Objective To construct pGL4.14-1uc eukaryotic expression vector for HLA-B27 promoter gene and explore the activity regulation of this promoter in Hela cells.Methods The HLA-B27 gene promot- er(-419 bp～1 bp)was amplified by polymerase chain reaction and was cloned into pGL4.14-luc vector to construct eukaryotic expression vector pGL4.14/B27 pro-luc.The purified pGL4.14/B27 pro-luc was stablely transfected into HeLa cells and the activity of HLA-B27 gene promoter was detected by luminometer.Results About 432 bp gene fragment was amplified by PCR from genomie DNA and pGL4.14/B27 pro-luc vector was constructed successfully.The activity of HLA-B27 promoter was 1.67?0.20,1.79?0.71,2.94?0.68,1.98?0.45 in Hela stable cells after treated with TNF-?,IFN-?,IFN-?and IFN-?for 48 hours.Conclusion TNF-?. IFN-?,IFN-?and IFN-?can regulate the B27 promoter activity.The high specific activity of constructed HLA-B27 promoter eukaryotic expression vector may be a good method for further research.

Traditional processes are mostly adopted in traditional Chinese medicine (TCM) preparation production and the quality of products is mostly controlled by terminal. Potential problems of the production in the process are unpredictable and is relied on experience in most cases. Therefore, it is hard to find the key points affecting the preparation process and quality control. A pattern of research and development of traditional Chinese medicine preparation process based on the idea of Quality by Design (QbD) was proposed after introducing the latest research achievement. Basic theories of micromeritics and rheology were used to characterize the physical property of TCM raw material. TCM preparation process was designed in a more scientific and rational way by studying the correlation among enhancing physical property of raw material, preparation process and product quality of preparation. So factors affecting the quality of TCM production would be found out and problems that might occur in the pilot process could be predicted. It would be a foundation for the R&D and production of TCM preparation as well as support for the "process control" of TCMIs gradually realized in the future.
Drug Compounding ; methods ; standards ; Drugs, Chinese Herbal ; chemistry ; isolation & purification ; standards ; Humans ; Medicine, Chinese Traditional ; standards ; trends ; Quality Control ; Research ; standards ; trends ; Research Design ; standards ; Technology, Pharmaceutical ; instrumentation ; methods ; standards

Twelve compounds were isolated from the herb of Chenopodium ambrosioides, and their structures were identified by spectroscopic methods as kaempferol-7-O-alpha-L-rhamnopyranoside (1), kaempferol-3,7-di-O-alpha-L-rhamnopyranoside (2), patuletin (3), quercetin-7-O-alpha-L-rhamnopyranoside (4), grasshopper ketone (5), 4-hydroxy-4-methyl-2-cyclohexen-1-one (6), syringaresinol (7), benzyl beta-D-glucopyranoside (8), dendranthemoside B (9), N-trans-feruloyl tyramine (10), N-trans-feruloyl 4'-O-methyldopamine (11), and 4-hydroxy-N-[2-(4-hydroxyphenyl) ethyl] benzamide (12). Among them,compounds 3, 6-8,10, and 12 were isolated from the genus Chenopodium for the first time, and compounds 2-12 were isolated from this plant for the first time.
Chenopodium ambrosioides ; chemistry ; Drugs, Chinese Herbal ; chemistry ; isolation & purification

Objective:To investigate the expression of Bcl-2 and Bax in renal tissues of patients with hepatitis B virus- associated glomerulonephritis(HBV-GN).Methods:Twenty HBV-GN specimens with complete nephrology data and 10 normal renal specimens were randomly chosen for the present study.Cell apoptosis was detected by means of terminal deoxynucleotidyl transferase mediated d-UTP nick end labeling(TUNEL)and the apoptotic index was calculated;immunohistochemistry was used to detect the protein expression of Bax and Bcl-2.ResuLts:The apoptotic index in HBV-GN group was obviously higher than that of the control group;the apoptotic cells were mainly distributed in the proximal and distal renal tubules and the collecting duct epithelial cells,seldom seen in the glomerular cells.The expression of Bcl-2 in HBV-GN patients was predominately present in the renal tubular epithelia cells(positive in the plasma,membrane and nuclear);the expression of Bax was found in both glomerular cells and renal tubular cells,mainly in tubular epithelial cells,seldom seen in Bowman's capsule or glomerular mesangial region.Conclusion:Apoptosis in the kidney of HBV-GN patients mainly occurs in the renal tubular epithelial cells;expression of Bax and Bcl-2 is mainly in the renal tubular epithelial cells,suggesting that the injury of tubular interstitial damage may be one of the important factors for the development of HBV-GN.

Background Q-value is used to express the asphricity of the anterior corneal surface.As a optical surface,the optial morphology of cornea is composed of a series of tangential sections that cut-off through the optical axial.Therefore,tangential section could represent the true optical morphology of the corneal surface,and Q-value calculation by tangential radius can better represent the asphericity.Objective This study was to calculate Q-value of meridian section of the anterior corneal surface by tangential radius of the tangential topography and analyze the corneal asphericity in horizontal interval.Methods Ninety right eyes of 90 myopia subjects aged 16-30years and with mean spherical equivalent of (-5.45 ± 2.75) D received corneal topography examination using Orbscan Ⅱ system.The subjects were assigned to low myopia group,moderate myopia group and high myopia group based on their diopter and 30 eyes for each group.The tangential curvatures on meridian section at a 0.1 mm interval from apex to periphery of the anterior surface were accepted and the Q value of the semimeridian section was calculated by linear regression mathematical formulas of the tangential radius.Mydriatic optometry,intraocular pressure,keratometer and fundus examinations were performed on the subjects.Written informed consent was obtained from each subject before any medical examination.Results The coefficients of determination in all the semimeridians were over 0.5.The average calculated Q-values in the nasal and temporal horizontal interval were -0.32±0.11 and-0.30±0.12,with a significant difference between them (t =2.009,P＜0.05).The vertex radius of curvature was (7.78±0.27)mm and (7.72±0.25)mm respectively in the nasal and temporal horizontal interval,showing a significant difference(t=-1.016,P＞0.05).No significant difference was seen in Q values of both nasal and temporal areas among three myopic groups (nasal:F =0.192,P =0.825 ; temporal:F =0.912,P =0.406).The average Q value of the nasal and temporal principal meridian was-0.33±0.14 and-0.30±0.13 respectively,and the r0 was 7.76±0.30 and 7.74±0.24 respectively.A weak positive correlation was found between r0 and mean Q value of nasal meridian (r=0.320,P＜0.05),but no significant correlation was found between r0 and mean Q-value of temporal meridian (r=0.104,P＞0.05).No significant differenees were seen in the Q values between nasal meridian and nasal zone (t=0.349,P＞0.05) as well as between temporal meridian and temporal zone(t=-0.373,P＞0.05).Conclusions The study analyzes the calculated Q-value of the semimeridian section in borizontal area with myopia by linear regression mathematical formulas of tangential radius on tangential topography.The anterior surface of the cornea is proved to be prolate ellipse in shape in the subjects with myopia.

Fluorescence differential display (FDD) technique was used to identify genes that are specifically or preferentially expressed in different developmental stages of cotton fiber cells. One hundred and nine differentially displayed cDNA fragments were isolated using 9, 21 and 27 DPA (days postanthesis) fibers as experimental materials. By a combination of two rounds of reverse Northern hybridization and Northern blot analyses, a number of such cDNA fragments were proved to represent fiber-specific/preferential genes. Sequencing determination and database searching indicated that most of these genes are novel. This work is an important step towards cloning the full-length cDNAs and characterizing the cellular functions of aforementioned genes in fiber development.
Blotting, Northern ; Cotton Fiber ; Fluorescence ; Gene Expression Profiling ; methods ; Gossypium ; genetics ; growth & development ; Polymerase Chain Reaction

OBJECTIVELandmark trials have demonstrated that statins can reduce the risk of coronary events. Despite the widespread use of statins in the settings of primary and secondary prevention of CHD, withdrawal of statins is a frequent problem in clinical practice. Several recent clinical studies have suggested that withdrawal of statin therapy might be associated with an increase in thrombotic vascular events and the onset of acute coronary syndromes. However, the effects of discontinuing of statins treatment on endothelial function and underlying mechanism are unknown. Objectives We investigated the effects after withdrawal of simvastatin on brachial artery endothelial function in patients unreached cholesterol target with coronary heart disease (CHD) or CHD risk factors.

METHODSWe included 33 patients with established CHD or CHD risk factors, whose serum cholesterol did not achieve NCEP target level. They were administered simvastatin (20 mg) for 4 weeks. Endothelial dependent flow-mediated vasodilation (FMD) was assessed in the brachial artery using high-resolution ultrasound at baseline, after 4 weeks of simvastatin and after termination of therapy 1 week. We evaluated fasting serum lipid profiles and vasoactive substances simultaneously, included nitric oxide (NO), endothelin (ET), 6-keto-PGF1(alpha) and thromboxane B(2) (TXB(2)), which were measured as plasma prostacyclin and TXA(2) respectively.

RESULTSSimvastatin treatment reduced low density lipoprotein cholesterol (LDL-C) and total cholesterol (TC) levels and improved endothelial-dependent vasodilation in patients after 4 weeks. Withdrawal of simvastatin, however, FMD showed a significant reduction [(4.82 +/- 0.71)% vs (11.51 +/- 0.87)%, P < 0.01], that remained in low level after 1 week, and the FMD were even lower than the baseline values [(4.82 +/- 0.71)% vs (5.89 +/- 0.65)%, P < 0.01]. After terminating simvastatin treatment, serum NO and plasma 6-keto-PGF1(alpha) levels decreased, as well as plasma ET and serum LDL-C levels increased. But there was no significant difference between plasma TXB(2) levels before and after withdrawal of simvastatin (P > 0.05). Overall, there were significant positive correlations between withdrawal-induced changes in FMD and serum NO level (r = 0.674, P = 0.004), whereas no correlations were shown between the changes in FMD and serum LDL-C level (r = -0.414, P = 0.083).

CONCLUSIONSAbrupt withdrawal of simvastatin therapy resulted in the significant adverse impact on brachial artery endothelial function in patients unreached cholesterol target with CHD or CHD risk factors. Termination of therapy may suppress endothelial NO production and impair endothelial function that is independent of lipid-lowering effect.

Aged ; Brachial Artery ; drug effects ; Cholesterol, LDL ; blood ; Coronary Disease ; drug therapy ; physiopathology ; Endothelium, Vascular ; physiopathology ; Female ; Humans ; Hypolipidemic Agents ; administration & dosage ; Male ; Middle Aged ; Nitric Oxide ; blood ; Risk Factors ; Simvastatin ; administration & dosage ; Vasodilation

To identify the structure of three related substances in potassium sodium dehydroandrographolide succinate (PSDS), an HPLC preparation method was used to separate the impurities. These main impurities were identified using LC-ESI/TOFMS, LC-ESI/MSn, NMR, UV and IR. One of the main impurities was a hydrolyzed and oxidized product of PSDS, which has not been reported previouely. The other two impurities were hydrolyzed products of PSDS after losing different succinic acids. The results indicate that PSDS can be easily hydrolyzed and oxidized. It should be stored at cool and dry places.
Andrographis ; chemistry ; Antiviral Agents ; chemistry ; isolation & purification ; Chromatography, High Pressure Liquid ; Diterpenes ; chemistry ; isolation & purification ; Drug Contamination ; Drugs, Chinese Herbal ; chemistry ; isolation & purification ; Molecular Structure ; Plants, Medicinal ; chemistry ; Spectrometry, Mass, Electrospray Ionization ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

OBJECTIVETo compare indirect immunofluorescence assay (IIFA) and enzyme-linked immunosorbent assay (ELISA) for detecting antinuclear antibodies (ANA) and anti-double-stranded DNA antibodies (anti-dsDNA).

METHODSA total of 125 serum samples were obtained from patients with established or suspected autoimmune disease, and 82 samples were used for ANA detection and 57 for anti-dsDNA detection using both IIFA and ELISA. Fourteen samples were examined for both ANA and anti-dsDNA. In cases where discrepancy occurred in the results by the two methods, extractable nuclear antigens were detected using immunoblotting.

RESULTSThe positivity rate of ANA detected by IIFA and ELISA was significantly different (87.8% and 73.17%, respectively, P<0.01), but the positivity rate of anti-dsDNA was similar between IIFA and ELISA (77.19% and 71.93%, respectively, P>0.05). The percent agreement between the two testing methods with different cutoff values of ANA and anti-dsDNA showed significant differences (P<0.01), and for some uncommon patterns, the percent agreement of the two methods was lowered in ANA detection but remained unchanged for anti-dsDNA with different ANA patterns. High percent agreements of the two methods were obtained with the cutoff ANA titer of 1:100 and the cutoff anti-dsDNA value of weak positivity, but they demonstrated a significant difference in testing low-titer ANA and anti-dsDNA.

CONCLUSIONIIFA is more sensitive than ELISA in detecting the total ANA and anti-dsDNA. ELISA prescreening combined with IIFA can obtain the information of the nuclear pattern and allow the observation of the titer alterations. The combination of two or more testing methods can greatly enhance the accuracy of the results.

Antibodies, Antinuclear ; analysis ; DNA ; immunology ; Enzyme-Linked Immunosorbent Assay ; Fluorescent Antibody Technique, Indirect ; Humans

OBJECTIVETo investigate the relationship between the genotypes of hepatitis B virus and the clinical and liver pathological features of patients with chronic hepatitis in the Zhoushan Islands.

METHODSOne hundred eighty HBV DNA positive chronic hepatitis patients with HBV markers were enrolled in this study. They were at least second generation Zhoushan Island residents. One hundred forty-seven of them were males and 33 were females with an average age of 39.0+/-11.3. Among the 180 patients, 17 had ASC, 57 had mild CHB, 48 moderate CHB, 9 severe CHB, 6 SHB, 39 LC, and 4 had HCC. The genotypes of their serum HBV were detected by using PCR integrated with Tagman MGB probe technology, and their serum HBV markers, HBV DNA and liver functions were also examined. Out of 180 patients, 129 accepted a liver biopsy. A pathological evaluation was then performed.

RESULTSHBVs of genotype C, 135 cases (75.0%), of B, 40 cases (22.2%), and of B+C, 5 cases (2.8%) were found among these 180 patients. No genotype A or D HBV were found. The proportions of genotype C virus were 7/17, 86/114, 34/39, 6/6 in ASC, CHB, LC and SHB patients. In the hepatocellular carcinoma patients, there were 2 each of genotype B and C. Among the 99 patients with genotype C HBV, 84 cases (84.8%) showed moderate and severe inflammation histologically in their livers and among the 30 patients with B, 7 cases (23.3%) showed moderate to severe inflammation in their livers (z = 6.47, P less than 0.01). The proportion of genotype C HBV was significantly different from that of genotype B HBV in those that showed moderate and severe (S3-4) liver fibrosis. In patients infected with genotype C HBV who had moderate and severe liver pathological changes, their clinical manifestations reflected better the histological alterations of their livers.

CONCLUSIONGenotypes C, B and B+C HBV were found in CHB patients in the Zhoushan Islands of China, and type C was the predominant one. The liver pathological damage level of genotype C HBV infected patients is more serious than that of genotype B.

Adult ; China ; epidemiology ; DNA, Viral ; genetics ; Female ; Genome, Viral ; Genotype ; Hepatitis B virus ; classification ; genetics ; Hepatitis B, Chronic ; epidemiology ; pathology ; Humans ; Liver ; pathology ; Male ; Middle Aged