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1.
Article in English | WPRIM | ID: wpr-289694

ABSTRACT

<p><b>OBJECTIVE</b>To explore the regulatory mechanism of Xiaoyin Recipe () on the T helper 1/T helper 2 (Th1/Th2) immune balance.</p><p><b>METHODS</b>Thirty-six experimental animals were divided into three groups, 12 rats in each group: blank control group (B group), negative control group (N group), and Xiaoyin Recipe treatment group (T group). The latter two groups received immunization of experimental autoimmune thyroiditis (EAT), and T group were treated with Xiaoyin Recipe for a month. Then, the expression of Th1-Th2-related genes in peripheral blood mononuclear cells (PBMCs) were screened with Oligo GEArray Rat Th1-Th2-Th3 Microarray. The expressions of tumor necrosis factor-α (TNF-α), interleukin-10 (IL-10), T-box expressed in T-cells (T-bet), and GATA-binding protein-3 (GATA-3) were detected by real-time polymerase chain reaction (RT-PCR).</p><p><b>RESULTS</b>Gene array screening showed that compared to N group, in T group after Xiaoyin Recipe treatment, 3 genes were upregulated in EAT rats, including interleukin-27 receptor alpha (IL-27rα), glomulin (Glmn), and GATA-3, while 38 genes were downregulated, such as CD28, IL-18, signal transducer, and activator of transcription 1 (STAT1), T-bet, TNF receptor superfamily member 4 (TNFRSF4), TNF ligand superfamily member 5 (TNFSF5), and TNF receptor superfamily member 5 (TNFRSF5). While RT-PCR showed that there was an increased level of TNF-α mRNA (P<0.01), an elevated ratio of T-bet/GATA-3, and a decreased level of IL-10 mRNA in PBMC of N and T group compared to B group (P <0.01); and after treatment with Xiaoyin Recipe, IL-10 mRNA level increased (P <0.01), while TNF-α mRNA level and T-bet/GATA-3 ratio decreased in T group compared to N group (P <0.01).</p><p><b>CONCLUSION</b>Xiaoyin Recipe for psoriasis could induce a Th1/Th2 balance drift toward Th2 in PBMC of EAT rats and thus improve the conditions.</p>


Subject(s)
Animals , Autoantibodies , Blood , Drugs, Chinese Herbal , Pharmacology , Therapeutic Uses , Female , GATA3 Transcription Factor , Genetics , Metabolism , Gene Expression Regulation , Interleukin-10 , Genetics , Metabolism , Psoriasis , Blood , Drug Therapy , Allergy and Immunology , Pathology , Rats , Rats, Wistar , T-Box Domain Proteins , Genetics , Metabolism , Th1-Th2 Balance , Th2 Cells , Allergy and Immunology , Thyroiditis, Autoimmune , Blood , Drug Therapy , Allergy and Immunology , Pathology , Tumor Necrosis Factor-alpha , Genetics , Metabolism
2.
Article in Chinese | WPRIM | ID: wpr-328576

ABSTRACT

The objective of this study was to establish an RNAi approach that can specifically target aqp1 gene sequence in vitro, and to assess the inhibitory effect of this siRNA on K562 cells. The siRNA targeting aqp1-mRNA was designed and transfected into K562 cells by using Lipofectamine(TM) 2000 reagent. Phase-contrast microscopy was used to analyze morphology changes of K562 cells. Cell viability was determined by MTT assay, and flow cytometry and DNA ladder analysis were carried out to identify siRNA-induced apoptosis. The expression levels of aqp1-mRNA at different transfection time were detected by RT-PCR. The results showed that the siRNA was successful by established. The transfected K562 cells displayed the significant apoptosis. The aqp1-siRNAs could obviously inhibit the activity of K562 cells. Cellular DNA fragmentation was observed in the siRNA group after transfection for 48 hours, the apoptosis rates at 24, 48 and 72 hours after transfection were 24.2%, 36.1% and 42.9% respectively. The aqp1-mRNA expression in the cells treated by aqp1-siRNA for 24, 48 and 72 hours were significantly reduced by 33%, 46% and 57% respectively. It is concluded that the aqp1-siRNA can efficiently and specifically inhibited the proliferation and inducing apoptosis of K562 cells. Gene aqp1 can be a potential target point for therapy of malignant tumor.


Subject(s)
Apoptosis , Aquaporin 1 , Genetics , Cell Proliferation , Cell Survival , Gene Targeting , Humans , K562 Cells , RNA Interference , RNA, Small Interfering , Genetics , Pharmacology
3.
Chinese Journal of Epidemiology ; (12): 795-799, 2010.
Article in Chinese | WPRIM | ID: wpr-341033

ABSTRACT

Objective To study the change of special antibodies titer IgG, IgM and nucleocaspid to SARS coronavirus (CoV) and observing the expression of stomach and enteric involvement on SARS-CoV infection by monoclonal antibody against N protein of SARS-CoV in the 7- year recovery period among family clustering cases of severe acute respiratory syndrome. Methods Special antibody titer to SARS-CoV of 14 patients from 5 different families and their 10 kinfolks continuously tested by IFA and antigen-capturing ELISA methods. Samples were taken in the 1st-7th year periods after SARS patients infected by SARS-CoV, being diluted and measured on it titers of three kinds of antibodies. Immunochemical staining with monoclonal antibody (mAb) against N protein of SARS-CoV was used to determine the stomach and enteric tissues among 5 SARS patients with their nucleocaspid antibody titer ascended obviously after 1st-7th year. Results When testing the IgG antibody titer of the 14 SARS patients by IFA method, the average titer was 1/71 (95%CI:1/58-1/85) in the 1st year, but began to descend in the following years, and the IgG antibody of the most SARS patients disappeared in the 7th year. Regarding the IgM titer, it disappeared in most of the SARS patients 1 year later. The average value of nucleocaspid antibody titer was 1/146 (95% CI:1/122-1/171) in the 1st year, and it descended as the IgG antibody titer did. In 5 cases, differences appeared.The nucleocaspid antibody titer was between 1/156 and 1/210 in 3 cases, and 2 cases were normal.Immunochemical staining with mAb against N protein of SARS-CoV was identified in the stomach and enteric tissues of 5 SARS patients with the nucleocaspid antibody titer increased significantly, 1st-7th year later. The five patients were detected by gastroscopy detection and cell immunohistochemistry test. 3 cases showed N protein antibody positive in the serum, and positive immunohistochemical expression in most of the cytoplasm in the gastric tissue mucous gland epithelial cells. 1 case also expressed in the intestinal tissue slurry columnar epithelium and interstitial cells. The other two cases showed negative on both serum N protein antibody and immunohistochemical expression. The biopsy results of the 5 patients were as follows: 1 case diagnosed as "signet-ring cell carcinoma of the stomach and rectum multiple transfer", 1 case of gastric polyp, 1 case of superficial antral gastritis and 2 cases were normal. Conclusion By testing the special IgG, IgM, nucleocaspid antibody to SARS-CoV of the 14 family clustering cases , we found that they all decreased in the 7th year, and most of them disappeared. The nucleocaspid antibody titer was related to pathogenetic condition. SARS-CoV was proved to be still present in stomach and enteric tissues of SARS patients with the nucleocaspid antibody titer increased significantly after the 7th year.

4.
Article in Chinese | WPRIM | ID: wpr-336018

ABSTRACT

<p><b>OBJECTIVE</b>To explore the feasibility of using gene chip method to identify pathogens in blood cultures.</p><p><b>METHODS</b>Clinical blood samples were obtained and cultured using an automated blood culture system. A gene chip diagnostic kit was used to detect the pathogenic bacteria in these blood cultures following the procedures of target gene extraction and amplification, hybridization and result analysis. The conventional method was also used to isolate and identify the bacteria from the clinical blood cultures, and the results of the two methods were compared.</p><p><b>RESULTS</b>In the 86 clinical blood samples, 74 were positive and 12 negative according to the conventional method, while 48 were positive and 38 negative as found by the gene chip method, showing significant differences in the results (P<0.05). The two methods only had a concordance rate of 69.77%.</p><p><b>CONCLUSION</b>The gene chip diagnostic kit has low concordance rate with the conventional method for detecting pathogens in clinical blood cultures and awaits further improvement.</p>


Subject(s)
Bacteria , Genetics , Bacterial Typing Techniques , Methods , Blood , Microbiology , DNA, Bacterial , Genetics , Humans , Oligonucleotide Array Sequence Analysis , Methods , RNA, Ribosomal, 16S , Genetics
5.
Chinese Journal of Hepatology ; (12): 359-362, 2009.
Article in Chinese | WPRIM | ID: wpr-310086

ABSTRACT

<p><b>OBJECTIVE</b>To test whether nuclear factor kappa B plays an important role in the apoptosis-inhibitory effect of hepatitis B virus (HBV) P22(e) protein.</p><p><b>METHODS</b>HepG2 cells were transfected with recombination plasmid pEGFP-HBVP22(e). The Act-D and TNF alpha were used to induce apoptosis. NF-kappa B inhibitor ALLN were used to inhibit the signaling pathway. The activation of NF-kappa B was EMSA, and the nulear translocation of NF-kappa B was determined by immuno-staining.</p><p><b>RESULTS</b>Laser scanning confocal microscopy and EMSA indicated that HBV P22(e) protein enhanced the nuclear translocation of NF-kappa B after apoptosis induction. ALLN treatment inhibited the nuclear translocation of NF-kappa B, and blocked the apoptosis-inhibiting effect of HBV P22(e) protein.</p><p><b>CONCLUSION</b>This study indicates that HBV P22(e) protein inhibits apoptosis of hepatocyte via the NF-kappa B signaling pathway.</p>


Subject(s)
Apoptosis , Carcinoma, Hepatocellular , Metabolism , Hep G2 Cells , Hepatitis B Core Antigens , Metabolism , Hepatitis B virus , Genetics , Humans , Leupeptins , Pharmacology , Liver Neoplasms , Metabolism , NF-kappa B , Metabolism , Plasmids , Signal Transduction , Transfection , Viral Core Proteins , Metabolism
6.
Chinese Journal of Hepatology ; (12): 816-820, 2007.
Article in Chinese | WPRIM | ID: wpr-354619

ABSTRACT

<p><b>OBJECTIVE</b>To prepare oligo microarrays for hepatitis virus detection and genotyping.</p><p><b>METHODS</b>By analyzing the DNA or cDNA of HBV, HDV and 4 different genotypes of HCV with the BLAST program, a group of specific sequences for the candidate probes was specified. Array Designer 3.0 software was applied to analyze the candidates to select probes with high specificity, identical length and similar melting temperature (Tm). Altogether 16, 8 and 68 oligonucleotide probes were designed for diagnosis of HBV, HDV, and genotyping HCV. Following the synthesizing and purification, oligo probes were deposited on oligonucleotide chips as microarrays for hepatitis virus detection and genotyping. The samples were labeled by RD-PCR method. Hybridization results were analyzed to cross out those probes with low specificity and sensitivity, and those with signal to noise ratios (SNR) less than 4.0.</p><p><b>RESULTS</b>Two types of gene chips were successfully developed: microarrays for HBV and HDV simultaneous detection and for HCV genotyping.</p><p><b>CONCLUSION</b>Using oligo probes to construct gene chips for clinical diagnosis of hepatitis virus is a simple and effective method. It may be widely used in detecting hepatitis viruses and their genotyping in clinical settings.</p>


Subject(s)
Base Sequence , DNA Fingerprinting , DNA, Viral , Genetics , Genotype , Hepatitis B virus , Classification , Genetics , Molecular Sequence Data , Oligonucleotide Array Sequence Analysis , Oligonucleotide Probes , Sensitivity and Specificity
7.
Chinese Journal of Biotechnology ; (12): 392-396, 2003.
Article in Chinese | WPRIM | ID: wpr-259180

ABSTRACT

Genes encoding nucleocaspid (N) and membrane (M) protein of SARS coronavirus were obtained by RT-PCR and were cloned into expression vector pET22b and pBV222. DNA sequencing showed that the genes cloned from a patient in Beijing were identical to the gene sequences from reported Toronto strain. The genes were over-expressed in E. coli either as inclusion body or as soluble form. The recombinant proteins were purified by ion-exchange, or ion-exchange followed by metal chelate affinity chromatography. The recombinant N protein was demonstrated highly antigenic and could be employed as antigen to detect SARS antibodies in ELISA system for SARS diagnosis.


Subject(s)
Chromatography, Affinity , Chromatography, Ion Exchange , Enzyme-Linked Immunosorbent Assay , Escherichia coli , Genetics , Metabolism , Nucleocapsid Proteins , Genetics , Metabolism , Reverse Transcriptase Polymerase Chain Reaction , SARS Virus , Genetics , Metabolism , Viral Structural Proteins , Genetics , Metabolism
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