Objective: To investigate the differential expression of serum soluble TNF-related apoptosis-inducing ligand (sTRAIL) between ankylosing spondylitis (AS) and rheumatoid arthritis patients (RA) and to discuss its clinical significance. Methods: Sixty AS patients, including 38 HLA-B27-positive ones and 22 HLA-B27-negative ones, 20 rheumatoid arthritis (RA) patients and 30 healthy individuals were included in the present study. The AS patients were divided into active group and inactive group based on bath ankylosing spondylitis disease activity index (BASDAI). The concentrations of serum sTRAIL were measured by ELISA in all groups. Erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) were detected automatically by ESR automatic analyzer and specific protein analyzer. Results: The serum sTRAIL concentration was significantly higher in AS patients (both HLA-B27-positive and -negative AS patients) than in RA patients and healthy controls (P<0.01); no significant difference was found between HLA-B27-positive and -negative AS patients. Serum sTRAIL concentration was significantly higher in active AS group than in inactive AS group (P<0.01). Serum sTRAIL and CRP concentrations were correlated with each other in HLA-B27 positive AS patients (r = 0.609, P = 0.000), but not in HLA-B27-negative ones. Serum sTRAIL concentration was not correlated with ESR in AS patients. Conclusion: Serum sTRAIL is obviously up-regulated in AS patients, which is not associated with the status of HLA-B27. However, the association between sTRAIL and CRP is influenced by the status of HLA-B27.

Objective To establish a specific fluorescence quantitative method for determining the mRNA expression of Toll-like receptor3(TLR3)in peripheral blood mononuclear cells(PBMCs).Methods Using the Beacon Designer 2.1 software,specific primers and Taqman-MGB probe were designed.The plasmid pMD18-T-TLR3 was constructed as calibrator and the amplified fragment was obtained by reverse- transcript-PCR(RT-PCR).RNA quantification based on cycle threshold values(Ct)was used to establish the standard curve.According to which,the TLR3 mRNA levels in 30 normal individuals,20 patients with primary biliary cirrhosis(PBC)and 20 ones with chronic liver cirrhosis induced by HBV were calculated automatically by software after the fluorescence of PCR product was detected continuously during amplification.Results The linear detection range of the assay for TLR3 gene and ?-actin was 10~2-10~8(r= -0.9974)and 10~3～10~8(r=-0.9984),respectively.The coefficient of variation of both intra-and inter- assay reproducibility for high concentration sample were 6.7% and 8.7%,respectively,and those for low concentration sample were 12.3% and 14.0%.The TLR3 mRNA expression level ranges from 3.46?10~2- 4.51?10~3 copies/?g RNA,4.92?10~2-1.42?10~4 copies/?g RNA and 2.58?10~2-7.17?10~3 copies/?g RNA for 30 healthy individuals,20 PBC patients and 20 ones with chronic liver cirrhosis induced by HBV, respectively.Conclusion We have successfully set up a FQ-RT-PCR method for detecting TLR3 mRNA, which may be used as an excellent tool for the clinic and basic study on the expression of TLR3 gene.

The leakage of genetically engineered microorganism(GEM) at the beginning of operation was the important content for ecological risk assessment when GEM was inoculated in a membrane bioreactor(MBR) for bioaugmentation. The effects of different operating conditions on leaking density and intercepting efficiency of GEM were investigated in a submerged microfiltration MBR at the beginning of operation. The interception characteristics were also discussed. The results showed that different operating conditions had different influences on intercepting efficiency:higher sludge concentration was profit for interception,while higher aeration intensity and flux had opposite effects on interception. When the inoculated density of GEM was 1.0?1010 CFU/mL,the leaking densities varied from 1.0?102 CFU/mL to 2.5?102 CFU/mL under different operating conditions at the beginning of operation and the maximum intercepting efficiency could be higher than 8 lg. The main factors determining intercepting efficiency at the beginning of operation were membrane module interception,sludge adsorption as well as suspended GEM transfer inhibition and their contribution shares under certain conditions were 82.3%,14.9% and 2.8%,respectively. Gel layer formation during MBR stable operation was helpful to increase intercepting efficiency. The contribution shares for GEM interception of membrane module,sludge and gel layer were 75.3%,10.7% and 14.0%,respectively,under certain conditions.

According to the characteristics of sludge samples from full-scale wastewater treatment bioreac-tors, the essential total DNA extraction method for most environmental samples, lysozyme-SDS-phenol/ chloroform method, was modified to improve sample pretreatment, intensify cell lysis and enhance the effi-ciency of impurity removal. Obtain a general total DNA extraction method for industrial sludge samples. Such a method was applied for total DNA extraction of sludge samples from several running full-scale an- aerobic or aerobic bioreactors in Shijiazhuang, China. The results indicated that the modified method was suitable for all the sludge samples in this study, showing the satisfying generality. The extracted total DNA of all sludge samples were pure, with about 1.8 of A260/ A280 ratio. The method was also efficient; with average total DNA yield of over 0.7 mg/g and maximum yield of 0.85 mg/g. Moreover, all the extracted to- tal DNA samples could serve as templates directly to amplify 16S rDNA by PCR. The PCR products could be separated well by denaturing gradient gel electrophoresis (DGGE) and the DGGE band patterns were clear enough to be used for further analysis. All these facts indicated that the total DNA extraction method provided in this study could meet the requirements of sludge samples research, from full-scale wastewater treatment bioreactors, using molecular biology technologies.

Derris eriocarpa, a traditional Chinese medicine belonging to the family of Leguminosae, is widely distributed mainly over Yunnan, Guangxi and Guizhou of China. Modern pharmacological researches on this herb showed that it had extensive bioactivities, such as promoting urination, removing dampness and cough and reducing inspissated mucus and other biological activities. The extensive studies on the chemical constituents of this plant have resulted in the isolation of triterpenoids, steroids, fatty acid and others, but the flavone compounds haven't reported before. In our further research on the ethyl acetate of this plant, nine flavone compounds were obtained by column chromatography on silica gel, Sephadex LH-20, semi-prep HPLC, polyamide column chromatography and recrystallization for separation and purification. The structures were determined on the basis of extensive spectroscopic analysis, including MS, NMR experiments and comparison with spectroscopic data in the literature, respectively, as diosmetin (1), 3, 3'-di-O-methylquercetin (2), afromosin (3), 6, 3'-dihydroxy-7, 4'-dimethoxyisoflavone (4), odoratin (5), 7, 3'-dihydroxy-8, 4'-dimethoxyisoflavone (6), 6, 4'-dihydroxy-7, 3'-dimethoxyisoflavone (7), 5, 7, 4'-trihydroxy-3, 3', 5'-trimethoxyflavone (8), and alpinumisoflavone (9). All these compounds were isolated from Derris eriocarpa How for the first time. And the in vitro assays showed that compound 2 possessed moderate inhibitory activity against human cancer cells K562 and HEL.
Derris ; chemistry ; Flavonoids ; chemistry ; isolation & purification ; pharmacology ; Humans ; K562 Cells

Clinical Laboratory Techniques ; Hepatitis, Autoimmune ; diagnosis ; Humans

OBJECTIVETo apply fluorescent real-time reverse transcriptase-polymerase chain reaction (RT-PCR) in detecting influenza viruses.

METHODSA total of 207 oral swab samples were obtained in 16 collections from SARS patients and suspected influenza outbreak cases. They were subjected to influenza virus detection by fluorescent real-time RT-PCR, MDCK cell culture, and hemagglutinin inhibition assay.

RESULTSOut of 207 samples, 79 (38.16%) were positive for influenza viruses when tested by fluorescent real-time PCR, and 62 (29.95%) positive when tested by MDCK cell culture. There was a statistically significant difference between them (chi square=8.64, P less than 0.005). From 104 cases in 9 collections dual serum samples were obtainable. When tested with hemagglutinin inhibition assay, 64 cases (61.54%) showed a 4-fold increase against H3N2 antigen.

CONCLUSIONThis study showed that fluorescent real-time PCR is a reliable, sensitive, and fast method for detecting influenza viruses.

Cell Culture Techniques ; Humans ; Influenza A Virus, H3N2 Subtype ; isolation & purification ; Influenza A virus ; isolation & purification ; Influenza, Human ; virology ; Reverse Transcriptase Polymerase Chain Reaction ; methods ; Sensitivity and Specificity ; Severe Acute Respiratory Syndrome ; virology

Fragments encoding amino acids 76-130 in the linear conserved region (LCR) of A/Hubei/1/2010 (H5N1) HA2 was fused to hepatitis B core antigen (HBc) to generate a LCR-HBe virus-like particle (VLP). Results showed that the fusion protein of LCR-HBc was highly expressed in this prokaryotic expression system. The purified LCR-HBc particle stimulated high levels of IgG production in mice with a titer of > 1:12 800, and provided 50% cross-protection against lethal challenge by H1N1 viruses.
Amino Acid Sequence ; Animals ; Female ; Hemagglutinin Glycoproteins, Influenza Virus ; immunology ; Influenza A Virus, H5N1 Subtype ; immunology ; Influenza Vaccines ; immunology ; Interferon-gamma ; biosynthesis ; Lung ; pathology ; Mice ; Mice, Inbred BALB C ; Molecular Sequence Data

OBJECTIVETo investigate the expression of the interferon-induced transmembrane-1 (IFITM1) gene in colorectal cancer (CRC) tissue and the serum anti-IFITM1 antibody responses of the patients and assess their value in clinical diagnosis of CRC.

METHODSSemi-quantitative RT-PCR was performed to detect IFITM1 mRNA expression in the specimens of normal colonic mucosa, CRC tissue, inflammatory polyps, adenomatous polyps, gastric cancer, esophageal carcinoma and liver cancer tissues. Serum samples were collected from the patients to detect anti-IFITM1 antibody responses using Western blotting. The clinicopathological features of the carcinoma expressing IFITM1 gene were analyzed.

RESULTSIFITM1 mRNA was expressed in 47.4 % (18/38) of the CRC specimens, a rate significantly higher than that in adenomatous polyps [15% (3/20)] and gastric cancer [4.8% (1/21)]; no obvious IFITM1 expression was found in normal colonic mucosa, inflammatory polyp, esophageal carcinoma or liver cancer tissues (P<0.001 or P<0.05). IFITM1 mRNA was strongly expressed in CRC at the expression level of 0.8048-/+0.2273, which was significantly higher than that in adenomatous polyps (0.4447-/+0.0989, P<0.001). No anti-IFITM1 antibody response was detected in healthy human sera, but in the CRC patients, the serum antibody response was detected at the rate of 36.8% (14/38), significantly higher than the rate of 9.5% (2/21) in gastric cancer (P<0.05). No antibody response was detected in esophageal carcinoma, liver cancer, inflammatory polyp or adenomatous polyps. Most of the IFITM1-expressing CRC had a diameter exceeding 5 cm, often invading the serous membrane with metastasis to the lymph nodes and the distant organs; these tumors were identified mostly as well-differentiated adenocarcinoma in Dukes stage C or D.

CONCLUSIONIFITM1 gene may play an important role in the pathogenesis, development and metastasis of CRC, and may serve as a potential biomarker for clinical diagnosis of CRC.

Antibodies ; blood ; Antigens, Differentiation ; Biomarkers, Tumor ; genetics ; metabolism ; Colorectal Neoplasms ; diagnosis ; genetics ; Humans ; Membrane Proteins ; genetics ; metabolism ; RNA, Messenger ; genetics ; immunology ; metabolism

OBJECTIVE: To investigate the relationship among 3 polymorphisms of GP IIb and the function of GP IIb T13959 G in the platelet transfusion refractoriness(PTR). METHODS: The 26th exon, the 30th exon and the 21st intron of gene GP IIb in 110 individuals were amplified by polymerase chain reaction (PCR), and the PCR products were analyzed with single-strand conformation polymorphism(SSCP) and sequenced to investigate whether there was linkage among the polymorphisms of the gene. Human platelet antigen-3 (HPA-3) gene frequency was detected by Fok I enzyme in 147 patients with hematologic diseases, and was compared with that in 110 normal individuals. Forty-four patients who received apheresis platelet transfusion repeatedly were randomly divided into the HPA-3 homotype group and the control group. The antibodies of the platelet were detected after 3 times of platelet transfusion. RESULTS: There were polymorphisms of gene GP IIb in the 26th, 30th exon and the 21st intron, and the mutations were: T changed into G in 13,959 th of the 26th exon; C changed into T in 16,997 th of the 30th exon; the 9 bps deletion occurred in 11,996-12,004 th of the 21st intron. The 3 polymorphisms had synchronization in the individuals. The results of Fok I enzyme indicated that the frequency of HPA-3a was 83.6% (92/110)and 81.9%(119/147), and that of HPA-3b was 16.4%(18/110) and 19.1%(28/147) in the normal individuals and the patients respectively. There was no significant difference between the patients and normal individuals (P>0.05). After the platelet transfusion, the antibodies of all the cases of the homotype platelet transfusion were negative, but the antibodies in 2 cases of the control group were positive, and there was antibody to HPA-3a in one of the antibodies positive cases. CONCLUSION (1)There is close linkage among the polymorphisms of gene GP IIb, which is T->G in 13 959 th of the 26th exon, C->T in 16,997 th of the 30th exon, and the 9 bps deletion in 11,996-12,004 th in the 21st intron. (2)The gene frequency of HPA-3a/3b is similar in the normal individuals and patients with hematologic diseases. (3) HPA-3 system may be one of the reasons for PTR in Chinese.
Adolescent ; Adult ; Aged ; Antigens, Human Platelet ; immunology ; physiology ; Asian Continental Ancestry Group ; genetics ; Case-Control Studies ; Child ; Exons ; Female ; Gene Frequency ; Genotype ; Humans ; Immune Tolerance ; Introns ; Male ; Middle Aged ; Platelet Membrane Glycoprotein IIb ; genetics ; immunology ; Platelet Transfusion ; Polymorphism, Single-Stranded Conformational ; Young Adult