High-performance liquid chromatographic coupled with variable wavelength detection (HPLC-VWD) has been developed for simultaneous determination of 5 analytes including ellagic acid, quercetin, kaempferol-3-O-beta-D-rutinoside, tiliroside and kaempferol, and high-performance liquid chromatographic with an evaporative light scattering detector (HPLC-ELSD) has been established to determine goshonoside-F5 in extract of Rubi Fructus. Chromatographic separations were carried out on an Agilent ZORBAX SB-C18 column (4.6 mm x 250 mm, 5.0 microm). All calibration curves of reference standards revealed good linearity (R2 > 0.999 5) within the concentration ranges tested. The method limits of detection ranged 0.297-90.144 ng and the method limits ofquantitation ranged 0.990-300.480 ng, respectively. Recoveries of 6 analytes were from 97.11% to 101.7%, with RSD less than 2.1%. The result shows that amounts of the 6 analytes in the samples from 16 localities were found to be different. The higher latitude of growing environment, the more ellagic acid in herb. The content of total flavonoids in sample from east localities were higher than that in middle and west localities, and the content of goshonoside-F5 in Bozhou, Anhui province was higher than others. This method was found to be simple, accurate, sensitive with good repeatability. Those results might serve as a sound foundation for further study, quality control and application of Rubi Fructus.
Chromatography, High Pressure Liquid ; Ellagic Acid ; analysis ; Flavonoids ; analysis ; Geography ; Rosaceae ; chemistry

Objective To establish and evaluate intestinal epithelial barrier model using Caco-2 cell so as to play a foundation for next study of barrier permeability.Methods Caco-2 cells were cultured in vitro then seeded into Transwell cell culture inserts.The permeability of the intestinal epithelial barrier was detected by transepithelial electrical resistance(TEER)and lucifer yellow flux,and verified by transmission electron micro-scope.Different concentrations of PAF(0,50,100,and 200 nmol /L)were exposed for 24 hours to Caco-2 mono-layer when cultured 21 days.The tight junction was observed under transmission electron microscope.Assess-ment of ZO-1 protein localization and expression were detected by immunofluorescence and Western blot analy-sis.Results Cultured Caco-2 cell confluencd as monolayer with time passed.From 5th day,TEER increased, then reached 600Ω?cm2 at 15th day and lasted to 21 st day,there was little flux of lucifer yellow,transmission e-lectron microscopy also found cells differentiated better,had well-arranged villi and polarity alined as monolayer, forming completed tight junction which was the marker of intestinal epithelial barrier model in vitro.TEER de-creased and lucifer yellow flux increased in cells exposed to PAF.The permeability reached the peak when ex-posed to 100 nmol /L PAF(P <0.01 ),tight junction disrupted,ZO-1 protein expression downregulated,abnor-mal localization and distribution was assessed by immunofluorescence staining.Conclusion Cultured Caco-2 cells for 2-3w can be used to study intestinal epithelial barrier as a model in vitro.PAF increased intestinal epi-thelial permeability,which would correlate to the decreased protein expression and abnormal distribution of ZO-1.

Objective To investigate the feasibility to establish experimental model of autoimmune myocarditis and to study the value of echocardiographic assessment of left ventricular structure and function.Methods Seventy-two male 6 weeks old Lewis rats were randomly divided into 3 groups:normal control group,negative control group and positive group.Positive group were immunized with porcine cardiac myosin at days 0,7,30.Results ①The positive group showed weight loss,increased heart weight and myocardial necrosis with inflammatory infiltration.②The development of experimental autoimmune myocarditis included acute,subacute and chronic stages.The left ventricular diameter,ventricular wall thickness,left ventricular fractional shortening and ejection fraction of positive group differed significantly from those of other two groups.Conclusions Lewis rats immunized with porcine cardiac myosin may be a desirable experimental model of experimental autoimmune myocarditis,echocardiography can evaluate changes of cardiac structure and function accurately.

ObjectiveTo evaluate the effectiveness of transcatheter Amplatzer device closure on patent ductus arteriosus(PDA),and to give some evidences for the clinical application.MethodsAll studies in the world regard to the controlled trials(CT) about transcatheter Amplatzer device closure and cardiac surgery on PDA were searched and made synthetic evaluation by means of Meta-analysis.RevMan 4.2.2 software was used for statistical analysis.Cases relative risk(RR)and its 95% confidence interval(CI)of procedure failure,the incidence of complication and residual shunt were calculated.ResultsTotally 5 studies including 349 cases were analyzed.Operation failure of Amplatzer device occlusion was higher than cardiac surgery [5 CT,349 cases,3.0% vs 0,RR=4.29,95%CI(0.77,23.95)](P=0.10).Incidence of complication of Amplatzer device occlusion was lower than cardiac surgery[5 CT,343 cases,3.1% vs 38.0%,RR=0.11,95%CI(0.05,0.23)](P

Objective To investigate the diagnosis value of the tissue strain imaging in myocardial dysfunction in systemic lupus erythematosus (SLE). Methods Sixty-two patients and sixty controls underwent conventional and tissue Doppler echocardiography. Peak strain and strain rate value during systolic and diastolic phases as well as E/A, left ventricular fraction shortening (LVFS), left ventricular ejection fraction(LVEF) were measured in both SLE and the control groups. Results ①E/A,LVFS and LVEF did not differ between SLE patients and controls( P ＞0.05). The systolic peak strain and strain rate of SLE patients were lower than those of controls but without significant differences( P ＞0.05). ②The diastolic peak strain and strain rate of SLE patients were significantly lower than those of controls (P ＜0.01). ③ The diastolic peak strain and strain rate of antiheart antibody (AHA) positive patients were significantly lower than those of negative ones( P ＜0.05). Conclusions Strain and strain rate combining with AHA can sensitively detect myocardial dysfunction of SLE.

Objective To evaluate the respiratory mechanics and inflammatory status in elderly patients with stable chronic obstructive pulmonary diseases (COPD).Methods The arterial blood gases (ABGs),respiratory drive and its derivatives,mechanics of respiratory muscles,resistance and compliance of airway,interleukin-8 (IL-8)and interferon-?(IFN-?)were measured in 42 cases withstable COPD and 40 subjects of normal control.Results The elderly patients with stable COPD had great changes in the following parameters while compared with the control group:peak inspiratory pressure(PIMAX) [(4.48?2.11)vs(6.10?2.91)kPa],maximum expiratory pressure (PEMAX)[(6.30?3.20)vs(9.15?93.30)kPa],0.1s mouth occlusion pressure(P_(0.1)) with its correction index,airway resistance,compliance,ABGs,the levels of IL-8[(218.46?91.14) vs (161.84?14.40)ng/L]and IFN-?[(2435.82?639.92)vs(1652.40?95.08)ng/L],which might aggravate the progress of COPD consistently.Conclusions The elderly patiends with stable COPD has marked changes in respiratory drive,airway resistance,and airway compliance,respiratory mechanic and inflammatory status.The intervention should be performed in the elderly stable COPD patients.

Objective To observe whether the expression of interferon-regulatory factor genes are re- lated to systemic lupus erythematosus (SLE).Methods The clinical data of 45 SLE patients and 37 normal controls were collected.Total RNA of peripheral blood was extracted and transcripted into cDNA.Sybr green dye based real-time quantitative PCR method was used to compare the expression (indicated as-??Ct value) of IRFI,IRF4,IRF8 in patients with SLE and those in the controls.Results The levels of IRF1,IRF4 and IRF8 mRNA were-3.90?0.19,-8.04?0.25 and 3.60?0.15 respectively in normal controls.In SLE patients, IRF4 mRNA expression was -8.82?0.18,higher than that in normal (P=0.011).But IRF8 mRNA expression was 3.09?0.13,lower than that in normal (P=0.012).Conclusion Abnormal IRF mRNA expression is found in the peripheral blood of SLE patients.IRFs may play roles in the pathogenesis of SLE by affecting the differen- tiation of Th cells.

Objective To evaluate the value of volume measured by multi-slice spiral CT in preoperative N staging of gastric canc-er.Methods CT data of 1 93 cases of gastric cancer proven pathologically were collected and analyzed.Volume of the tumor was cal-culated in the portal phase,and the correlation between the results and N staging was evaluated.ROC curve was used to get diagnos-tic value to differentiate N stages.Results Intra-observer Kappa value was 0.77 (P < 0.05 ),0.72 (P < 0.05 ),Inter-observer Kappa value was 0.69 (P <0.05).The tumor volume data was positively correlated with different N stages (r=0.568,P <0.05). ROC curve showed that the volume could help differentiate between stage N0 and stage N1 - N3 (cutoff 12.06 cm3 ,sensitivity 55%,specificity 95%),stage N0-N1 and stage N2-N3 (cutoff 22.35 cm3 ,sensitivity 66%,specificity 86%),stage N0-N2 and stage N3 (cutoff 25.95 cm3 ,sensitivity 62%,specificity 89%)respectively.Conclusion The volume of gastric cancer measured by CT plays an important part in predicting lymph node metastasis staging and optimizing individualized clinical strategy for patients.

The progress in the research of the active ingredients of licorice flavonoid and the pharmacological activities was reviewed. Licorice flavonoid constituents mainly included flavones, flavonals, isoflavones, chalcones, bihydroflavones and bihydrochalcones. Pharmacological investigation concluded that they had antioxidant, antibacterial, antitumer and inhibiting HIV activities. It is important to study further the flavonoid constituents and pharmacological activities.
Animals ; Anti-HIV Agents ; pharmacology ; Antineoplastic Agents, Phytogenic ; pharmacology ; Antioxidants ; pharmacology ; Flavonoids ; chemistry ; isolation & purification ; pharmacology ; Glycyrrhiza ; chemistry ; Humans ; Molecular Conformation ; Molecular Structure ; Plants, Medicinal ; chemistry

OBJECTIVETo investigate the effect of vascular endothelial growth factor 165 (VEGF 165) gene on vascularization of dermal substitute in vivo.

METHODSHuman umbilical vein endothelial cells (HUVECs) were cultured in M199 medium containing FBS in the volume fraction of 10% (briefly called complete medium). (1) HUVECs were divided into non-transfection group (without transfection), empty vector group [transfected with pIRES2-enhanced green fluorescent protein (EGFP) plasmid], and VEGF plasmid group (transfected with pIRES2-EGFP-VEGF plasmid) according to the random number table, with 6 wells in each group. At post transfection hour (PTH) 24, the expression of green fluorescent protein (GFP) in each group was observed under inverted phase contrast fluorescence microscope, and the expression rate of GFP was detected with flow cytometer. Cells in non-transfection group were tested with the same methods as listed above. The cells in stable transfection in empty vector group and VEGF plasmid group were sifted by neomycin. The mRNA and protein expression levels of VEGF 165 in cells and the protein amount of VEGF 165 in the supernatant of cell culture medium in 3 groups were respectively determined by real-time fluorescent quantitation PCR, Western blotting, and enzyme-linked immunosorbent assay. (2) Forty-eight male nude mice were divided into 4 groups according to the random number table, with 12 mice in each group. Mice in saline group were subcutaneously implanted with dermal substitutes which had been cultured in saline for 2 days on both sides of back (the same site below); mice in medium group were subcutaneously implanted with dermal substitutes which had been cultured in complete medium for 2 days; mice in non-transfected cells group were subcutaneously implanted with dermal substitutes that had been cultured in complete medium with non-transfected HUVECs for 2 days; mice in transfected cells group were subcutaneously implanted with dermal substitutes that had been cultured in complete medium with HUVECs stably transfected with VEGF plasmid for 2 days. The dermal substitutes in every group were taken out on post operation day (POD) 3, 7, 14, and 21. Distributions of microvessels and HUVECs in dermal substitutes were observed by immunohistochemical staining, and the microvessel number was counted on POD 14; the expression level of VEGF 165 protein in dermal substitutes was determined by Western blotting. The experiments were all done in triplicate. Data were processed with one-way analysis of variance and LSD method.

RESULTS(1) Obvious green fluorescence was only observed in the two groups with transfected cells at PTH 24. Expression rates of GFP in the cells of non-transfection group, empty vector group, and VEGF plasmid group were respectively 0, (85.2 ± 3.2) %, and (93.1 ± 2.4) %. In the non-transfection group, empty vector group, and VEGF plasmid group, the relative expression amounts of VEGF 165 mRNA were respectively 1, 1.05 ± 0.09, and 3.02 ± 0.13 (F = 5.28, P < 0.05); the relative expression amounts of VEGF 165 protein were respectively 0.78 ± 0.16, 0.76 ± 0.13, and 1.92 ± 0.18 (F = 7.62, P < 0.05); the protein quantity of VEGF 165 in cell supernatant was respectively (62.4 ± 2.7), (73.1 ± 3.8), (117.5 ± 3.1) pg/mL (F = 15.08, P < 0.05). The mRNA and protein levels of VEGF 165 and VEGF 165 protein amount in supernatant were significantly higher in VEGF plasmid group than in the other two groups, with P values all below 0.05. (2) The number of HUVECs in dermal substitutes of transfected cells group was significantly higher than that of the other three groups on POD 14. The numbers of microvessels of dermal substitutes on POD 14 in saline group, medium group, non-transfected cells group, transfected cells group were respectively 4.2 ± 1.1, 5.2 ± 1.1, 6.6 ± 0.9, 13.8 ± 0.8 per 200 times visual field (F = 17.96, P < 0.01). The microvessel number in transfected cells group was significantly higher than that of the other three groups, with P values all below 0.05. The relative expression ratio of VEGF 165 protein of dermal substitutes in transfected cells group was significantly higher than that in saline group as of POD 7. On POD 14 and 21, the relative expression ratios of VEGF 165 proteins in non-transfected cells group (1.652 ± 0.086, 2.152 ± 0.062) and transfected cells group (2.403 ± 0.091, 2.879 ± 0.047) were significantly higher than those of saline group (1.299 ± 0.027, 1.362 ± 0.103), with P values all below 0.05. And the index level of transfected cells group was significantly higher than that in non-transfected cells group (with P values below 0.05). The VEGF 165 protein content in dermal substitutes increased with time extension in all groups.

CONCLUSIONSTransfection of VEGF 165 gene in HUVEC could effectively facilitate vascularization of dermal substitutes in vivo by high expression of VEGF 165 protein.

Animals ; Cells, Cultured ; Dermis ; blood supply ; Human Umbilical Vein Endothelial Cells ; Humans ; Male ; Mice ; Mice, Nude ; Plasmids ; Transfection ; Vascular Endothelial Growth Factor A ; genetics ; metabolism