Membrane proteins are the main undertakers of biofilm function, and also the most important target group for innovative drug discovery and research. About 60% of drugs targets are membrane proteins. Due to the obvious aggregation and denaturation tendency of membrane proteins in aqueous solution, it is difficult to simulate the membrane like environment to maintain the correct conformation of membrane proteins in vitro, which results in the slower-growing research on the structure and function of membrane proteins and related ligand drugs than that of water-soluble proteins. Membrane protein stabilization technology is the premise of establishing high specificity, high sensitivity and high throughput drug screening methods for membrane protein ligands, which is of great significance. In this paper, some techniques for stable separation and purification of membrane proteins are reviewed, including detergents, artificial membranes, polymers, lentiviral particles and so on, as well as their specific applications in drug screening.

Objective To investigate the effect of different Chinese herbs on cell proliferation and cartilage oligomeric matrix protein(COMP)expression in chondrocyte culture.Methods Chondrocytes isolated from rabbit knee cartilage were cultured for 3 generations with the density of 2?10~4/cm~2 and were verified by collagenⅡimmunohistochemical staining.Rabbit sera containing herbs were obtained after animals orally ad- ministrated herbs at the dosage equivalent to human.At 5% and 10% serum density,cells were cultured in the medium that contained liver-softening herbal compound sera.Subgroups setting at 1,3 and 5 hours after herb intervention were observed.Rabbit and bovine sera were control groups.Seven days after intervention,chon- drocytes proliferation was observed using the MTT assay kit.For the study of COMP expression,chondrocytes were isolated from human knee cartilage supematant.Superuatant COMP level was tested by enzyme-linked immunoabsorbent assays(ELISA)after directly adding compound and extract from liver-softening herbs to the culture at the final concentration of 10 mg/ml for 3 days.Results Liver-softening herbal compound group had significant effect on cell proliferation compared to control,of which,3-hour subgroup was more significant than 1-and 5-hour subgroups(P

Objective To evaluate the application value of X-ray,echocardiogram,pulmonary perfusion scintigraphy,EBCT,Magnetic resonance Pulmonary angiography in diagnosis of PTE.Methods Twenty-five consecutive patients clinically diagnosed of having PTE were examined from july 2003 through March 2004. Patients underwent X-ray chest plain film, echoeardiogram, electronic beam computed tomographie (EBCT)angiography,ventilation-perfusion (V-P)seintigraphy,Magnetic resonance Pulmonary angiography (MRPA)and puhnonary angiography according to a strict diagnostic protocol.Two of the independent readers reviewed the pulmonary angiography and record all of the lobe and segmental involved in PTE and compared with other image method.Results Pulmonary angiography:all of the patients success underwent the technique,the pulmonary artery branch with PTE was in 556 of 775 branches (71.7%). Chest radiography had hints of diagnosis in 12 of 25 patients.Nine patients diagnosed with echocardiogram. Right heart enlargement was in 21,and pulmonary hypertension in 18.V-P scintigraphy revealed 247 segmental involved with PTE of 500 (52.0% ),and the sensitivity was 64.66% compare with the pulmonary angiography.There were 523 pulmonary branches involved PTE with EBCT pulmonary angiograpy of 775 branches,and the sensitivity was 94.06%.MRPA: 8 of 10 patients succeed in the technique, 155 branches of 248 were detected with PTE(62.5% ),the sensitivity was 81.29%.Conclusions EBCT is a high sensitivity method in diagnosis of PTE.Chest radiography and echocardiogram are the first-line modality of PTE.V-P scintigrapby is the valid compensation in diagnosis subsegmental pulmonary artery with PTE when EBCT miss diagnosis.Gd-CE-MRPA may be the second-line modality in diagnosis of PTE.

ABC transporters on the intestinal barrier, blood-brain barrier and on tumor cells will affect drug bioavailability, transport across the blood-brain barrier and multidrug resistance. The active ingredients of traditional Chinese medicines can affect the function and expression of ABC transporters. When combined with pharmaceuticals the potential interaction between the two can change the efficacy of the medicines. We review the ABC transporter superfamily and their distribution with regard to their relationship and interactions with traditional Chinese medicine on the intestinal barrier and the blood-brain barrier, as well as their role in tumor multidrug resistance mediated by ABC transporters. We summarize the research progress over the past five years.

OBJECTIVETo establish a reliable model for drug screening and therapy by culturing rat femoral head and inducing cartilage degeneration quickly in vitro.

METHODSThe femoral heads from the same SD rats of two-month old were divided into control group and experimental group respectively. They were cultured with DMEM medium plus 10% fetal bovine serum or DMEM medium plus 10% fetal bovine serum plus 50 ng/ml IL-1β for three days. Femoral heads were fixed in 4% paraformaldehyde, decalcified, dehydrated, embedded in paraffin and cut into slices. Specimens were stained with Toluidine blue and Safranine O-Fast Green FCF. The protein expression levels of type II collagen, MMP13, Sox9 and ADAMTS5 were analyzed by immunofluorescence.

RESULTSBoth the Toluidine blue and Safranine O staining were pale in the margin of femoral heads which were stimulated with IL-1β for three days compared to that in control group. The Fast Green FCF staining was positive at the edge of the femoral head in experimental group, which indicated that cartilage became degenerated. The expression levels of both type H collagen and Sox9 were decreased significantly while the expression levels of MMP13 and ADAMTS5 were increased in experimental group.

CONCLUSIONThe model of cartilage degeneration is established by culturing and inducing the degeneration of the femoral heads quickly in vitro.

Animals ; Cartilage Diseases ; genetics ; metabolism ; Collagen Type II ; genetics ; metabolism ; Disease Models, Animal ; Femur Head ; metabolism ; Humans ; In Vitro Techniques ; Interleukin-1beta ; genetics ; metabolism ; Male ; Matrix Metalloproteinase 13 ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley ; SOX9 Transcription Factor ; genetics ; metabolism

Objective To develop a new type of individual first aid kit for plateau conditions to treat acute high altitude reaction.Methods The theories of military preventive medicine as well as the practices of plateau medical protection and treatment were involved in the design and manufacturing of the kit with emphases on kit body architecture design and selection of equipped medicinal materials.Totally 2 100 servicemen had 1 000 ones divided into a control group and 1 100 ones into an observation group.The two groups both underwent conventional treatment,while the observation group applied the kit besides.The incidence rates and curative effects of acute high altitude reaction were compared in the two groups.Results The kit improved first aid of acute high altitude reaction in time consumed and speed.One week after entering the plateau the scores of headache,gastrointestinal symptom,fatigue and dizziness were (1.5±0.5),(1.7±0.4),(1.3±0.6) and (1.6±0.7) respectively in the observation group,and (2.3±0.6),(2.4±0.5),(2.2±0.5) and (2.4±0.6) respectively in the control group.One month after entering the plateau,the observation group had the recovery rate being 99.2％ and the mean therapy time being (8.1±3.3)d,and the control group had the recovery rate being 91.2％ and the mean therapy time being (15.2±6.4)d.The observation group behaved better than the control group in disease grading,therapy time,recovery rate significantly (P＜0.05).Conclusion The kit gains advantages in convenience,portability and curative effect when used to treat acute high altitude reaction,and thus is worthy promoting practically.

Objective To survey the distribution and amibiotie resistance of bacterium isolated from hospitalized patients with hematolegical diseases.Methods Bacterial strains were isolated from patients with hematological disease.Antimicrobial susceptibility testing was done by the method of minimum inhibitory concentration.Results A total of 56 bacterial strains were isolated from all kinds of specimens,including blood,phlegm and urine.14.3% were gram-positive and 85.7% were gram-negative bacterium.Escheriehia coli and pseudomonas aeruginosa account for 21.4% of gram-negative bacterium,respectively.Detection rates of ESBLs in E.coli and K.pneumoniae were 75.0%(8/12)and 33.3%(2/6),respectively.Conclusion Our data may have great significance for the empirical use of antimicrobial agents in the treatment of infections in patients with hematological disease.

OBJECTIVETo investigate regulation function of anodonta glucan HBP-A on chondrocytes through Wnt pathway in vitro.

METHODSRat chondrocytes were cultured and differentiated induced with IL-1beta (10 ng/ml) in vitro. Chondrocytes were divided into five groups:IL-13 group,IL-1beta + IWP-2 (5 microM,Wnt pathway inhibitor) group, IL-1beta + HBP-A (0.3 mg/ml) group and IL-1beta + IWP-2 + HBP-A group. Wnt-3a, beta-catenin (24 h,48 h,72 h) and MMP-13(72 h) genes expression were detected by Rt-PCR, while beta-catenin, MMP-13, Sox-9 and coll-II (48 h) protein expression were measured by Western-blot.

RESULTSAfter induction of IL-1beta, gene expression of Wnt-3a, beta-catenin and MMP-13 were increased,so were the protein expression of beta-catenin and MMP-13. In contrast,protein expression of Sox-9 and Coll-II were declined. Following addition of HBP-A, Wnt-3a, beta-catenin and MMP-13 were shown as induction of IL-1beta, but protein expression of Sox-9 and Coll-II were upgraded. Combining HBP-A with IWP-2 led to the lowest level in Wnt-3a, beta-catenin gene and beta-catenin protein expression and highest expression of Sox-9 protein.

CONCLUSIONHBP-A could not only delay the differentiation of chondrocytes through downgrading the signal expression of Wnt/beta-catenin,but also adjust the expression of Wnt-3a, beta-catenin and Sox-9 when combinated with the Wnt inhibitor.

Animals ; Anodonta ; chemistry ; Cell Differentiation ; drug effects ; Cells, Cultured ; Chondrocytes ; cytology ; drug effects ; metabolism ; Glucans ; pharmacology ; Interleukin-1beta ; metabolism ; Rats ; Wnt Signaling Pathway ; drug effects ; Wnt3A Protein ; genetics ; metabolism ; beta Catenin ; metabolism

OBJECTIVETo construct eukaryotic expression plasmids containing green fluorescent protein gene and CYP19 wild-type or its variants (W39R, R264C, W39R-R264C) and to observe its expression in MCF-7 and Bcap-37 cells.

METHODSThe aromatase WT cDNA sequence was obtained by RT-PCR amplification and cloned into the eukaryotic expression vector pcDNA3.1(+). pcDNA3.1(+)-CYP19-GFP plasmid was then used as the template for site-directed mutation to create variant constructs (W39R, R264C, W39R-R264C). pcDNA3.1(+)-CYP19-GFP was transfected and expressed in MCF-7 and Bcap-37 cells.

RESULTThe construction of pcDNA3.1(+)-CYP19-GFP plasmid was confirmed by enzyme digestion and DNA sequencing. pcDNA3.1(+)-CYP19(W39R)-GFP, pcDNA3.1(+)-CYP19(R264C)-GFP, pcDNA3.1(+)- CYP19(W39R-R264C)-GFP plasmids were confirmed by DNA sequencing. The MCF-7 and Bcap-37 cells transfected with the pcDNA3.1(+)-CYP19-GFP plasmid expressed reporter gene of GFP.

CONCLUSIONThe eukaryotic expression plasmids have been constructed and expressed in MCF-7 and Bcap-37 cells successfully, which lays the foundation for the research of biological activities of CYP19 variant allozymes.

Aromatase ; genetics ; Cell Line, Tumor ; Genetic Vectors ; Green Fluorescent Proteins ; genetics ; Humans ; Mutagenesis, Site-Directed ; Plasmids ; genetics ; Recombinant Fusion Proteins ; genetics ; Transfection

OBJECTIVETo study the chemical constituents and activity of total flavonoids contained in Yushen Tang,in order to lay a foundation for defining active consituents of the prescription and their mechanism.

METHODThe activity of total flavonoids contained in Yushen Tang were evaluated by in vitro H2O2-induced human umbilical vein endothelial cell injury experiment. The chemical constituents were separated and purified by such methods as silica column chromatography, macroporous resin chromatography, sephadex and HPLC preparation,and their structures were identified on the basis of their spectral data and physicochemical properties.

RESULTTotal flavonoids contained in Yushen Tang showed the effects in inhibiting hydrogen peroxide-induced human umbilical vein endothelial cell (HUVEC) injury and down-regulating PAI-1 expression (P<0.05). Nine flavonoids were separated from total flavonoids contained in Yushen Tang and identified as calycosin (1), linarin (2), 3',4',7 -trihydroxyflavone-7-O-beta-D-glucopyranoside (3), 7,3'-dihydroxy-4'-methoxyflavone-7-glucoside (4), quercetin (5), kaempferol (6), rutin (7), pratensein-7-O-beta-D-glucoside (8) and 7-O-glucosyl liquiritigenin (9).

CONCLUSIONTotal flavonoids contained in Yushen Tang showed the effect in inhibiting HUVEC injury. All of the nine flavonoids were separated from Yushen Tang for the first time, and compounds 1,3,4,8,9 may be derived from astragalus mongholicus, while compounds 4,5-7 may be derived from herba cepbalanoplosis segeti and hairyvein agrimony. Among them, compounds 3,4 and 9 were seperated from single ingredient of the prescription for the first time.

Animals ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; Flavonoids ; chemistry ; pharmacology ; Gene Expression ; drug effects ; Human Umbilical Vein Endothelial Cells ; Humans ; Kidney Diseases ; drug therapy ; genetics ; metabolism ; Male ; Molecular Structure ; Plasminogen Activator Inhibitor 1 ; genetics ; metabolism ; Rats ; Rats, Wistar