluate the effect of enteral nutrition emulsion on the immunologic function and intestinal mucous barrier in diabetic patients. Methods Eighty diabetic patients were randomly divided into con-trol group (n=40) and enteral nutrition group (n=40). The urine lactulose (L) and mannitol (M) levels and the blood immunologic indicators recorded on day 1 and day 8. Results The L/M ratio was significantly lower in enteral nutrition group than in control group on day 1 and day 8 ( P < 0. 05 ). The IgG level was significantly higher in enteral nutrition group than in control group on day 8 ( P = 0. 02 ). Conclusion Enteral nutrition emulsion can decrease the permeability of intestinal mucous membrane and improve the immunologic function in diabetic pa-tients.

Objective:To develop an HPLC-DAD-ELSD method for the determination of nitidine chloride, 5-ethoxychelerythrine, bergeninum and ardisiacrispin A in Shangtong tinctures ( STD) . Methods: A Hypersil C18 column was used as the chromatographic column, the flow rate was 0. 8 ml·min-1 . For nitidine chloride and 5-ethoxychelerythrine, the mobile phase A consisted of acetoni-trile,the mobile phase B consisted of 0. 1% formic acid-triethylamine (pH 4. 5),and the DAD detection wavelength was at 273 nm. For bergeninum and ardisiacrispin A, the mobile phase consisted of methanol-water(25∶75), the temperature of drift tube was set at 95℃, and the gas flow (N2) was set at 2. 5 SLPM·min-1. Results:There was a good linear relationship between the concentration and peak area for nitidine chloride and 5-ethoxychelerythrine within the range of 0. 021-0. 426 μg (r=0. 999 5) and 0. 075-1. 494 μg (r=0. 999 8), respectively. The average recovery was 99. 22%(RSD=0. 64%) and 98. 61%(RSD=0. 46%), respectively. There was a good linear relationship between the concentration and peak area for bergeninum and ardisiacrispin A within the range of 0. 215-4. 304 μg(r=0. 999 3) and 0. 286-5. 728 μg(r=0. 999 7), respectively. The average recovery was 99. 15%(RSD=0. 77%) and 99. 25%(RSD=0. 56%) accordingly. Conclusion:The method is accurate, sensitive and reproducible, and can be used in the de-termination of nitidine chloride, 5-ethoxychelerythrine, bergeninum and ardisiacrispin A in STD.

OBJECTIVE:To establish a method for simultaneous determination of the contents of ligustroflavone,specnuezhe-nide,demethylwedelolactone and wedelolactone in Zibu ganshen pill. METHODS:Dual-wavelength HPLC was performed on the column of Elite C18 with mobile phase of acetonitrile-0.5%acetic acid(gradient elution)at flow rate of 0.9 ml/min,column temper-ature was 25 ℃,detection wavelengths were 224 nm(0-30 min) and 351 nm (30-50 min),and the injection volume was 20 μl. RESULTS:The linear range was 6.75-135.00μg/ml(r=0.999 5)for ligustroflavone,6.54-130.80μg/ml(r=0.999 8)for specnuezhe-nide,4.90-98.00μg/ml(r=0.999 4)for demethylwedelolactone and 6.42-128.40μg/ml(r=0.999 6)for wedelolactone;RSDs of pre-cision,stability and reproducibility tests were no more than 1.25%;average recoveries were 96.15%-99.96%(RSD<2%,n=6). CONCLUSIONS:The method is rapid,sensitive and accurate,and can be used for the contents determination of ligustroflavone, specnuezhenide,demethylwedelolactone and wedelolactone in Zibu ganshen pill.

Objective To study the risk factors of Pseudomonas aeruginosa (PA)infection and homology of PA isolated from nasopharyngeal carcinoma patients undergoing radiotherapy,and prevent and control the spread of in-fection.Methods Bacteria isolated from clinical specimens were identified by BD Phoenix automated microbiology system. Gene homology were analyzed with randomly amplified polymorphic DNA (RAPD)technique.Results Forty-nine strains of PA were mainly isolated from 43 nasopharyngeal carcinoma patients,the major specimens were nasopharyngeal swab (46.94% ),sputum(32.65% ),and oral secretion (10.20% ).All these strains were amplified 46 electrophore-sis diagrams,and 19 genotypes were identified. The highly homologous genotypes of type H and J strains were mainly isolated from patients in the second section (57.14% ,4/7)and fourth section (60.00% ,3/5)of radiotherapy department respectively. Conclusion Localized epidemic of highly homologous PA exists in different sections of ward,transfer of patients between different sections is the risk factor for homology PA infection/colonization. Gen-otyping technique such as RAPD for analyzing the homology of pathogenic bacteria in healthcare-associated infection has important value in preventing and control Ling infection spread.

BACKGROUND: Revascularization is a challenge for the tissue-engineered bone carrying cells after implanted into human body. Previous studies have found that tanshinol can improve the functions of endothelial progenitor cells and exert vascular protective effects.OBJECTIVE: To prepare the β-calcium phosphate (β-TCP) scaffold with tanshinol coating, and to observe its cytocompatibility.METHODS: The β-TCP scaffolds coated with 10-7, 10-6 and 10-5 mol of tanshinol were constructed by negative pressure absorption method. The distribution of tanshinol coating on the scaffold was observed using scanning electron microscopy,and the inner ingredients were analyzed by infrared spectrum. Human endothelial progenitor cells (hEPCs) were cultured in the extracts of β-TCP and β-TCP scaffolds with 10-7, 10-6, 10-5 mol of tanshinol coatings, respectively. The cell proliferation was detected at 2, 4, 6, 8 and 10 days of culture; the levels of nitric oxide and vascular endothelial growth factor in the supernatants were detected at 1, 7 and 14 days of culture; the lumen formation on the matrigel was observed after 14-day culture. hEPCs were respectively seeded onto the β-TCP and β-TCP scaffolds with different dosages of tanshinol coating,and then the cell growth was observed under scanning electron microscope at 7 days.RESULTS AND CONCLUSION: The tanshinol coating evenly distributed on the inner surface of the pores, and its crystalline structure became dense with dosage increasing. Infrared spectrum analysis revealed no changes in the characteristic absorption peak of tanshinol and TCP in the scaffold. The β-TCP scaffolds with tanshinol coating could promote the proliferation of hEPCs, especially the scaffolds with 10-6 and 10-5 mol tanshinol coating. Compared with the β-TCP scaffold, the scaffolds with 10-6 and 10-5 mol tanshinol coating significantly upregulated the nitric oxide level at 14 days of culture, and significantly increased the level of vascular endothelial growth factor at 7 and 14 days of culture (P <0.05). Although it could be found in all β-TCP scaffolds with tanshinol coating, the lumen formation was the maturest in the scaffold with 10-5 mol tanshinol coating. These results suggest the β-TCP scaffolds with tanshinol coating can promote the proliferation and endothelial differentiation of hEPCs, and hold a good cytocompatibility.

Objective To detect the levels of peptidylarginine deiminase 4 (PAD4) mRNA and neutrophil extracellular traps (NETs) of the peripheral blood neutrophils in rheumatoid arthritis (RA) patients and to study the association between PAD4 and NETs in RA.The anti-cyclic citrullinated peptide (anti CCP) antibodies,disease activity score (DAS28) score,erythrocyte sedimentation rate (ESR) were recorded.Its value in the pathogenesis of RA was explored.T test,rank-sum test,Pearson and Spearman correlative analysis were used for statical analysis.Methods The serum double-stranded DNA (dsDNA)/NETs in 43 RA patients and 20 healthy controls were detected by PicoGreen dsDNA Quantitation Kits (Invitrogen).Real-time quantitative polymerase chain reaction (RT-PCR) was used to determine the expression of PAD4 mRNA in neutrophils,and the relationship between dsDNA/NETs and their clinical indexes were analyzed.Results ① The level of peripheral blood neutrophils PAD4 mRNA in RA was significantly higher than healthy controls [1.493(0.831,2.607)] vs [0.631(0.358,1.489)],(Z=-2.07,P=0.039).The levels of serum PAD4 mRNA in RA treated with disease-modifying antirheumatic drugs (DMARDs) were lower than untreated with DMARDs but no significant difference was found (Z=-1.19,P=0.234).② The levels of serum dsDNA/NETs in RA patients were significantly higher than in healthy controls (t=3.4,P=0.001).③ The levels of serum dsDNA/NETs in RA were positively correlated with DAS28 score,ESR,CRP and PAD4 mRNA (r=0.36,P=0.036;r=0.345,P=0.042;r=0.36,P=0.043;r=0.42,P=0.017) but not with anti-CCP antibodies (r=0.277,P=0.154).Conclusion ① PAD4 may play an important role in the pathogenesis of RA.② NETs maybe associated with disease activity and play an important role in RA pathogenesis,inhibit NETs generation or improve the ability to clear NETs,perhaps can treat RA.③ PAD4 probably play an important role in rheumatoid arthritis by influencing the formation of the NETs.

Objective To explore the significance of aryl hydrocarbon receptor (AhR) in patients with rheumatoid arthritis (RA) through detecting the levels of AhR and its response gene cytochrome P4501 A1 (CYP1 A1) in peripheral blood mononuclear cells (PBMCs).Methods Peripheral blood samples were collected from 35 patients with RA and 20 healthy subjects.The expression of AhR and CYP1A1 at mRNA level were detected by real-time PCR.The percentages of AhR-positive cells in PBMCs were detected by flow cytometry (FCM).The effects of leflunomide (LEF) on the expression of AhR and CYP1A1 were analyzed.The detailed clinical data of RA patients were recorded.The disease activity score (DAS) was calculated.Correlation analysis between AhR/CYP1A1 level and clinical data was conducted.Results (1) Both the expression of AhR at mRNA level and the percentage of AhR-positive cells in PBMCs from RA patients without LEF treatment were significantly higher than those from healthy subjects [(3.61±1.65) vs.(2.00±1.27),P=0.002; (34.21±11.30)％ vs.(18.83±7.32)％,P＜0.01].There were no statistically significant differences in the expression of AhR at mRNA level and the percentages of AhR-positive cells between patients with or without LEF treatment [(3.83 ± 1.62) vs.(3.61 ± 1.65),P =0.670 ; (36.69±10.61)％ vs.(34.21±11.30)％,P=0.462].(2) Non-LEF treatment group showed a higher relative expression of CYP1 A1 at mRNA level than that from control group [1.33 (0.08,7.86) vs.(0.62 ±0.29),z=-3.922,P＜0.01],but there was no statistical difference between LEF treatment group and non-LEF treatment group [(2.62±2.08) vs.1.33(0.08,7.86) z=-0.133,P=0.894].(3) Neither the expression of AhR and CYP1A1 at mRNA level nor the percentages of AhR-positive cells showed significant correlations with clinical data.Conclusion AhR was highly expressed in PBMCs from patients with RA,which might participate in the progression of rheumatoid arthritis.But the high expression of AhR did not reflect disease activity.Moreover,the treatment of LEF showed no significant influences on the expression of AhR and CYP1A1 in PBMCs from patients with RA.

Objective To analyze the expression of purinergic receptor P2X ligand-gated ion channel 7 (P2X7R) on different cells and peripheral blood mononuclear cell (PBMC) and to investigate its correlation with inflammatory cytokines in patients with SLE.Methods Flow cytometry was used to detect surface expression of P2X7R on lymphocytes,CD4+ cells,and CD19+ cell in 29 SLE patients and 28 healthy human controls to compare the difference between the SLE patients and the controls in P2X7R expression.Enzyme linked immunosorbent assay (ELISA) was performed to detect P2X7R-related serum cytokines interleukin (IL)-1β,IL-6,tumor necrosis factor (TNF)-α level.T test,Wilcoxon rank sum test,Spearman's correlation analysis were used for statitical analysis.Results ① SLE patients had significantly higher expression of P2X7R on CD4+,CD8+ lymphocytes compared to controls [CD4+ cells∶ 2.21(3.55) vs 0.89(1.15),Z=-1.527,P=0.015； CD19+ cells∶ 11.53(20.01) vs 6.66 (6.27),Z=-2.091,P=0.037]； ② The levels of three cytokines in patients with SLE were significantly higher than those in control.The positive relationship between P2X7R expression in lymphocytes with the serum IL-6 level was found in SLE patients (r=0.449,P=0.015)；③ Patients with arthritis showed significantly higher expression of P2X7R on lym-phocytes compared to patients without arthritis (Z=-2.772,P=0.006).The expression of P2X7R on lymphocytes and CD19+ cell was significantly positively correlated with the SLEDAI score.Positive correlation with anti-β2GP Ⅰ in lymphocyteswas also found.Conclusion P2X7R may mediate the release of inflammatory cytokines involved in the pathogenesis of SLE,and may participate the development of arthritis,lupus nephritis and NPSLE in SLE patients.

Objective To explore the role of serum neutrophil extracellular traps (NETs) in rheumatoid arthritis (RA).Methods The serum circulating free DNA (cf-DNA) / NETs in 45 RA patients and 18 healthy controls were detected by Pico Green dsDNA Quantitation Kits,and the association between serum NETs and ESR,DAS28 score,anti-CCP antibodies was analyzed.T test,rank sum test,Pearson or Spearman correlation analysis were ued for statistical analysis.Results ① The levels of serum dsDNA/NETs in RA patients [0.523 0(0.282 0,1.637 0) μg/ml] were significantly higher than those in healthy controls [0.410 5 (0.140 0,0.966 0)μg/ml] (Z=-2.419,P=0.016).② The level of serum dsDNA/NETs in RA was positively correlated with ESR and DAS28 score (r=0.357,P=0.016; r=0.325,P=0.029),but not with anti-CCP antibodies(r=0.146,P=0.434).③ The levels of serum dsDNA/NETs in RA treated by DMARDs were lower than those of untreated with DMARDs[0.516 5(0.282 0,1.637 0) μg/ml,0.523 0 (0.369 0,1.485 0) μg/ml,Z=-1.215,P=0.225],but the difference was not significant.Conclusion NETs maybe associated with disease activity and play an important role in RA pathogenesis.